title: Patterns and Progress of Peninsular Malaysia's Amphibian Research in the 21st Century (2000-2020)

abstract: In this study, we review the status, patterns, and progress of Peninsular Malaysias amphibian research in the 21st century with the main goal of identifying areas for improvement that can help focus and prioritize future research initiatives. Between 2000-2020 we found 130 publications that can be broadly categorized into four groups: 1) checklists and biodiversity; 2) new species and taxonomy; 3) ecology and natural history; and 4) evolution and phylogenetics. An average of 6.5 papers was published per year and although the number of papers fluctuated, there was a significant upward trend in the number of papers published. Almost half (49%) of all papers published comprised checklists and biodiversity-related papers. This was followed by new species and taxonomy (25%, 33 papers), evolution and phylogenetics (14%, 18 papers), and ecology and natural history (12%, 16 papers). Amphibian research was conducted most frequently in the states of Kedah, Pahang, and Perak, and most infrequently in the states of Malacca, Negeri Sembilan, Selangor/Kuala Lumpur, Perlis, and Kelantan. Despite being a megadiverse country and a biodiversity hotspot, not a single conservation-centric paper has ever been published on Peninsular Malaysian amphibians, highlighting the urgent need for future research to focus on conservation.

category: scientific communication and education


title: Detection of Locomotion Deficit in a Post-Traumatic Syringomyelia Rat Model Using Automated Gait Analysis Technique

abstract: Syringomyelia (SM) is a spinal cord disorder in which a cyst (syrinx) filled with fluid forms in the spinal cord post-injury/disease, in patients syrinx symptoms include loss of pain and temperature sensation or locomotion deficit. Currently, there are no small animal models and connected tools to help study the functional impacts of SM. The objective of this study was to determine the detectability of subtle locomotion deficits due to syrinx formation/expansion in post-traumatic syringomyelia (PTSM) rat model using the recently reported method of Gait Analysis Instrumentation, and Technology Optimized for Rodents (GAITOR) with Automated Gait Analysis Through Hues and Areas (AGATHA) technique. First videos of the rats were collected while walking in an arena (using GAITOR) followed by extracting meaningful locomotion information from collected videos (using AGATHA protocol. PTSM injured rats demonstrated detectable locomotion deficits in terms of duty factor imbalance, paw placement accuracy, step contact width, stride length, and phase dispersion parameters compared to uninjured rats due to SM. We concluded that this technique could detect mild and subtle locomotion deficits associated with PTSM injury, which also in future work could be used further to monitor locomotion responses after different treatment strategies for SM.

category: bioengineering


title: Circularly permuted LOV domain as an engineering module for optogenetic tools

abstract: We re-engineered a commonly-used light-sensing protein, LOV domain, using a circular permutation strategy to allow photoswitchable control of the C-terminus of a peptide. We demonstrate that the use of circularly permuted LOV domain on its own or together with the original LOV could expand the engineering capabilities of optogenetic tools.

category: synthetic biology


title: Cortical Representations of Concrete and Abstract Concepts in Language Combine Visual and Linguistic Representations

abstract: The human semantic system stores knowledge acquired through both perception and language. To study how semantic representations in cortex integrate perceptual and linguistic information, we created semantic word embedding spaces that combine models of visual and linguistic processing. We then used these visually-grounded semantic spaces to fit voxelwise encoding models to fMRI data collected while subjects listened to hours of narrative stories. We found that cortical regions near the visual system represent concepts by combining visual and linguistic information, while regions near the language system represent concepts using mostly linguistic information. Assessing individual representations near visual cortex, we found that more concrete concepts contain more visual information, while even abstract concepts contain some amount of visual information from associated concrete concepts. Finally we found that these visual grounding effects are localized near visual cortex, suggesting that semantic representations specifically reflect the modality of adjacent perceptual systems. Our results provide a computational account of how visual and linguistic information are combined to represent concrete and abstract concepts across cortex.

category: neuroscience


title: Impaired and Intact Aspects of Attentional Competition and Prioritization during Visual Working Memory Encoding in Schizophrenia

abstract:
ObjectivePeople with schizophrenia (PSZ) are impaired in the attentional prioritization of non-salient but relevant stimuli over salient but irrelevant distractors during visual working memory (VWM) encoding. Conversely, the guidance of top-down attention by external predictive cues is intact. Yet, it is unknown whether this preserved ability can help PSZ overcome impaired attentional prioritization in the presence of salient distractors.

MethodsWe employed a visuospatial change-detection task using four Gabor Patches with differing orientations in 69 PSZ and 74 healthy controls (HCS). Two patches flickered to reflect saliency and either a predictive or a non-predictive cue was displayed resulting in four conditions.

ResultsAcross all conditions, PSZ stored significantly less information in VWM than HCS (all p < 0.001). With a non-predictive cue, PSZ stored significantly more salient than non-salient information (t140 = 5.66, p < 0.001, dt = 0.5). With a predictive cue, PSZ stored significantly more non-salient information (t140 = 5.70, p < 0.001, dt = 0.5).

ConclusionOur findings support a bottom-up bias in schizophrenia with performance significantly better for visually salient information in the absence of a predictive cue. These results indicate that bottom-up attentional prioritization is disrupted in schizophrenia, but the top-down utilization of cues is intact. We conclude that additional top-down information significantly improves performance in PSZ when non-salient visual information needs to be encoded in working memory.


title: Corticotropin-Releasing Factor Induces Functional And Structural Synaptic Remodelling In Acute Stress

abstract: Biological responses to internal and external stress factors involve highly conserved mechanisms, using a tightly coordinated interplay of many factors. Corticotropin-releasing factor (CRF) plays a central role in organizing these lifesaving physiological responses to stress. We show that CRF rapidly and reversibly changes Schaffer Collateral input into hippocampal CA1 pyramidal cells (PC), by modulating both functional and structural aspects of these synapses. Host exposure to acute stress, in vivo CRF injection, and ex vivo CRF application all result in fast de novo formation and remodeling of existing dendritic spines. Functionally, CRF leads to a rapid increase in synaptic strength of Schaffer collateral input into CA1 neurons, e.g. increase in spontaneous neurotransmitter release, paired-pulse facilitation and repetitive excitability and improves long-term synaptic plasticity: LTP and LTD. In line with the changes in synaptic function, CRF increases the number of presynaptic vesicles, induces redistribution of vesicles towards the active zone increases active zone size, and improves the alignment of the pre- and post-synaptic compartments. Together, CRF rapidly enhances synaptic communication in the hippocampus, potentially playing a crucial role in the enhanced memory consolidation in acute stress.


title: Perineuronal nets in HVC and plasticity in male canary song

abstract: Songbirds learn their vocalizations during developmental sensitive periods of song memorization and sensorimotor learning. Some seasonal songbirds, called open-ended learners, recapitulate transitions from sensorimotor learning and song crystallization on a seasonal basis during adulthood. In adult male canaries, sensorimotor learning occurs each year in autumn and leads to modifications of the syllable repertoire during successive breeding seasons. We previously showed that perineuronal nets (PNN) expression in song control nuclei decreases during this sensorimotor learning period. Here we explored the causal link between PNN expression in adult canaries and song modification by enzymatically degrading PNN in HVC, a key song control system nucleus. Three independent experiments identified limited effects of the PNN degradation in HVC on the song structure of male canaries. They clearly establish that presence of PNN in HVC is not required to maintain general features of crystallized song. Some suggestion was collected that PNN are implicated in the stability of song repertoires but this evidence is too preliminary to draw firm conclusions and additional investigations should consider producing PNN degradations at specified time points of the seasonal cycle. It also remains possible that once song has been crystallized at the beginning of the first breeding season, PNN no longer play a key role in determining song structure which could be tested by treatments with ChABC at key step in ontogeny. It would in this context be important to develop multiple stereotaxic procedures allowing the simultaneous bilateral degradation of PNN in several song control nuclei for extended periods.


title: A hybridoma-derived monoclonal antibody with high homology to the aberrant myeloma light chain

abstract: The identification of antibody variable regions in the heavy (VH) and light (VL) chains from hybridomas is necessary for the production of recombinant, sequence-defined monoclonal antibodies (mAbs) and antibody derivatives. This process has received renewed attention in light of recent reports of hybridomas having unintended specificities due to the production of non-antigen specific heavy and/or light chains for the intended antigen. Here we report a surprising finding and potential pitfall in variable domain sequencing of an anti-human CD63 hybridoma. We amplified multiple VL genes from the hybridoma cDNA, including the well-known aberrant Sp2/0 myeloma VK and a unique, full-length VL. After finding that the unique VL failed to yield a functional antibody, we discovered an additional full-length sequence with surprising similarity (~95% sequence identify) to the non-translated myeloma kappa chain but with a correction of its key frameshift mutation. Expression of the recombinant mAb confirmed that this highly homologous sequence is the antigen-specific light chain. Our results highlight the complexity of PCR-based cloning of antibody genes and strategies useful for identification of correct sequences.

category: molecular biology


title: Concurrent invasions by European starlings (Sturnus vulgaris) suggest selection on shared genomic regions even after genetic bottlenecks

abstract: A species success during the invasion of new areas hinges on an interplay between demographic processes and the outcome of localized selection. Invasive European Starlings (Sturnus vulgaris) established populations in Australia and North America in the 19th century. Here, we compare whole-genome sequences among native and independently introduced European Starling populations from three continents to determine how demographic processes interact with rapid adaptive evolution to generate similar genetic patterns in these recent and replicated invasions. Our results confirm that a post-bottleneck expansion may in fact support local adaptation. We find that specific genomic regions have differentiated even on this short evolutionary timescale, and suggest that selection best explains differentiation in at least two of these regions. This infamous and highly mobile invader adapted to novel selection (e.g., extrinsic factors), perhaps in part due to the demographic boom intrinsic to many invasions.

category: evolutionary biology


title: Natural variation in the consequences of gene overexpression and its implications for evolutionary trajectories

abstract: Copy number variation (CNV) through gene or chromosome amplification provides a route for rapid phenotypic variation and supports long-term evolution of gene functions. Although the evolutionary importance of CNV is known, little is understood about how genetic background influences CNV tolerance. Here, we measured fitness costs of over 4,000 over-expressed genes in 15 Saccharomyces cerevisiae strains representing different lineages, to explore natural variation in tolerating gene overexpression (OE). Strain-specific effects dominated the fitness costs of gene OE. We report global differences in the consequences of gene OE, independent of the amplified gene, as well as gene-specific effects that were dependent on the genetic background. Natural variation in the response to gene OE could be explained by several models, including strain-specific physiological differences, resource limitations, and regulatory sensitivities. This work provides new insight on how genetic background influences tolerance to gene amplification and the evolutionary trajectories accessible to different backgrounds.

category: genomics


title: Genetic structure correlates with ethnolinguistic diversity in eastern and southern Africa

abstract: African populations are the most diverse in the world yet are sorely underrepresented in medical genetics research. Here, we examine the structure of African populations using genetic and comprehensive multigenerational ethnolinguistic data from the Neuropsychiatric Genetics of African Populations-Psychosis study (NeuroGAP-Psychosis) consisting of 900 individuals from Ethiopia, Kenya, South Africa, and Uganda. We find that self-reported language classifications meaningfully tag underlying genetic variation that would be missed with consideration of geography alone, highlighting the importance of culture in shaping genetic diversity. Leveraging our uniquely rich multi-generational ethnolinguistic metadata, we track language transmission through the pedigree, observing the disappearance of several languages in our cohort as well as notable shifts in frequency over three generations. We find suggestive evidence for the rate of language transmission in matrilineal groups having been higher than that for patrilineal ones. We highlight both the diversity of variation within the African continent, as well as how within-Africa variation can be informative for broader variant interpretation; many variants appearing rare elsewhere are common in parts of Africa. The work presented here improves the understanding of the spectrum of genetic variation in African populations and highlights the enormous and complex genetic and ethnolinguistic diversity within Africa.


title: Linguistic laws are not the law in chimpanzee sexual solicitation gestures.

abstract: Two language laws have been identified as manifestations of universal principles of animal behaviour, both acting on the organisation of numerous vocal and behavioural communicative systems. Zipfs law of brevity describes a negative relationship between behavioural length and frequency of behaviour. Menzeraths law defines a negative correlation between the number of behaviours in a sequence and average length of the behaviour composing it. Both laws have been linked with the information-theoretic principle of compression, which tends to minimise code length. We investigate the presence of these two laws in the repertoire of chimpanzee sexual solicitation gestures. We find that chimpanzee solicitation gestures do not follow either Zipfs law of brevity or Menzeraths law consistently. For the second time in ape gestural communication, evidence supporting Zipfs law of brevity was absent, and, here, the presence of Menzeraths law appears individually driven. Ape gesture does not appear to manifest a principle of compression or pressure for efficiency that has been previously proposed to be universal. Importantly, the same signals were shown to adhere to these laws when used in a different behavioural context; highlighting that signallers consider signalling efficiency broadly, and diverse factors play important roles in shaping investment in signal production.

category: animal behavior and cognition


title: Mitochondria serve as axonal shuttle for Cox7c mRNA through mechanism that involves its mitochondrial targeting signal

abstract: Localized protein synthesis plays a key role in spatiotemporal regulation of the cellular proteome. Neurons, which extend axons over long distances, heavily depend on this process. However, the mechanisms by which axonal mRNAs are transported to protein target sites are not fully understood. Here, we describe a novel role for mitochondria in shuttling a nuclear encoded mRNA along axons. Fractionation analysis and smFISH revealed that the mRNA encoding Cox7c protein is preferentially associated with mitochondria from a neuronal cell line and from primary motor neuron axons. Live cell imaging of MS2-tagged Cox7c or Cryab control mRNA in primary motor neurons further confirmed the preferential colocalization of Cox7c mRNA with mitochondria. More importantly, Cox7c demonstrated substantial cotransport with mitochondria along axons. Intriguingly, the coding region, rather than the 3UTR, was found to be the key domain for the cotransport. Furthermore, we show that puromycin treatment as well as hindering the synthesis of the mitochondrial targeting signal (MTS) reduced the colocalization. Overall, our results reveal a novel mRNA transport mode which exploits mitochondria as a shuttle and translation of the MTS as a recognition feature. Thus, mitochondria may play a role in spatial regulation of the axonal transcriptome and self-sustain their own proteome at distant neuronal sites.

category: cell biology


title: The unfolded protein response of the endoplasmic reticulum supports mitochondrial biogenesis by buffering non-imported proteins

abstract: Almost all mitochondrial proteins are synthesized in the cytosol and subsequently targeted to mitochondria. The accumulation of non-imported precursor proteins occurring upon mitochondrial dysfunction can challenge cellular protein homeostasis. Here we show that blocking protein translocation into mitochondria results in the accumulation of mitochondrial membrane proteins at the endoplasmic reticulum, thereby triggering the unfolded protein response (UPRER). Moreover, we find that mitochondrial membrane proteins are also routed to the ER under physiological conditions. The levels of ER-resident mitochondrial precursors is enhanced by import defects as well as metabolic stimuli that increase the expression of mitochondrial proteins. Under such conditions, the UPRER is crucial to maintain protein homeostasis and cellular fitness. We propose the ER serves as a physiological buffer zone for those mitochondrial precursors that cant be immediately imported into mitochondria while engaging the UPRER to adjust the ER proteostasis capacity to the extent of precursor accumulation.


title: A new type of ERGIC-ERES membrane contact mediated by TMED9 and SEC12 is required for autophagosome biogenesis

abstract: Under stress, the endomembrane system undergoes reorganization to support autophagosome biogenesis, which is a central step in autophagy. How the endomembrane system remodels has been poorly understood. Here we identify a new type of membrane contact formed between the ER-Golgi intermediate compartment (ERGIC) and the ER-exit site (ERES) in the ER-Golgi system, which is essential for promoting autophagosome biogenesis induced by different stress stimuli. The ERGIC-ERES contact is established by the interaction between TMED9 and SEC12 which generates a short distance opposition (as close as 2-5 nm) between the two compartments. The tight membrane contact allows the ERES-located SEC12 to transactivate COPII assembly on the ERGIC. In addition, a portion of SEC12 also relocates to the ERGIC. Through both mechanisms, the ERGIC-ERES contact promotes formation of the ERGIC-derived COPII vesicle, a membrane precursor of the autophagosome. The ERGIC-ERES contact is physically and functionally different from the TFG-mediated ERGIC-ERES adjunction involved in secretory protein transport, and therefore defines a unique endomembrane structure generated upon stress conditions for autophagic membrane formation.


title: CD9 tetraspanins convey robustness to CXCR4b signalling during collective cell migration

abstract: Collective cell migration is essential for embryonic development and homeostatic processes. During zebrafish development, the posterior lateral line primordium (pLLP) navigates along the embryo flank by collective cell migration. The chemokine receptors, Cxcr4b and Cxcr7b, as well as their cognate ligand, Cxcl12a, are essential for this process. We corroborate that knockdown of the zebrafish cd9 tetraspanin orthologue, cd9b, results in mild pLL abnormalities. Through generation of CRISPR and TALEN mutants, we show that cd9a and cd9b function partially redundantly in pLLP migration, which is delayed in the cd9b single and cd9a; cd9b double mutants. This delay led to a transient reduction in neuromast numbers. Loss of both Cd9a and Cd9b sensitized embryos to reduced Cxcr4b and Cxcl12a levels. Together these results provide evidence that Cd9 modulates collective cell migration of the pLLP during zebrafish development. One interpretation of these observations is that Cd9 contributes to more effective chemokine signalling.

category: developmental biology


title: Tetraspanin Cd9b and Cxcl12a/Cxcr4b have a synergistic effect on the control of collective cell migration


title: Hif1α is required for Wnt regulated gene expression during Xenopus tropicalis tail regeneration

abstract: Regeneration of complex tissues is initiated by an injury-induced stress response, eventually leading to activation of developmental signaling pathways such as Wnt signaling. How early injury cues are interpreted and coupled to activation of these developmental signals and their targets is not well understood. Here, we show that Hif1, a stress induced transcription factor, is required for tail regeneration in Xenopus tropicalis. We find that Hif1 is required for regeneration of differentiated axial tissues, including axons and muscle. Using RNA-sequencing, we find that Hif1 and Wnt converge on a broad set of genes required for posterior specification and differentiation, including the posterior hox genes. We further show that Hif1 is required for transcription via a Wnt-responsive element, a function that is conserved in both regeneration and early neural patterning. Our findings indicate a regulatory role for Hif1 in Wnt mediated gene expression across multiple tissue contexts.


title: Semaphorin3f as an intrinsic regulator of chamber-specific heart development

abstract: During development a pool of precursors form a heart with atrial and ventricular chambers that exhibit distinct transcriptional and electrophysiological properties. Normal development of these chambers is essential for full term survival of the fetus, and deviations result in congenital heart defects. The large number of genes that may cause congenital heart defects when mutated, and the genetic variability and penetrance of the ensuing phenotypes, reveals a need to understand the molecular mechanisms that allow for the formation of chamber-specific cardiomyocyte differentiation. We find that in the developing zebrafish heart, mRNA for a secreted Semaphorin (Sema), Sema3fb, is expressed by all cardiomyocytes, whereas mRNA for its receptor Plexina3 (Plxna3) is expressed by ventricular cardiomyocytes. In sema3fb CRISPR zebrafish mutants, ventricular chamber development is impaired; the ventricles of mutants are smaller in size than their wild type siblings, apparently because of differences in cell size and not cell numbers, with ventricular cardiomyocytes failing to undergo normal developmental hypertrophy. Analysis of chamber differentiation indicates defects in chamber specific gene expression at the border between the ventricular and atrial chambers, with spillage of ventricular chamber genes into the atrium, and vice versa, and a failure to restrict bmp4a mRNA to the atrioventricular canal. The disrupted atrioventricular border region in mutants is accompanied by hypoplastic heart chambers and impaired cardiac function. These data suggest a model whereby cardiomyocytes secrete a Sema cue that, through spatially restricted expression of the receptor, signals in a ventricular chamber-specific manner to establish a distinct border between atrial and ventricular chambers that is important for functional development of the heart.


title: Epi-mutations for spermatogenic defects by maternal exposure to Di (2-ethylhexyl) phthalate

abstract: Exposure to environmental factors during fetal development may lead to epigenomic modifications in fetal germ cells, altering gene expression and promoting diseases in successive generations. In mouse, maternal exposure to Di (2-ethylhexyl) phthalate (DEHP) is known to induce defects in spermatogenesis in successive generations, but the mechanism(s) of impaired spermatogenesis are unclear. Here, we showed that maternal DEHP exposure results in DNA hypermethylation of promoters of spermatogenesis-related genes in fetal testicular germ cells in F1 mice, and hypermethylation of Hist1h2ba, Sycp1 and Taf7l, which are crucial for spermatogenesis, persisted from fetal testicular cells to adult spermatogonia, resulting in the downregulation of expression of these genes. Forced methylation of these gene promoters silenced expression of these loci in a reporter assay. Expression and methylation of those genes tended to be downregulated and increased, respectively in F2 spermatogonia following maternal DEHP exposure. These results suggested that DEHP-induced hypermethylation of Hist1h2ba, Sycp1 and Taf7l in fetal germ cells results in downregulation of these genes in spermatogonia and subsequent defects in spermatogenesis, at least in the F1 generation.


title: A Modular Platform for Streamlining Automated Cryo-FIB Workflows

abstract: Lamella micromachining by focused ion beam milling at cryogenic temperature (cryo-FIB) has matured into a preparation method widely used for cellular cryo-electron tomography. Due to the limited ablation rates of low Ga+ ion beam currents required to maintain the structural integrity of vitreous specimens, current preparation protocols are time-consuming and labor intensive. The improved stability of new generation cryo-FIB instruments now enables automated operations. Here, we present an open-source software tool, SerialFIB, for creating automated and customizable cryo-FIB preparation protocols. The software encompasses a graphical user interface for easy execution of routine lamellae preparations, a scripting module compatible with available python packages, and interfaces with 3-dimensional correlative light and electron microscopy (CLEM) tools. The software enables the streamlining of advanced cryo-FIB protocols such as multi-modal imaging, CLEM guided preparation and in situ lamella lift-out procedures. Our software therefore provides a foundation for further development of advanced cryogenic imaging and sample preparation protocols.

category: biophysics


title: The folding space of protein β2-microglobulin is modulated by a single disulfide bond

abstract: Protein beta-2-microglobulin ({beta}2m) is classically considered the causative agent of dialysis related amyloidosis (DRA), a conformational disorder that affects patients undergoing long-term hemodialysis. Together with the wild type form, the {Delta}N6 structural variant, and the D76N mutant, have been extensively used as model systems of {beta}2m aggregation. In all of them, the native structure is stabilized by a disulfide bridge between the sulphur atoms of the cysteine residues 25 (at B strand) and 80 (at F strand), which has been considered fundamental in {beta}2m fibrillogenesis. Here, we use extensive Discrete Molecular Dynamics simulations of a full atomistic structurebased model to explore the role of this disulfide bridge as a modulator of the folding space of {beta}2m. In particular, by considering different models for the disulfide bridge, we explore the thermodynamics of the folding transition, and the formation of intermediate states that may have the potential to trigger the aggregation cascade. Our results show that the dissulfide bridge affects folding transition and folding thermodynamics of the considered model systems, although to different extents. In particular, when the interaction between the sulphur atoms is stabilized relative to the other intramolecular interactions, or even locked (i.e. permanently established), the WT form populates an intermediate state featuring a well preserved core, and two unstructured termini, which was previously detected only for the D76N mutant. The formation of this intermediate state may have important implications in our understanding of {beta}2m fibrillogenesis.


title: A specific set of heterogeneous native interactions yields efficient knotting in protein folding

abstract: Native interactions are crucial for folding, and non-native interactions appear to be critical for efficiently knotting proteins. Therefore, it is important to understand both their roles in the folding of knotted proteins. It has been proposed that non-native interactions drive the correct order of contact formation, which is essential to avoid backtracking and efficiently self-tie. In this study we ask if non-native interactions are strictly necessary to tangle a protein, or if the correct order of contact formation can be assured by a specific set of native, but otherwise heterogeneous, interactions. In order to address this problem we conducted extensive Monte Carlo simulations of lattice models of proteinlike sequences designed to fold into a pre-selected knotted conformation embedding a trefoil knot. We were able to identify a specific set of heterogeneous native interactions that drives efficient knotting, and is able to fold the protein when combined with the remaining native interactions modeled as homogeneous. This specific set of heterogeneous native interactions is strictly enough to efficiently self-tie. A distinctive feature of these native interactions is that they do not backtrack, because their energies ensure the correct order of contact formation. Furthermore, they stabilize a knotted intermediate state, which is enroute to the native structure. Our results thus show that - at least in the context of the adopted model - non-native interactions are not necessary to knot a protein. However, when they are taken into account into protein energetics it is possible to find specific, non-local non-native interactions that operate as a scaffold that assists the knotting step.


title: Anomalous propulsion regime in axonemal-propelled cargoes

abstract: Bio-hybrid micro-swimmers, composed of biological entities integrated with synthetic constructs, actively transport cargo by converting chemical energy into mechanical work in a fluid at low Reynolds number, where viscous drag dominates over inertia. Here, using isolated and demembranated flagella from green algae Chlamydomonas reinhardtii (C. reinhardtii), we build efficient axonemally-driven micro-swimmers that consume ATP to propel micron-sized beads. Depending on the calcium concentration, we observed two main classes of motion: Whereas beads move along curved trajectories at calcium concentrations below 0.03 mM, they are propelled along straight paths when the calcium concentration increases. In this regime, they reached velocities of approximately 20 m/sec, comparable to human sperm velocity in vivo. We relate this transition to the properties of beating axonemes, in particular the reduced static curvature with increasing calcium concentration. To quantify the motion, we used mode decomposition of the flagellar waveform, and we studied both analytically and numerically the propulsion of the bead as a function of the axonemal waveform and bead-axoneme attachment geometry. While our analysis semi-quantitatively describes the experimental results, it also reveals the existence of a counter-intuitive propulsion regime where the speed of the axonemally-driven bead increases with the size of the bead. Moreover, we demonstrated that asymmetric, sideways attachment of the axoneme to the bead can also contribute to the rotational velocity of the micro-swimmer. The uncovered mechanism has potential applications in the fabrication of synthetic micro-swimmers, and in particular, bio-actuated medical micro-robots for targeted drug delivery.


title: Bio-hybrid micro-swimmers propelled by flagella isolated from C. reinhardtii


title: Kel1 is a phosphorylation-regulated noise suppressor of the pheromone signaling pathway

abstract: Mechanisms have evolved that allow cells to detect signals and generate an appropriate response. The accuracy of these responses relies on the ability of cells to discriminate between signal and noise. How cells filter noise in signaling pathways is not well understood. Here, we analyze noise suppression in the yeast pheromone signaling pathway and show that the poorly characterized protein Kel1 serves as a major noise suppressor and prevents cell death. At the molecular level, Kel1 prevents spontaneous activation of the pheromone response by inhibiting membrane recruitment of Ste5 and Far1. Only a hypophosphorylated form of Kel1 suppresses signaling, reduces noise and prevents pheromone-associated cell death, and our data indicate that the MAPK Fus3 contributes to Kel1 phosphorylation. Taken together, Kel1 serves as a phospho-regulated suppressor of the pheromone pathway to reduce noise, inhibit spontaneous activation of the pathway, regulate mating efficiency, and to prevent pheromone-associated cell death.


title: Near-infrared imaging in fission yeast by genetically encoded biosynthesis of phycocyanobilin

abstract: Near-infrared fluorescent protein (iRFP) is a bright and stable fluorescent protein with excitation and emission maxima at 690 nm and 713 nm, respectively. Unlike the other conventional fluorescent proteins such as GFP, iRFP requires biliverdin (BV) as a chromophore because iRFP originates from bacteriophytochrome. Here, we report that phycocyanobilin (PCB) functions as a brighter chromophore for iRFP than BV, and biosynthesis of PCB allows live-cell imaging with iRFP in the fission yeast Schizosaccharomyces pombe. We initially found that fission yeast cells did not produce BV, and therefore did not show any iRFP fluorescence. The brightness of iRFP attached to PCB was higher than that of iRFP attached to BV in vitro and in fission yeast. We introduced SynPCB, a previously reported PCB biosynthesis system, into fission yeast, resulting in the brightest iRFP fluorescence. To make iRFP readily available in fission yeast, we developed an endogenous gene tagging system with iRFP and all-in-one integration plasmids, which contain genes required for the SynPCB system and the iRFP-fused marker proteins. These tools not only enable the easy use of iRFP in fission yeast and the multiplexed live-cell imaging in fission yeast with a broader color palette, but also open the door to new opportunities for near-infrared fluorescence imaging in a wider range of living organisms.


title: Progressive Increases in Mesenchymal Cell Diversity Modulate Lung Development and are Attenuated by Hyperoxia

abstract: Early in life, the lung mesenchyme is extremely dynamic. We profiled the transcriptomes and locations of mesenchymal cells (MC) in the perinatal murine lung and observed cellular progressions of fibroblasts, myofibroblasts, airway smooth muscle (ASM), and mural cells, poised to modulate the extracellular matrix (ECM) with both subtype- and temporal-specific patterns. Within one day after birth, embryonic fibroblast precursors branched into two subtypes, alveolar and adventitial fibroblasts. ASM and myofibroblasts derived from a common precursor population that specifically expressed Crh, a hormone central to glucocorticoid release. Vascular smooth muscle cells and pericytes differentiated gradually. Fibroblast, myofibroblast, and pericyte proliferation peaked at 7 days after birth. Paracrine signaling among cell types decreased after birth and was hierarchical, with pericytes at the interface between endothelial cells and other MC. Postnatal hyperoxia delayed the developmental progression in cell type abundance and gene expression, mirroring the arrested development that characterizes the neonatal lung disease, bronchopulmonary dysplasia. Hyperoxia decreased pericyte and myofibroblast abundances and proliferation, dysregulated signaling, altered ECM modulation and, in males specifically, gave rise to a novel population of contractile fibroblasts. This study identifies the mesenchymal populations orchestrating lung development at the critical transition to air-breathing life and their distinct reactions to hyperoxia.


title: Buffered EGFR signaling regulated by spitz to argos expression ratio is critical for patterning the Drosophila eye

abstract: The Epidermal Growth Factor Receptor (EGFR) signaling pathway plays a critical role in regulating tissue patterning. Drosophila EGFR (DER) signaling achieves specificity through multiple ligands and feedback loops to finetune signaling spatiotemporally. The principal Drosophila EGF, cleaved Spitz, and the negative feedback molecule, Argos are diffusible and can act both in a cell autonomous and non-autonomous manner. The relative expression dose of Spitz and Argos early in development has been shown to be critical in patterning the Drosophila eye, but the exact identity of the cells expressing these genes in the larval eyedisc has been elusive. Using single molecule RNA Fluorescence in situ Hybridization (smFISH), we reveal an intriguing differential expression of spitz and argos in the Drosophila third instar eye imaginal disc indicative of directional DER signaling. By genetically tuning DER signaling, we show that rather than absolute levels of expression, the ratio of expression to be critical for determining the adult eye phenotype. Proper ommatidial patterning is robust to thresholds around a tightly maintained wildtype ratio, and breaks down beyond. This provides a powerful instance of developmental buffering.


title: The FusX TALE Base Editor (FusXTBE) for rapid mitochondrial DNA programming of human cells in vitro and zebrafish disease models in vivo

abstract: Functional analyses of mitochondria have been hampered by few effective approaches to manipulate mtDNA and a lack of existing animal models. Recently a TALE-derived base editor was shown to induce C-to-T (or G-to-A) sequence changes in mtDNA. We report here the FusX TALE Base Editor (FusXTBE) to facilitate broad-based access to TALE mitochondrial base editing technology. TALE Writer is a de novo in silico design tool to map potential mtDNA base editing sites. FusXTBE was demonstrated to function with comparable activity to the initial base editor in human cells in vitro. Zebrafish embryos were used as a pioneering in vivo test system, with FusXTBE inducing 90+% editing efficiency in mtDNA loci, the first example of majority mtDNA heteroplasmy induction in any system. Gene editing specificity as precise as a single nucleotide was observed in vivo for a protein-coding gene. Non-destructive genotyping enables single animal mtDNA analyses for downstream biological functional genomics applications. FusXTBE is a new gene editing toolkit for exploring important questions in mitochondrial biology and genetics.

category: genetics


title: Modeling The Effect of Forest Fire and Deforestation on Mid-Sized Wild Cat (Felids) Occupancy in SE Asian Tropical Rainforest

abstract: Felids are mammal groups that also experiencing the effects of forest fire and deforestation rate. By using camera detection method, two felid species, Prionailurus bengalensis and Pardofelis marmorata, of tropical rainforests in SE Asia have been studied. The studied area was a rainforest in Sumatra that has experienced several forest fires with annual deforestation rates of 1.69%-2.89%. Occupancy model using Akaike Information Criterion (AIC) is in agreement that deforestation rate is the best explanatory covariate explaining the declining occupancy of those felid species. P. marmorata was known more sensitive to the both deforestation rate and forest fire frequency covariate effects since it has similar AIC values. While P. bengalensis was slightly affected by forest fires. Values of Area Under The Curve (AUC) of Receiver Operating Characteristic (ROC) were > 0.5 and these indicate adequate probability of forest fire effects on felid occupancy. Cut off value of occupancy of P. bengalensis was higher than P. marmorata. For P. bengalensis, the cut off value was 1.75 leading to a sensitivity and specificity of 62%. This is the threshold value for the prediction of numbers of P. bengalensis individual occurred where both sensitivity and specificity are maximized and as an effect of forest fire, and this can be used to classify areas as occupied by P. bengalensis.

category: ecology


title: A practical guide for estimating animal density using camera traps: Focus on the REST model

abstract: Camera traps are increasingly popular in wildlife research and have the potential to be reliable and cost-effective for estimating animal density. Although density estimation using this automatic technique has long been restricted to species with individually recognizable markings, several analytical approaches have been proposed to target animals lacking such markings (i.e., unmarked populations). Among these approaches, the random encounter and staying time (REST) model may be an efficient and cost-effective approach, even though the procedures for the implementations have not yet been shared with researchers. This paper presents a working protocol for implementing the REST model our research group has been developing. We also present the R code to perform parameter estimation with a maximum likelihood and Bayesian approach. We suggest that this model has potential for further development. We strongly hope that this paper will encourage many researchers to use the REST for density estimation in a wide variety of species in various habitats and make a significant contribution to advancing wildlife conservation and management.


title: Estimating animal density with camera traps: a practitioner's guide of the REST model


title: In pursuit of a better broiler: A comparison of the inactivity, behavior, and enrichment use of fast- and slower-growing broiler chickens

abstract: Selection for rapid growth has produced heavier, more efficient broiler chickens, but has also introduced health and welfare issues, which may cause or be caused by inactivity. Rapid growth may also limit the performance of motivated behaviors, whereas the provision of enrichment may increase these behaviors and general activity. This study aimed to evaluate the inactivity, behavior, and enrichment use of 2 fast- (CONV) and 12 slower-growing broiler strains [categorized as fastest (FAST), moderate (MOD) and slowest slow (SLOW), based on their growth rates]. To evaluate inactivity, one male and one female from 153 pens were outfitted with omni-directional accelerometers from day 21 until processing (14-24 birds/strain from 8-12 pens/strain). Additionally, to supplement inactivity data, five-minute continuous behavioral observations of four focal birds per pen (2 males, 2 females) were conducted on days 26, 42, and 56 (72-148 observations of 8-12 pens/strain) to quantify the duration and frequency of various behaviors; at the same time, 5-11 instantaneous scan samples were also performed for pen-based enrichment use. Inactivity peaked at 78-80% of the day for all strains; however, those with slower growth rates reached these levels at older ages. Compared to slower-growing strains at the same age, faster-growing strains were more inactive, spent more time sitting and feeding, spent less time standing and walking, and used enrichments less; these differences mostly occurred at younger ages. Generally, at the same age, strains with similar growth rates (within the same category) behaved similarly, with only a few exceptions. Results suggest that not all strains identified as  slow-growing broilers behave differently from fast-growing broilers, nor do they all behave similarly to each other. As such, results suggest that improved broiler welfare, particularly with respect to reduced inactivity, the performance of a wider range of normal, motivated behaviors, and/or increased enrichment use, is related to the broiler strains specific growth rate.


title: Light-weight Electrophysiology Hardware and Software Platform for Cloud-Based Neural Recording Experiments

abstract:
ObjectiveNeural activity represents a functional readout of neurons that is increasingly important to monitor in a wide range of experiments. Extracellular recordings have emerged as a powerful technique for measuring neural activity because these methods do not lead to the destruction or degradation of the cells being measured. Current approaches to electrophysiology have a low throughput of experiments due to manual supervision and expensive equipment. This bottleneck limits broader inferences that can be achieved with numerous long-term recorded samples.

ApproachWe developed Piphys, an inexpensive open source neurophysiological recording platform that consists both hardware and software. It is easily accessed and controlled via a standard web interface through Internet of Things (IoT) protocols.

Main ResultsWe used a Raspberry Pi as the primary processing device and Intan bioamplifier. We designed a hardware expansion circuit board and software to enable voltage sampling and user interaction. This standalone system was validated with primary human neurons, showing reliability in collecting real-time neural activity.

SignificanceThe hardware modules and cloud software allow for remote control of neural recording experiments as well as horizontal scalability, enabling long-term observations of development, organization, and neural activity at scale.


title: Ultrafast amplitude modulation for molecular and hemodynamic ultrasound imaging

abstract: Ultrasound is playing an emerging role in molecular and cellular imaging thanks to new micro- and nanoscale contrast agents and reporter genes. Acoustic methods for the selective in vivo detection of these imaging agents are needed to maximize their impact in biology and medicine. Existing ultrasound pulse sequences use the nonlinearity in contrast agents response to acoustic pressure to distinguish them from mostly linear tissue scattering. However, such pulse sequences typically scan the sample using focused transmissions, resulting in a limited frame rate and restricted field of view. Meanwhile, existing wide-field scanning techniques based on plane wave transmissions suffer from limited sensitivity or nonlinear artifacts. To overcome these limitations, we introduce an ultrafast nonlinear imaging modality combining amplitude-modulated pulses, multiplane wave transmissions and selective coherent compounding. This technique achieves contrast imaging sensitivity comparable to much slower gold-standard amplitude modulation sequences and enables the acquisition of larger and deeper fields of view, while providing a much faster imaging framerate of 3.2kHz. Additionally, it enables simultaneous nonlinear and linear image formation, and allows concurrent monitoring of phenomena accessible only at ultrafast framerates, such as blood volume variations. We demonstrate the performance of this ultrafast amplitude modulation (uAM) technique by imaging gas vesicles, an emerging class of genetically encodable biomolecular contrast agents, in several in vitro and in vivo contexts. These demonstrations include the rapid discrimination of moving contrast agents and the real-time monitoring of phagolysosomal function in the mouse liver.


title: Sequence-based modeling of genome 3D architecture from kilobase to chromosome-scale

abstract: The structural organization of the genome plays an important role in multiple aspects of genome function. Understanding how genomic sequence influences 3D organization can help elucidate their roles in various processes in healthy and disease states. However, the sequence determinants of genome structure across multiple spatial scales are still not well understood. To learn the complex sequence dependencies of multiscale genome architecture, here we developed a sequence-based deep learning approach, Orca, that predicts genome 3D architecture from kilobase to whole-chromosome scale, covering structures including chromatin compartments and topologically associating domains. Orca also makes both intrachromosomal and interchromosomal predictions and captures the sequence dependencies of diverse types of interactions, from CTCF-mediated to enhancer-promoter interactions and Polycomb-mediated interactions. Orca enables the interpretation of the effects of any structural variant at any size on multiscale genome organization and provides an in silico model to help study the sequence-dependent mechanistic basis of genome architecture. We show that the models accurately recapitulate effects of experimentally studied structural variants at varying sizes (300bp-80Mb) using only sequence. Furthermore, these sequence models enable in silico virtual screen assays to probe the sequence-basis of genome 3D organization at different scales. At the submegabase scale, the models predicted specific transcription factor motifs underlying cell-type-specific genome interactions. At the compartment scale, based on virtual screens of sequence activities, we propose a new model for the sequence basis of chromatin compartments: sequences at active transcription start sites are primarily responsible for establishing the expression-active compartment A, while the inactive compartment B typically requires extended stretches of AT-rich sequences (at least 6-12kb) and can form  passively without depending on any particular sequence pattern. Orca thus effectively provides an "in silico genome observatory" to predict variant effects on genome structure and probe the sequence-based mechanisms of genome organization.

category: bioinformatics


title: PaReBrick: PArallel REarrangements and BReakpoints identification toolkit

abstract:
MotivationHigh plasticity of bacterial genomes is provided by numerous mechanisms including horizontal gene transfer and recombination via numerous flanking repeats. Genome rearrangements such as inversions, deletions, insertions, and duplications may independently occur in different strains, providing parallel adaptation. Specifically, such rearrangements might be responsible for multi-virulence, antibiotic resistance, and antigenic variation. However, identification of such events requires laborious manual inspection and verification of phyletic pattern consistency.

ResultsHere we define the term "parallel rearrangements" as events that occur independently in phylogenetically distant bacterial strains and present a formalization of the problem of parallel rearrangements calling. We implement an algorithmic solution for the identification of parallel rearrangements in bacterial population, as a tool PaReBrick. The tool takes synteny blocks and a phylogenetic tree as input and outputs rearrangement events. The tool tests each rearrangement for consistency with a tree, and sorts the events by their parallelism score and provides diagrams of the neighbors for each block of interest, allowing the detection of horizontally transferred blocks or their extra copies and the inversions in which copied blocks are involved. We proved PaReBricks efficiency and accuracy and showed its potential to detect genome rearrangements responsible for pathogenicity and adaptation in bacterial genomes.

AvailabilityPaReBrick is written in Python and is available on GitHub


title: Peptimetric: Quantifying and visualizing differences in peptidomic data

abstract: Finding new sustainable means of diagnosing and treating diseases is one of the most pressing issues of our time. In recent years, several endogenous peptides have been found to be both excellent biomarkers for many diseases and to possess important physiological roles which may be utilized in treatments. The detection of peptides has been facilitated by the rapid development of biological mass spectrometry and now the combination of fast and sensitive high resolution MS instruments and stable nano HP-LC equipment sequences thousands of peptides in one single experiment. In most research conducted with these advanced systems, proteolytically cleaved proteins are analyzed and the specific peptides are identified by software dedicated for protein quantification using different proteomics workflows. Analysis of endogenous peptides with peptidomics workflows also benefit from the novel sensitive and advanced instrumentation, however, the generated peptidomic data is vast and subsequently laborious to visualize and examine, creating a bottleneck in the analysis. Therefore, we have created Peptimetric, an application designed to allow researchers to investigate and discover differences between peptidomic samples. Peptimetric allows the user to dynamically and interactively investigate the proteins, peptides, and some general characteristics of multiple samples, and is available as a web application at https://peptimetric.herokuapp.com. To illustrate the utility of Peptimetric, weve applied it to a peptidomic dataset of 15 urine samples from diabetic patients and corresponding data from healthy subjects.


title: UMGAP: the Unipept MetaGenomics Analysis Pipeline

abstract: Shotgun metagenomics is now commonplace to gain insights into communities from diverse environments, but fast, memory-friendly, and accurate tools are needed for deep taxonomic analysis of the metagenome data. To meet this need we developed UMGAP, a highly versatile open source command line tool implemented in Rust for taxonomic profiling of shotgun metagenomes. It differs from state-of-the-art tools in its use of protein code regions identified in short reads for robust taxonomic identifications, a broad-spectrum index that can identify both archaea, bacteria, eukaryotes and viruses, a non-monolithic design, and support for interactive visualizations of complex biodiversities.


title: Protein detection in blood with single-molecule imaging

abstract:
The ability to identify and characterize individual biomarker protein molecules in patient blood samples could enable diagnosis of diseases at an earlier stage, when treatment is typically more effective. Single-molecule imaging offers a promising approach to accomplish this goal. However, thus far single-molecule imaging methods have only been used to monitor protein molecules in solutions or cell lysates, and have not been translated into the clinical arena. Furthermore, the detection limit of these methods has been confined to the picomolar (10-12 M) range. In many diseases, the circulating concentrations of biomarker proteins fall several orders of magnitude below this range. Here we describe Single-Molecule Augmented Capture (SMAC), a single-molecule imaging technique to visualize, quantify, and characterize individual protein molecules of interest down to the subfemtomolar (<10-15 M) range, even in complex biologic fluids. We demonstrate SMAC in a wide variety of applications with human blood samples, including the analysis of disease-associated secreted proteins, membrane proteins, and rare intracellular proteins. Using ovarian cancer as a model, a lethal malignancy in which high-grade disease is driven almost universally by alterations in the TP53 gene and frequently only diagnosed at a late, incurable stage, we found that mutant pattern p53 proteins are released into the blood in patients at an early stage in disease progression. SMAC opens the door to the application of single-molecule imaging in non-invasive disease profiling and allows for the analysis of circulating mutant proteins as a new class of highly specific disease biomarkers. The SMAC platform can be adapted to multiplex or high-throughput formats to characterize heterogeneous biochemical and structural features of circulating proteins-of-interest.

One Sentence SummaryA single-molecule imaging approach detects individual disease-specific protein molecules, including mutant intracellular proteins, in blood samples.


title: A coarse-grained approach to model the dynamics of the actomyosin cortex

abstract: The dynamics of the actomyosin machinery is at the core of many important biological processes. Several relevant cellular responses such as the rhythmic compression of the cell cortex are governed, at a mesoscopic level, by the nonlinear interaction between actin monomers, actin crosslinkers and myosin motors. Coarse grained models are an optimal tool to study actomyosin systems, since they can include processes that occur at long time and space scales, while maintaining the most relevant features of the molecular interactions. Here, we present a coarse grained model of a two-dimensional actomyosin cortex, adjacent to a three-dimensional cytoplasm. Our simplified model incorporates only well characterized interactions between actin monomers, actin crosslinkers and myosin, and it is able to reproduce many of the most important aspects of actin filament and actomyosin network formation, such as dynamics of polymerization and depolymerization, treadmilling, network formation and the autonomous oscillatory dynamics of actomyosin. Furthermore, the model can be used to predict the in vivo response of actomyosin networks to changes in key parameters of the system, such as alterations in the anchor of actin filaments to the cell cortex.


title: Protein resonance assignment by solid-state NMR based on 1H-detected 13C-based double-quantum spectroscopy at fast MAS

abstract: Solid-state NMR spectroscopy is a powerful technique to study insoluble and non-crystalline proteins and protein complexes at atomic resolution. The development of proton (1H) detection at fast magic-angle spinning (MAS) has considerably increased the analytical capabilities of the technique, enabling the acquisition of 1H-detected fingerprint experiments in few hours. Here an approach based on double-quantum (DQ) 13C spectroscopy, detected on 1H, is introduced at fast MAS (70 kHz) to perform the sequential assignment of insoluble proteins of small size, without any specific deuteration requirement. By combining two three-dimensional 1H detected experiments correlating a 13C DQ dimension respectively to its intra-residue and sequential 15N-1H pairs, a sequential walk through DQ (C+CO) resonance is obtained. Our approach takes advantage of fast MAS to achieve an efficient sensitivity and the addition of a DQ dimension provides spectral features useful for the resonance assignment process.


title: Viscoelasticity of basal plasma membranes and cortices derived from MDCK II cells

abstract: In mature epithelial cells, however, cells adhere to one another through tight junctions, adherens junctions and desmosomes thereby displaying a pronounced apical-basal polarity. In vivo, the apical membrane has a larger surface area and faces the outer surface of the body or the lumen of internal cavities, whereas the basolateral membrane is oriented on the side away from the lumen and forms focal adhesions with the extracellular matrix. The mechanical properties of cells are largely determined by the architecture and dynamics of their viscoelastic cortex, which consists of a contractile, cross-linked actin mesh attached to the plasma membrane via linker proteins. Measuring the mechanical properties of adherent, polarized epithelial cells is usually limited to the upper, i.e., apical side of the cells due to their accessibility on culture dishes. Moreover, contributions from the cell interior comprising various filament types, organelles, and the crowded cytoplasm usually impede examination of the cortex alone. Here, we investigate the viscoelastic properties of basolateral membranes derived from polarized MDCK II epithelia in response to external deformation and compare them to living cells probed at the apical side. Therefore, we grew MDCK II cells on porous surfaces to confluency and removed the upper cell body by sandwich cleavage. The free-standing, defoliated cortices were subject to force indentation and relaxation experiments permitting a precise assessment of cortical viscoelasticity. A new theoretical framework to describe the force cycles is developed and applied to obtain the time-dependent area compressibility modulus of cell cortices from adherent cells. Compared to the viscoelastic response of living cells the basolateral membranes are substantially less fluid and stiffer but obey to the same universal scaling law if excess area is taken into account.


title: Single molecule characterization of the binding kinetics of a transcription factor and its modulation by DNA sequence and methylation

abstract: The interaction of transcription factors with their response elements in DNA is emerging as a highly complex process, whose characterization requires measuring the full distribution of binding and dissociation times in a well-controlled assay. Here, we present a single-molecule assay that exploits the thermal fluctuations of a DNA hairpin, to detect the association and dissociation of individual, unlabeled transcription factors. We demonstrate this new approach by following the binding of Egr1 to its consensus motif and the three binding sites found in the promoter of the Lhb gene, and find that both association and dissociation are modulated by the 9 bp core motif and the sequences around it. In addition, CpG methylation modulates the dissociation kinetics in a sequence and position-dependent manner, which can both stabilize or destabilize the complex. Together, our findings show how variations in sequence and methylation patterns synergistically extend the spectrum of a proteins binding properties, and demonstrate how the proposed approach can provide new insights on the function of transcription factors.


title: Plasmid-encoded H-NS controls extracellular matrix composition in a modern Acinetobacter baumannii urinary isolate

abstract:
Acinetobacter baumannii is emerging as a multidrug-resistant (MDR) nosocomial pathogen of increasing threat to human health worldwide. The recent MDR urinary isolate UPAB1 carries the plasmid pAB5, a member of a family of large conjugative plasmids (LCP). LCP encode several antibiotic resistance genes and repress the type VI secretion system (T6SS) to enable their dissemination, employing two TetR transcriptional regulators. Furthermore, pAB5 controls the expression of additional chromosomally encoded genes, impacting UPAB1 virulence. Here we show that a pAB5-encoded H-NS transcriptional regulator represses the synthesis of the exopolysaccharide PNAG and the expression of a previously uncharacterized three-gene cluster that encodes a protein belonging to the CsgG/HfaB family. Members of this protein family are involved in amyloid or polysaccharide formation in other species. Deletion of the CsgG homolog abrogated PNAG production and Cup pili formation, resulting in a subsequent reduction in biofilm formation. Although this gene cluster is widely distributed in Gram-negative bacteria, it remains largely uninvestigated. Our results illustrate the complex cross-talks that take place between plasmids and the chromosomes of their bacterial host, which in this case can contribute to the pathogenesis of Acinetobacter.

IMPORTANCEThe opportunistic human pathogen Acinetobacter baumannii displays the highest reported rates of multidrug resistance among Gram-negative pathogens. Many A. baumannii strains carry large conjugative plasmids like pAB5. In recent years, we have witnessed an increase in knowledge about the regulatory cross-talks between plasmids and bacterial chromosomes. Here we show that pAB5 controls the composition of the bacterial extracellular matrix, resulting in a drastic reduction in biofilm formation. The association between biofilm formation, virulence, and antibiotic resistance is well-documented. Therefore, understanding the factors involved in the regulation of biofilm formation in Acinetobacter has remarkable therapeutic potential.

category: microbiology


title: NAP1L1 and NAP1L4 binding to Hypervariable Domain of Chikungunya Virus nsP3 Protein is bivalent and requires phosphorylation

abstract:
Chikungunya virus (CHIKV) is one of the most pathogenic members of the Alphavirus genus in the Togaviridae family. Within the last two decades, CHIKV has expanded its presence to both hemispheres and is currently circulating in both Old and New Worlds. Despite the severity and persistence of the arthritis it causes in humans, no approved vaccines or therapeutic means have been developed for CHIKV infection. Replication of alphaviruses, including CHIKV, is determined not only by their nonstructural proteins, but also by a wide range of host factors, which are indispensable components of viral replication complexes (vRCs). Alphavirus nsP3s contain hypervariable domains (HVDs), which encode multiple motifs that drive recruitment of cell- and virus-specific host proteins into vRCs. Our previous data suggested that NAP1 family members are a group of host factors that may interact with CHIKV nsP3 HVD. In this study, we performed a detailed investigation of the NAP1 function in CHIKV replication in vertebrate cells. Our data demonstrate that i) the NAP1-HVD interactions have strong stimulatory effects on CHIKV replication; ii) both NAP1L1 and NAP1L4 interact with the CHIKV HVD; iii) NAP1 family members interact with two motifs, which are located upstream and downstream of the G3BP-binding motifs of CHIKV HVD; iv) NAP1 proteins interact only with a phosphorylated form of CHIKV HVD and HVD phosphorylation is mediated by CK2 kinase; v) NAP1 and other families of host factors redundantly promote CHIKV replication and their bindings have additive stimulatory effects on viral replication.

IMPORTANCECellular proteins play critical roles in the assembly of alphavirus replication complexes (vRCs). Their recruitment is determined by the viral nonstructural protein 3 (nsP3). This protein contains a long, disordered hypervariable domain (HVD), which encodes virus-specific combinations of short linear motifs interacting with host factors during vRC assembly. Our study defined the binding mechanism of NAP1 family members to CHIKV HVD and demonstrated a stimulatory effect of this interaction on viral replication. We showed that interaction with NAP1L1 is mediated by two HVD motifs and requires phosphorylation of HVD by CK2 kinase. Based on the accumulated data, we present a map of the binding motifs of the critical host factors currently known to interact with CHIKV HVD. It can be used to manipulate cell specificity of viral replication and pathogenesis, and to develop a new generation of vaccine candidates.


title: A community-science approach identifies genetic variants associated with three color morphs in ball pythons (Python regius)

abstract: Color morphs in ball pythons (Python regius) provide a unique and largely untapped resource for understanding the genetics of coloration in reptiles. Here we use a community-science approach to investigate the genetics of three color morphs affecting production of the pigment melanin. These morphs--Albino, Lavender Albino, and Ultramel--show a loss of melanin in the skin and eyes, ranging from severe (Albino) to moderate (Lavender Albino) to mild (Ultramel). To identify genetic variants causing each morph, we recruited shed skins of pet ball pythons via social media, extracted DNA from the skins, and searched for putative loss-of-function variants in homologs of genes controlling melanin production in other vertebrates. We report that the Albino morph is associated with missense and non-coding variants in the gene TYR. The Lavender Albino morph is associated with a deletion in the gene OCA2. The Ultramel morph is associated with a missense variant and a putative deletion in the gene TYRP1. Our study is one of the first to identify genetic variants associated with color morphs in ball pythons and shows that pet samples recruited from the community can provide a resource for genetic studies in this species.


title: Phospholipases D1 and D2 regulate cell cycling in primary prostate cancer cells and are differentially associated with the nuclear matrix

abstract: Phospholipases D1 and D2 (PLD1/2) have been implicated in tumorigenesis. We previously detected higher expression of PLD in the nuclei of patient-derived prostate cancer (PCa) cells and prostate cancer cell lines. Here we have examined whether PLD1 or PLD2 are associated with the nuclear matrix and influence cell cycling. PLD1/PLD2 were detected by qualitative immunofluorescence in cultured PCa cells and extracted with a standardised protocol to reveal nuclear matrix-associated proteins. The effects of isoform-specific inhibition of PLD1or PLD2 on PCa cell cycle progression were analysed by flow cytometry. PLD2 mainly co-localised with the nucleolar marker fibrillarin in PCa cells. However, even after complete extraction, some PLD2 remained associated with the nuclear matrix. Inhibiting PLD2 effectively reduced PCa cell cycling into and through S phase. In contrast, PLD1 inhibition effects were weaker, and a subpopulation of cycling patient-derived PCa cells was unaffected by PLD1 inhibition. When associated with the nuclear matrix PLD2 could generate phosphatidic acid to regulate nuclear mTOR and control downstream transcriptional events. The association of PLD2 with the nucleolus also implies a role in stress regulation. The cell cycling results highlight the importance of PLD2 inhibition as a novel potential prostate cancer therapeutic mechanism by differential regulation of cell proliferation.

category: cancer biology


title: The misleading certainty of uncertain data in biological network processes

abstract: Mathematical models are often used to explore network-driven cellular processes from a systems perspective. However, a dearth of quantitative data suitable for model calibration leads to models with parameter unidentifiability and questionable predictive power. Here we introduce a Bayesian and Machine-Learning based Measurement Model approach to explore how quantitative and non-quantitative data constrain models of apoptosis execution within a missing data context. We find two orders of magnitude more ordinal (e.g. immunoblot) data are necessary to achieve accuracy comparable to quantitative (e.g. fluorescence) data. Notably, ordinal and nominal (e.g. immunostain) non-quantitative data synergize to reduce model uncertainty and improve accuracy. Further, model prediction accuracy and certainty strongly depend on rigorous data-driven formulations of the measurement, and the size and make-up of the datasets. Finally, we demonstrate the potential of a data-driven Measurement Model approach to identify model features that could lead to informative experimental measurements and improve model predictive power.

category: systems biology


title: Predictive uncertainty in mechanistic models of cellular processes calibrated to experimental data


title: A Model for Self-Organization of Sensorimotor Function: The Spinal Monosynaptic Loop

abstract: Recent spinal cord literature abounds with descriptions of genetic preprogramming and the molecular control of circuit formation. In this paper we explore to what extent circuit formation based on learning rather than preprogramming could explain some prominent aspects of the spinal cord connectivity patterns observed in animals. To test this we developed an artificial organism with a basic musculoskeletal system and proprioceptive sensors, connected to a neural network. We adjusted the initially randomized gains in the neural network according to a Hebbian plasticity rule while exercising the model system with spontaneous muscle activity patterns similar to those observed during early fetal development. The resulting connection matrices support functional self-organization of the mature pattern of Ia to motoneuron connectivity in the spinal circuitry. More coordinated muscle activity patterns such as observed later during neonatal locomotion impaired projection selectivity. These findings imply a generic functionality of a musculoskeletal system to imprint important aspects of its mechanical dynamics onto a neural network, without specific preprogramming other than setting a critical period for the formation and maturation of this general pattern of connectivity. Such functionality would facilitate the successful evolution of new species with altered musculoskeletal anatomy and it may help to explain patterns of connectivity and associated reflexes that appear during abnormal development.


title: Multiframe Evolving Dynamic Functional Connectivity (EVOdFNCs): A Method for Constructing and Investigating Functional Brain Motifs

abstract: The study of brain network connectivity as a time-varying property began relatively recently and to date has remained primarily concerned with capturing a handful of discrete static states that characterize connectivity as measured on a timescale shorter than that of the full scan. Capturing group-level representations of temporally evolving patterns of connectivity is a challenging and important next step in fully leveraging the information available in large resting state functional magnetic resonance imaging (rs-fMRI) studies. We introduce a flexible, extensible data-driven framework for the stable identification of group-level multiframe (movie-style) dynamic functional network connectivity (dFNC) states. Our approach employs uniform manifold approximation and embedding (UMAP) to produce a continuity-preserving planar embedding of high-dimensional time-varying measurements of whole-brain functional network connectivity. Planar linear exemplars summarizing dominant dynamic trends across the population are computed from local linear approximations to the 2D embedded trajectories. A high-dimensional representation of each 2D exemplar segment is obtained by averaging the dFNC observations corresponding to the n planar nearest neighbors of {tau} evenly spaced points along the 2D line segment representation (where n is the UMAP number-of-neighbors parameter and {tau} is the temporal duration of trajectory segments being approximated). Each of the 2D exemplars thus "lifts" to a multiframe high-dimensional dFNC trajectory of length {tau}. The collection of high-dimensional temporally evolving dFNC representations (EVOdFNCs) derived in this manner are employed as dynamic basis objects with which to characterize observed high-dimensional dFNC trajectories, which are then expressed as weighted combination of these basis objects. Our approach yields new insights into anomalous patterns of fluidly varying whole brain connectivity that are significantly associated with schizophrenia as a broad diagnosis as well as with certain symptoms of this serious disorder. Importantly, we show that relative to conventional hidden Markov modeling with single-frame unvarying dFNC summary states, EVOdFNCs are more sensitive to positive symptoms of schizophrenia including hallucinations and delusions, suggesting a more dynamic characterization is needed to help illuminate such a complex brain disorder.


title: Regions of visual cortex responding to tactile stimulation in an individual withlongstanding low vision are not causally involved in tactile processing performance

abstract:
Braille reading and other tactile discrimination tasks recruit the visual cortex of both blind and normally sighted individuals undergoing short-term visual deprivation. Prior functional magnetic resonance imaging (fMRI) work in patient  S, a visually impaired adult with the rare ability to read both highly magnified print visually and Braille by touch, found that foveal representations of Ss visual cortex were recruited during tactile perception, whereas peripheral regions were recruited during visual perception. Here, we test the causal nature of tactile responses in the visual cortex of S by combining tactile and visual psychophysics with repetitive transcranial magnetic stimulation (rTMS). First, we replicate this prior fMRI work in S. Second, we demonstrate that transient disruption of Ss foveal visual cortex has no measurable impact on Ss tactile processing performance compared to that of healthy controls - a pattern not predicted by the fMRI results. Third, stimulation of foveal visual cortex maximally disrupted visual processing performance in both S and controls, suggesting the possibility of preserved visual function within Ss foveal cortex. Finally, stimulation of somatosensory cortex induced the expected disruption to tactile processing performance in both S and controls. These data suggest that tactile responses in Ss foveal representation reflect unmasking of latent connections between visual and somatosensory cortices and not behaviourally relevant cross-modal plasticity. Unlike studies in congenitally blind individuals, it is possible that the absence of complete visual loss in S has limited the degree of causally impactful cross-modal reorganisation.

Significance statementPrior fMRI work in patient  S identified that foveal portions of Ss visual cortex respond more to tactile processing, whereas peripheral portions respond more to visual processing. Here, we tested whether this foveal processing was causally related to either tactile or visual processing. First, using fMRI we replicate prior work. Second, we demonstrate that TMS of the foveal representation and of somatosensory cortex interfered with visual and tactile discriminations respectively in controls and crucially also in S. The foveal representation in S, which is responsive to tactile stimulation, does not however play a causal role in mediating Ss ability to discriminate Braille characters and likely reflects the unmasking of latent connections between visual and somatosensory cortices.


title: A METHOD FOR INTEGRATIVE ANALYSIS OF LOCAL AND GLOBAL BRAIN DYNAMICS

abstract: The most common pipelines for studying time-varying network connectivity in resting state functional magnetic resonance imaging (rs-fMRI) operate at the whole brain level, capturing a small discrete set of "states" that best represent time-resolved joint measures of connectivity over all network pairs in the brain. This whole-brain hidden Markov model (HMM) approach "uniformizes" the dynamics over what is typically more than 1000 pairs of networks, forcing each time-resolved high-dimensional observation into its best-matched high-dimensional state. While straightforward and convenient, this HMM simplification obscures functional and temporal nonstationarities that could reveal systematic, informative features of resting state brain dynamics at a more granular scale. We introduce a framework for studying functionally localized dynamics that intrinsically embeds them within a whole-brain HMM frame of reference. The approach is validated in a large rs-fMRI schizophrenia study where it identifies group differences in localized patterns of entropy and dynamics that help explain consistently observed differences between schizophrenia patients and controls in occupancy of whole-brain dFNC states more mechanistically.


title: CARDIOVASCULAR AUTONOMIC NEUROPATHY IN TYPE 1 DIABETES IS ASSOCIATED WITH SEVERAL METABOLIC PATHWAYS: New Risk Markers on the horizon

abstract:
ObjectiveCardiovascular autonomic neuropathy (CAN) in diabetes is associated with increased mortality and morbidity and is a non-treatable complication. We investigated associations between circulating metabolites and presence of CAN in persons with type 1 diabetes (T1D).

MethodsCAN was assessed by cardiovascular reflex tests (CARTs) in 302 persons with T1D as heart rate response to: deep breathing; lying-to-standing test; and the Valsalva manoeuvre. More than 1 pathological CART defined the CAN diagnosis.

Serum metabolomics and lipidomics profiles were analysed with two complementary non-targeted massspectrometry methods. Cross-sectional associations between single metabolites and CAN were assessed by linear regression. Models were fitted with and without adjustments for relevant confounders and multiple testing.

ResultsParticipants were mean (IQR) aged 55(49, 63) years, 50% males, with diabetes duration 39(32, 47) years, HbA1c 63(55,69) mmol/mol and 34% had the CAN diagnosis.

A total of 75 metabolites and 106 lipids were examined. In crude models, CAN diagnosis was associated with higher levels of hydroxy fatty acids (2,4- and 3,4-dihydroxybutanoic acids, 4-deoxytetronic acid), creatinine, sugar derivates (ribitol, ribonic acid, myo-inositol), citric acid, glycerol, phenols, phosphatidylcholines and lower levels of free fatty acids and amino acid methionine (p<0.05). Upon adjustment, positive associations with CAN were retained with hydroxy fatty acids, tricarboxylic acid (TCA) cycle-based sugar derivates, and citric acid and phenols (Padjusted<0.05).

ConclusionsMetabolic pathways, including the TCA cycle, hydroxy fatty acids, phosphatidylcholines and sugar derivatives, were associated with CAN in T1D. These novel metabolic pathways associated with CAN could prove to be future modifiable risk factors.


title: Out-of-sight or out-of-site? Forays and detection probabilities in single-season occupancy models

abstract: Occupancy models have become popular in wildlife survey analyses because they account for the frequent failure to detect individuals of targeted species. Those individuals sometimes move outside sampling sites, i.e. exhibit temporary emigration. In such cases, occupancy models may become difficult to interpret or even misleading either at the species or the individual level, because they confound presence at the site, availability for detection given presence, and actual detection by the observer. We quantified the probabilities of these three components with spruce grouse (Falcipennis canadensis) in southern Quebec, Canada. We conducted call-response surveys of 24 grouse monitored by radio-telemetry. We defined sites empirically as circular areas of 83 m radius centered on the observer, corresponding to the maximum detection distance obtained. Based on telemetry locations, grouse were present at the site during 42 % of the surveys. Six stationary grouse were present during surveys, but were never detected. Thus, only individuals that moved in the presence of the observer (89 %) were considered available for detection. Individuals available for detection were detected in 51 % of the cases. We simulated detection histories and built single-season occupancy models, based on the empirical relationship between detection probability and the distance measured between observers and grouse. When temporary emigration was ignored, site occupancy was{psi} = 0.89, and the associated probability of detection was p = 0.23. When instances of temporary emigration were dropped, estimates were{psi} = 0.88 and p = 0.41. Using only grouse available for detection, estimates were{psi} = 0.87 and p = 0.42. Disentangling the components of detection probabilities had little impact on occupancy estimates, but showed a major effect of temporary emigration on estimated detection probabilities.


title: Distinct effects of host and neighbour tree identity on arbuscular and ectomycorrhizal fungi along a tree diversity gradient

abstract: Plant diversity and plant-related ecosystem functions have been in focus in biodiversity-ecosystem functioning studies. However, in this context, biotic interactions with mycorrhizal fungi have been understudied although they are crucial for plant-resource acquisition. We investigated the effects of tree species richness, tree mycorrhizal type on arbuscular (AMF) and ectomycorrhizal fungal (EMF) communities. We aimed to understand how dissimilarities in taxa composition and beta-diversity are related to target trees and neighbours of the same/different mycorrhizal type. We sampled a tree experiment with saplings ([~]7 years old), where tree species richness (monocultures, 2-species, and 4-species mixtures) and mycorrhizal type were manipulated. AMF and EMF richness significantly increased with increasing tree species richness. AMF richness of mixture plots resembled that of the sum of the respective monocultures, whereas EMF richness of mixture plots was lower compared to the sum of the respective monocultures. Specialisation scores revealed significantly more specialised AMF than EMF suggesting that, in contrast to previous studies, AMF were more specialised, whereas EMF were not. We further found that AMF communities were little driven by the surrounding trees, whereas EMF communities were. Our study revealed the drivers of mycorrhizal fungal communities and further highlights the distinct strategies of AMF and EMF.


title: Gene flow influences the genomic architecture of local adaptation in six riverine fish species

abstract: Understanding how gene flow influences adaptive divergence is important for predicting adaptive responses. Theoretical studies suggest that when gene flow is high, clustering of adaptive genes in fewer genomic regions would protect adaptive alleles from among-population recombination and thus be selected for, but few studies have tested this hypothesis with empirical data. Here, we used RADseq to generate genomic data for six fish species with contrasting life histories from six reaches of the Upper Mississippi River System, USA. We then conducted genome scans for genomic islands of divergence to examine the distribution of adaptive loci and investigated whether these loci were found in inversions. We found that gene flow varied among species, and adaptive loci were clustered more tightly in species with higher gene flow. For example, the two species with the highest overall FST (0.03 - 0.07) and therefore lowest gene flow showed little evidence of clusters of adaptive loci, with adaptive loci spread uniformly across the genome. In contrast, nearly all adaptive loci in the species with the lowest FST (0.0004) were found in a single large putative inversion. Two other species with intermediate gene flow (FST ~ 0.004) also showed clustered genomic architectures, with most islands of divergence clustered on a few chromosomes. These results provide important empirical evidence to support the hypothesis that increasingly clustered architectures of local adaptation are associated with high gene flow. Our study utilized a unique system with species spanning a large gradient of life histories to highlight the importance of gene flow in shaping adaptive divergence.


title: CeAid: A smartphone application for logging and plotting C. elegans assays

abstract: C. elegans is used as a model organism to study a wide range of topics in molecular and cellular biology. Conventional C. elegans assays often require a large sample size with frequent manipulations, rendering them labor-intensive. Automated high-throughput workflows may not be always the best solution to reduce benchwork labor, as they may introduce more complexity. Thus, most assays are carried out manually, where logging and digitizing experimental data can be as time-consuming as picking and scoring worms. Here we report the development of CeAid, C. elegans Application for inputting data, which significantly expedites the data entry process, utilizing swiping gestures and a voice recognition algorithm for logging data using a standard smartphone or Android device. This modular platform can also be adapted for a wide range of assays where recording data is laborious, even beyond worm research.


title: Haplotype-aware inference of human chromosome abnormalities

abstract:
Extra or missing chromosomes--a phenomenon termed aneuploidy--frequently arises during human meiosis and embryonic mitosis and is the leading cause of pregnancy loss, including in the context of in vitro fertilization (IVF). While meiotic aneuploidies affect all cells and are deleterious, mitotic errors generate mosaicism, which may be compatible with healthy live birth. Large-scale abnormalities such as triploidy and haploidy also contribute to adverse pregnancy outcomes, but remain hidden from standard sequencing-based approaches to preimplantation genetic testing (PGT-A). The ability to reliably distinguish meiotic and mitotic aneuploidies, as well as abnormalities in genome-wide ploidy may thus prove valuable for enhancing IVF outcomes. Here, we describe a statistical method for distinguishing these forms of aneuploidy based on analysis of low-coverage whole-genome sequencing data, which is the current standard in the field. Our approach overcomes the sparse nature of the data by leveraging allele frequencies and linkage disequilibrium (LD) measured in a population reference panel. The method, which we term LD-informed PGT-A (LD-PGTA), retains high accuracy down to coverage as low as 0.05x and at higher coverage can also distinguish between meiosis I and meiosis II errors based on signatures spanning the centromeres. LD-PGTA provides fundamental insight into the origins of human chromosome abnormalities, as well as a practical tool with the potential to improve genetic testing during IVF.

Significance StatementWhole chromosome gains and losses--termed aneuploidies--are the leading cause of human pregnancy loss and congenital disorders. Recent work has demonstrated that in addition to harmful meiotic aneuploidies, mitotic aneuploidies (which lead to mosaic embryos harboring cells with different numbers of chromosomes) may also be common in preimplantation embryos but potentially compatible with healthy birth. Here we developed and tested a method for distinguishing these forms of aneuploidy using genetic testing data from 8154 IVF embryos. We re-classified embryos based on signatures of meiotic and mitotic error, while also revealing lethal forms of chromosome abnormality that were hidden to existing approaches. Our method complements standard protocols for preimplantation and prenatal genetic testing, while offering insight into the biology of early development.


title: Variation in female songbird state determines signal state needed to evoke copulation

abstract: It is the female response to male signals that determines courtship success. In most songbirds, females control reproduction via the copulation solicitation display (CSD), an innate, stereotyped posture produced in direct response to male displays. Because CSD can be elicited in the absence of males by the presentation of recorded song, CSD production enables investigations into the effects of underlying signal features and behavioral state on female mating preferences. Using computer vision to quantify CSD trajectory in female brown-headed cowbirds (Molothrus ater), we show that both song quality and a females internal state predict CSD production, as well as the onset latency and duration of the display. We also show that CSD can be produced in a graded fashion based on both signal strength and internal state. These results emphasize the importance of underlying receiver state in determining behavioral responses and suggest that female responsiveness acts in conjunction with male signal strength to determine the efficacy of male courtship.


title: Variation in female songbird state determines signal strength needed to evoke copulation


title: Open-Source Thermometer, Temperature Controller, and Light Meter for Use in Animal Facilities and During Experiments

abstract: Experiments with biological samples require precise control of environmental conditions. In our work we use zebrafish (Danio rerio) to understand the neurobiology of sleep, which requires precise control of temperature and lighting. Like many labs, lighting and temperature in the animal facility are centrally controlled in the building. During behavioral experiments and microscopy sessions, we use custom-built heating systems and perform occasional manual checks of conditions. However, without a system to precisely record conditions, gradual changes in temperature can go unnoticed for a long time, and temporary failures may be missed entirely. Here we present the design and characterization of affordable open-source tools to record temperature and light conditions during animal experiments using an Arduino microcontroller or a Raspberry Pi compact computer. The waterproof temperature sensor has high stability over 50 days of recording and is precise to 0.1{degrees}C. The Arduino device can be used through a common serial port interface for which we present code in Python and MATLAB. The Raspberry Pi version can be accessed through a web interface, for which we provide an installation guide. We use the device to record and review temperature and lighting conditions in two zebrafish animal facilities. We use our platform to add a water heating system to maintain temperature at 28{degrees}C during in vivo light-sheet imaging of larval zebrafish. We show that a change in temperature from 28{degrees}C to 32{degrees}C affects resting heart rate of the animal, highlighting the importance of maintaining and recording conditions. The protocols presented here do not require advanced engineering, fabrication, or software skills, and provide an approach to accurately record and report experimental conditions.


title: Converting Muscle-mimetic Biomaterials to Cartilage-like Materials

abstract: Load-bearing tissues, such as muscle and cartilage, exhibit mechanical properties that often combine high elasticity, high toughness and fast recovery, despite their different stiffness ([~]100 kPa for muscles and one to several MPa for cartilage).1-7 The advance in protein engineering and protein mechanics has made it possible to engineer protein-based biomaterials to mimic soft load-bearing tissues, such as muscles.8-10 However, it is challenging to engineer protein biomaterials to achieve the mechanical properties exhibited by stiff tissues, such as articular cartilage,6,11 or to develop stiff synthetic extracellular matrices for cartilage stem/progenitor cell differentiation12. By employing physical entanglements13 of protein chains and force-induced protein unfolding,14,15 here we report the engineering of a highly tough and stiff protein hydrogel to mimic articular cartilage. By crosslinking an engineered artificial elastomeric protein from its unfolded state, we introduced chain entanglement into the hydrogel network. Upon renaturation, the entangled protein chain network and forced protein unfolding entailed this single network protein hydrogel with superb mechanical properties in both tensile and compression tests, showing a Youngs modulus of [~]0.7 MPa and toughness of 250 kJ/m3 in tensile testing; and [~]1.7 MPa in compressive modulus and toughness of 3.2 MJ/m3. The energy dissipation in both tensile and compression tests is reversible and the hydrogel can recovery its mechanical properties rapidly. Moreover, this hydrogel can withstand a compression stress of >60 MPa without failure, amongst the highest compressive strength achieved by a hydrogel. These properties are comparable to those of articular cartilage, making this protein hydrogel a novel cartilage-mimetic biomaterial. Our study opened up a new potential avenue towards engineering protein hydrogel-based substitute for articular cartilage, and may also help develop protein biomaterials with superb mechanical properties for applications in soft actuators and robotics.


title: Comparison of confound adjustment methods in the construction of gene co-expression networks

abstract: Adjustment for confounding sources of expression variation is an important preprocessing step in large gene expression studies, but the effect of confound adjustment on co-expression network analysis has not been well-characterized. Here, we demonstrate that the choice of confound adjustment method can have a considerable effect on the architecture of the resulting co-expression network. We compare standard and alternative confound adjustment methods and provide recommendations for their use in the construction of gene co-expression networks from bulk tissue RNA-seq datasets.


title: Survival analysis of pathway activity as a prognostic determinant in breast cancer

abstract: Most activities in a cell are dependent on sets of biomolecules, often referred to as pathways. Pathways are useful to study as their activities are frequently more directly related to the behavior of the cell than their components. Traditional pathway analysis gives significance to differences in the pathway components concentrations between sample groups. Here we instead advocate a singular value decomposition-based method for estimating individual samples pathway activities that allow us to investigate more advanced statistical models. Specifically, we investigate the pathway activities association with patients survival time based on the transcription profiles of the METABRIC dataset. Our implementation shows that pathway activities are better prognostic markers for survival time in METABRIC than the individual transcripts. We also demonstrate that we can regress out the effect of individual pathways on other pathways, which allows us to estimate the other pathways residual pathway activity on survival.


title: How to Assemble a Scale-Invariant Gradient

abstract: Intracellular protein gradients serve a variety of functions, such as the establishment of cell polarity and to provide positional information for gene expression in developing embryos. Given that cell size in a population can vary considerably, for the protein gradients to work properly they often have to be scaled to the size of the cell. Here we examine a model of protein gradient formation within a cell that relies on cytoplasmic diffusion and cortical transport of proteins toward a cell pole. We show that the shape of the protein gradient is determined solely by the cell geometry. Furthermore, we show that the length scale over which the protein concentration in the gradient varies is determined by the linear dimensions of the cell, independent of the diffusion constant or the transport speed. This gradient provides scale-invariant positional information within a cell, which can be used for assembly of intracellular structures whose size is scaled to the linear dimensions of the cell, as was recently reported for the cytokinetic ring and actin cables in budding yeast cells.