title: Relating genome completeness to functional predictions

abstract: Genome and transcriptome assemblies vary in their quality in terms of the connectedness of the assemblies and the amount of biological information captured. Interpreting de novo assemblies from new, poorly characterized, organisms in the context of complex traits can be challenging because, in the absence of a reference, it is difficult to know how much information is enough to claim the presence or absence of a trait. This study uses randomly downsampled proteome files to compare a genome completeness metric, BUSCO, to functional predictions of the complex trait of phagocytosis in known phagocytotic organisms broadly across the eukaryotic tree of life. We find that as additional proteins are added, BUSCO scores increase incrementally, while the phagocytosis prediction follows a sigmoidal curve. Generalizing our findings, we suggest a threshold of the number of BUSCOs detected above which one would expect an accurate prediction, positive or negative, of the complex trait of phagocytosis. While these findings are specific to a single trait, the methods can be extended to consider additional functional traits and predictive frameworks.


title: Daily ranging and den usage patterns structure fission-fusion dynamics and social associations in spotted hyenas

abstract: Environment structure often shapes social interactions. Spatial attractors that draw multiple individuals may play a particularly important role in dispersed groups, where individuals must first encounter one another to interact. We use GPS data recorded simultaneously from five spotted hyenas (Crocuta crocuta) within a single clan to investigate how communal dens and daily ranging patterns shape fission-fusion dynamics (subgroup splits and merges). We introduce a species-general framework for identifying and characterizing dyadic fission-fusion events and describe a taxonomy of ten possible configurations of these events. Applying this framework to the hyena data illuminates the spatiotemporal structure of social interactions within hyenas daily routines. The most common types of fission-fusion events involve close approaches between individuals, do not involve co-travel together, and occur at the communal den. Comparison to permutation-based reference models suggests that den usage structures broad-scale patterns of social encounters, but that other factors influence how those encounters unfold. We discuss the dual role of communal dens in hyenas as physical and social resources, and suggest that dens are an example of a general "social piggybacking" process whereby environmental attractors take on social importance as reliable places to encounter conspecifics, causing social and spatial processes to become fundamentally intertwined.

category: animal behavior and cognition


title: Resting together, foraging apart: Network structure of foraging bats is not explained by energy expenditure or foraging efficiency

abstract:
Group foraging animals may change their spatial relationships in response to the resource landscape. The strength of social network ties across foraging groups could then reflect the energy balance of individual group members through their effect on foraging efficiency. Greater spear-nosed bats (Phyllostomus hastatus) live in social groups where unrelated females develop group-specific social calls, defend each others offspring, and are hypothesized to coordinate social foraging to feed on balsa flowers. To test if social groups forage together, and if this increases foraging efficiency, we tracked 18 individuals from three social groups of P. hastatus on Isla Colon, Panama using high frequency GPS for 1 - 3 days each. We found that individuals commuted alone but then joined group members in small foraging patches with high densities of flowering balsas, 25 km from their roost on the mainland. Proximity-based social network measures did not predict foraging efficiency or energy expenditure, but individuals with higher social associations spent more time outside of the cave. Individuals did not coordinate movement, but close proximity among individuals increased resting bout duration and synchrony, particularly when in the same patch. These results show that stronger social proximity associations were more closely tied to resting behavior, and indicate that factors other than increased feeding efficiency may structure network relationships of group members while foraging.

Lay SummarySocial foraging can increase foraging efficiency on hard-to-find food sources. Miniaturized high-frequency GPS allowed us to test this on groups of greater spear-nosed bats. We estimated food intake rates, energy expenditure, and the strength of bat social relationships. In contrast to our expectations, we did not find a relationship between bat social relationships and energy expenditure or foraging efficiency. However, bats took longer resting breaks when group members were close. Why these bats commute 25 km individually across the ocean to then forage near each other thus requires further investigation.


title: Greater spear nosed bats in Panama do not use social proximity to improve foraging efficiency


title: Study of Metyltetraprole, an unusual agrofungicide targeting the Qo-site of cytochrome bc1 complex

abstract: The mitochondrial respiratory chain bc1 complex is a proven target of agrofungicides. Most of them are Qo-site antagonists (i.e QoIs), competing with the substrate ubiquinol, and likely share the same binding mode as the widespread Qo-site resistance mutation G143A confers cross-resistance. Metyltetraprole (MTP) presents an exception as studies with phytopathogenic fungi showed that the inhibitor was unaffected by G143A. Here, we used the yeast model to investigate its mode of action. Analysis of bc1 complex mutants supports a Qo-site binding for MTP. However the compound seems distinct to other QoIs, such as azoxystrobin, in various ways, namely; 1) G143A was without effect on MTP, as previously reported. 2) The level of MTP resistance of mutants was higher in bc1 complex activity assays than in growth assays while the opposite was observed with azoxystrobin. 3) Steady-state kinetics used to characterise the mode of action of MTP also revealed differences compared to other QoIs.

category: biochemistry


title: Rad24-RFC loads the 9-1-1 clamp by inserting DNA from the top of a wide-open ring, opposite the mechanism of RFC/PCNA

abstract: In response to DNA damage, the ring-shaped 9-1-1 clamp is loaded onto 5 recessed DNA to arrest the cell cycle and activate the DNA damage checkpoint. The 9-1-1 clamp is a heterotrimeric ring that is loaded in S. cerevisiae by Rad24-RFC, an alternative clamp loader in which Rad24 replaces the Rfc1 subunit in the RFC1-5 clamp loader of PCNA. Unlike RFC that loads the PCNA ring onto a 3-ss/ds DNA junction, Rad24-RFC loads the 9-1-1 ring onto a 5-ss/ds DNA junction, a consequence of DNA damage. The underlying 9-1-1 clamp loading mechanism has been a mystery. Here we report two 3.2-[A] cryo-EM structures of Rad24-RFC bound to DNA and either a closed or 27 [A] open 9-1-1 clamp. The structures reveal a completely unexpected mechanism by which a clamp can be loaded onto DNA. The Rad24 subunit specifically recognizes the 5-DNA junction and holds ds DNA outside the clamp loader and above the plane of the 9-1-1 ring, rather than holding DNA inside and below the clamp as in RFC. The 3 ssDNA overhang is required to obtain the structure, and thus confers a second DNA binding site. The bipartite DNA binding by Rad24-RFC suggests that ssDNA may be flipped into the open 9-1-1 ring, similar to ORC-Cdc6 that loads the Mcm2-7 ring on DNA. We propose that entry of ssDNA through the 9-1-1 ring triggers the ATP hydrolysis and release of the Rad24-RFC. The key DNA binding residues are conserved in higher eukaryotes, and thus the 9-1-1 clamp loading mechanism likely generalizes.


title: Cell-Penetrating D-Peptides Retain Antisense Morpholino Oligomer Delivery Activity

abstract:
Cell-penetrating peptides (CPPs) can cross the cell membrane to enter the cytosol and deliver otherwise non-penetrant macromolecules such as proteins and oligonucleotides. For example, recent clinical trials have shown that a CPP attached to phosphorodiamidate morpholino oligomers (PMO) resulted in higher muscle concentration, increased exon-skipping and dystrophin production relative to another study of the PMO alone in patients of Duchenne muscular dystrophy. Therefore, effective design and study of CPPs could help enhance therapies for difficult-to-treat diseases. So far, the study of CPPs for PMO delivery has been restricted to predominantly canonical L-peptides. We hypothesized that mirror-image D-peptides could have similar PMO delivery activity as well as enhanced proteolytic stability, facilitating their characterization and quantification from biological milieu. We found that several enantiomeric peptide sequences could deliver a PMO-biotin cargo with similar activities, while remaining stable against serum proteolysis. The biotin label allowed for affinity capture of fully intact PMO-peptide conjugates from whole cell and cytosolic lysates. By profiling a mixture of these constructs in cells, we determined their relative intracellular concentrations. When combined with PMO activity, these concentrations provide a new metric for delivery efficiency which may be useful for determining which peptide sequence to pursue in further pre-clinical studies.

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category: bioengineering


title: Alkaline phosphatase activity present in serum influences osteogenic differentiation cultures

abstract:
Fetal bovine serum (FBS) is a widely used supplement in cell culture medium, despite its known variability in composition which greatly affects cellular function and consequently the outcome of studies. In bone tissue engineering, the deposited mineralized matrix is one of the main outcome parameters, but using different brands of FBS can result in large variations. Alkaline phosphatase (ALP) is present in FBS. Not only is ALP used to judge the osteogenic differentiation of bone cells, it may affect deposition of mineralized matrix. The present study focused on the enzymatic activity of ALP in FBS of different suppliers and its contribution to mineralization in osteogenic differentiation cultures. It was hypothesized that culturing cells in a medium with high intrinsic ALP activity of FBS will lead to higher mineral deposition compared to media with lower ALP activity. The used FBS types were shown to have significant differences in enzymatic ALP activity. Our results indicate that the ALP activity of the medium not only affected the deposited mineralized matrix but also the osteogenic differentiation of cells as measured by a changed cellular ALP activity of human bone marrow derived mesenchymal stromal cells (hBMSC). In media with low inherent ALP activity, the cellular ALP activity was increased and played the major role in the mineralization process; while, in media with high intrinsic ALP activity contribution from the serum, less cellular ALP activity was measured and the ALP activity of the medium also contributed to mineral formation substantially. Our results highlight the diverse effects of ALP activity intrinsic to FBS on osteogenic differentiation and matrix mineralization and how FBS can determine the experimental outcomes, in particular for studies investigating matrix mineralization. Once again, the need to replace FBS with more controlled and known additives is highlighted.

Graphical abstract

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title: Alkaline phosphatase activity of serum affects osteogenic differentiation cultures


title: Localization free super-resolution microbubble velocimetry using a long short-term memory neural network

abstract: Ultrasound localization microscopy is a super-resolution imaging technique that exploits the unique characteristics of contrast microbubbles to side-step the fundamental trade-off between imaging resolution and penetration depth. However, the conventional reconstruction technique is confined to low microbubble concentrations to avoid localization and tracking errors. Several research groups have introduced sparsity- and deep learning-based approaches to overcome this constraint to extract useful vascular structural information from overlapping microbubble signals, but these solutions have not been demonstrated to produce blood flow velocity maps of the microcirculation. Here, we introduce Deep-SMV, a localization free super-resolution microbubble velocimetry technique, based on a long short-term memory neural network, that provides high imaging speed and robustness to high microbubble concentrations, and directly outputs blood velocity measurements at a super-resolution. Deep-SMV is trained efficiently using microbubble flow simulation on real in vivo vascular data and demonstrates real-time velocity map reconstruction suitable for functional vascular imaging at super-resolution. The technique is successfully applied to a wide variety of imaging scenarios, include flow channel phantoms, chicken embryo chorioallantoic membranes, and mouse kidney, tumor, and brain imaging.


title: Training data composition affects performance of protein structure analysis algorithms

abstract: The three-dimensional structures of proteins are crucial for understanding their molecular mechanisms and interactions. Machine learning algorithms that are able to learn accurate representations of protein structures are therefore poised to play a key role in protein engineering and drug development. The accuracy of such models in deployment is directly influenced by training data quality. The use of different experimental methods for protein structure determination may introduce bias into the training data. In this work, we evaluate the magnitude of this effect across three distinct tasks: estimation of model accuracy, protein sequence design, and catalytic residue prediction. Most protein structures are derived from X-ray crystallography, nuclear magnetic resonance (NMR), or cryo-electron microscopy (cryo-EM); we trained each model on datasets consisting of either all three structure types or of only X-ray data. We find that across these tasks, models consistently perform worse on test sets derived from NMR and cryo-EM than they do on test sets of structures derived from X-ray crystallography, but that the difference can be mitigated when NMR and cryo-EM structures are included in the training set. Importantly, we show that including all three types of structures in the training set does not degrade test performance on X-ray structures, and in some cases even increases it. Finally, we examine the relationship between model performance and the biophysical properties of each method, and recommend that the biochemistry of the task of interest should be considered when composing training sets.

category: bioinformatics


title: A multi-objective based clustering for identifying clonally-related sequences from high-throughput B cell repertoire data

abstract: The adaptive B cell response is driven by the expansion, somatic hypermutation, and selection of B cell clones. A high number of clones in a B cell population indicates a highly diverse repertoire, while clonal size distribution and sequence diversity within clones can be related to antigens selective pressure. Identifying clones is fundamental to many repertoire studies, including repertoire comparisons, clonal tracking and statistical analysis. Several methods have been developed to group sequences from high-throughput B cell repertoire data. Current methods use clustering algorithms to group clonally-related sequences based on their similarities or distances. Such approaches create groups by optimizing a single objective that typically minimizes intra-clonal distances. However, optimizing several objective functions can be advantageous and boost the algorithm convergence rate. Here we propose a new method based on multi-objective clustering. Our approach requires V(D)J annotations to obtain the initial clones and iteratively applies two objective functions that optimize cohesion and separation within clones simultaneously. We show that under simulations with varied mutation rates, our method greatly improves clonal grouping as compared to other tools. When applied to experimental repertoires generated from high-throughput sequencing, its clustering results are comparable to the most performing tools. The method based on multi-objective clustering can accurately identify clone members, has fewer parameter settings and presents the lowest running time among existing tools. All these features constitute an attractive option for repertoire analysis, particularly in the clinical context to unravel the mechanisms involved in the development and evolution of B cell malignancies.


title: Integration of probabilistic functional networks without an external Gold Standard

abstract:
BackgroundProbabilistic functional integrated networks (PFINs) are designed to aid our understanding of cellular biology and can be used to generate testable hypotheses about protein function. PFINs are generally created by scoring the quality of interaction datasets against a Gold Standard dataset, usually chosen from a separate high-quality data source, prior to their integration. Use of an external Gold Standard has several drawbacks, including data redundancy, data loss and the need for identifier mapping, which can complicate the network build and impact on PFIN performance.

ResultsWe describe the development of an integration technique, ssNet, that scores and integrates both high-throughput and low-throughout data from a single source database in a consistent manner without the need for an external Gold Standard dataset. Using data from Saccharomyces cerevisiae we show that ssNet is easier and faster, overcoming the challenges of data redundancy, Gold Standard bias and ID mapping, while producing comparable performance. In addition ssNet results in less loss of data and produces a more complete network.

ConclusionsThe ssNet method allows PFINs to be built successfully from a single database, while producing comparable network performance to networks scored using an external Gold Standard source.


title: CellDepot: A unified repository for scRNA-seq data and visual exploration

abstract: CellDepot serves as an integrated web application to assist users in exploring single-cell RNA-seq (scRNA-seq) datasets and comparing the datasets among various studies through a user-friendly interface with advanced visualization and analytical tools. To begin with, it provides an efficient data management system that users can upload single cell datasets and query the database by multiple attributes such as species and cell types. In addition, the advanced query function incorporated in MySQL database system and its conditional filtering, allows users to quickly query and compare the expression of gene(s) across the datasets of interest. Moreover, by embedding the cellxgene VIP tool, CellDepot enables fast exploration of individual dataset in the manner of interactivity and scalability to gain more refined insights such as cell composition, gene expression profiles, and differentially expressed genes among cell types. In summary, the web portal allows large scale single cell data sharing, analysis and visualization for supporting decision-making, and encouraging scientists to contribute to the single-cell community in a tractable and collaborative way. Finally, CellDepot is released as open-source software to motivate crowd contribution, broad adoption, and local deployment for private data.


title: Balance of osmotic pressures determines the volume of the cell nucleus

abstract: The volume of the cell nucleus varies across cell-types and species, and is commonly thought to be determined by the size of the genome and degree of chromatin compaction. However, this notion has been challenged over the years by multiple experimental evidence. Here, we consider the physical condition of mechanical force balance as a determining condition of the nuclear volume and use quantitative, order-of-magnitude analysis to estimate the forces from different sources of nuclear and cellular pressure. Our estimates suggest that the dominant pressure within the nucleus and cytoplasm originates from the osmotic pressure of proteins and RNA molecules that are localized to the nucleus or cytoplasm by out-of-equilibrium, active nucleocytoplasmic transport rather than from chromatin or its associated ions. This motivates us to formulate a physical model for the ratio of the cell and nuclear volumes in which osmotic pressures of localized proteins determine the relative volumes. In accordance with unexplained observations that are century-old, our model predicts that the ratio of the cell and nuclear volumes is a constant, robust to a wide variety of biochemical and biophysical manipulations, and is changed only if gene expression or nucleocytoplasmic transport are modulated.

category: biophysics


title: Chi3l1 is a modulator of glioma stem cell states and a therapeutic vulnerability for glioblastoma

abstract: Chi3l1 (Chitinase 3-like 1) is a secreted protein highly expressed in glioblastoma. Here, we show that exposure of glioma stem cells (GSCs) to Chi3l1 reduces the CD133+/SOX2+ cells and increases the CD44+/Chi3l1+ cells. Chi3l1 binds to CD44 and induces phosphorylation and nuclear translocation of beta-catenin, Akt and STAT3. Single cell RNA-seq and RNA velocity following incubation of GSCs with Chi3l1 show significant changes in GSC state dynamics driving GSCs towards a mesenchymal expression profile and reducing transition probabilities towards terminal cellular states. ATAC-seq reveals that Chi3l1 increases accessibility of promoters containing MAZ transcription factor footprint. Inhibition of MAZ directly regulates genes with highest expression in cellular clusters exhibiting significant cell state transitions. Finally, targeting Chi3l1 in vivo with a blocking antibody, resets the transcriptomic profile of glioblastoma and inhibits tumor growth. Our work implicates Chi3l1 as modulator of GSC cellular states and demonstrates pre-clinical efficacy of anti-Chi3l1 antibody treatment.

category: cancer biology


title: Combined PI3K and MLL inhibition synergizes in models of HR+ breast cancer

abstract:
The high frequency of aberrant PI3K pathway activation in hormone receptor-positive (HR+) breast cancer has led to the development, clinical testing, and approval of the p110-selective PI3K inhibitor alpelisib. The limited clinical efficacy of alpelisib and other PI3K inhibitors is partially attributed to the functional antagonism between PI3K and estrogen receptor (ER) signaling, which is mitigated via combined PI3K inhibition and endocrine therapy. We and others have previously demonstrated a chromatin-associated mechanism by which PI3K supports cancer development and antagonizes ER signaling through the modulation of H3K4 methylation. Here we show that inhibition of the H3K4 histone methyltransferase MLL1 in combination with PI3K inhibition impairs HR+ breast cancer clonogenicity and cell proliferation. While combined PI3K/MLL1 inhibition reduces AKT effector signaling and H3K4 methylation, MLL1 inhibition increases PI3K effector signaling and upregulates the expression of receptor tyrosine kinase signaling cascades upstream of and including AKT. These data reveal a feedback loop between MLL1 and AKT in which MLL1 inhibition reactivates AKT. We additionally show that combined PI3K and MLL1 inhibition synergizes to cause cell death in in vitro and in vivo models of HR+ breast cancer, which is enhanced by the additional genetic ablation of the H3K4 methyltransferase and AKT target MLL4. Together, our data provide evidence of a feedback mechanism connecting histone methylation with AKT and may support the preclinical development and testing of pan-MLL inhibitors.

Significance StatementPharmacological inhibition of PI3K provides limited efficacy in PIK3CA-mutated, HR+ breast cancers. Here the authors leverage PI3K/AKT-driven chromatin modification to identify MLL histone methyltransferases as a therapeutic target. Dual PI3K and MLL inhibition synergize to reduce clonogenicity and cell proliferation while enhancing apoptosis in in vitro models, and induces tumor regression in xenograft models of PI3K-activated, HR+ breast cancer. Furthermore, MLL1 inhibition reveals a feedback loop leading to AKT hyperactivation, which is relieved with combined PI3K/MLL inhibition. These findings demonstrate the utility of MLL inhibitors for the treatment of some solid cancers, as patients with HR+ breast cancer characterized by PIK3CA mutation may derive clinical benefit from combined PI3K/MLL inhibition.


title: Targeting MLL methyltransferases enhance the anti-tumor effects of PI3K inhibition in hormone receptor-positive breast cancer


title: EWS/FLI mediated reprogramming of 3D chromatin promotes an altered transcriptional state in Ewing sarcoma

abstract: Ewing sarcoma is a prototypical fusion transcription factor-associated pediatric cancer that expresses EWS/FLI or highly related fusions. EWS/FLI dysregulates transcription to induce and maintain sarcomagenesis, but the mechanisms utilized are not fully understood. We therefore sought to define the global effects of EWS/FLI on chromatin conformation and transcription in Ewing sarcoma. We found that EWS/FLI (and EWS/ERG) genomic localization is largely conserved across multiple patient-derived Ewing sarcoma cell lines. EWS/FLI binding is primarily associated with compartment activation, establishment of topologically-associated domain (TAD) boundaries, enhancer-promoter looping that involve both intra- and inter-TAD interactions, and gene activation. Importantly, local chromatin features provide the basis for transcriptional heterogeneity in regulation of direct EWS/FLI target genes across different Ewing sarcoma cell lines. These data demonstrate a key role of EWS/FLI in mediating genome-wide changes in chromatin configuration and support the notion that fusion transcription factors serve as master regulators through three-dimensional reprogramming of chromatin.


title: Synthetic glabridin derivatives mitigate steatohepatitis in a diet-induced biopsy-confirmed non-alcoholic steatosis hepatitis mouse model through paraoxonase-2

abstract: Limited therapeutic agents have been developed for non-alcoholic steatohepatitis (NASH), a common immunometabolic disease that can progress to hepatic cirrhosis and cancer. Glabridin and its derivatives are potential therapeutics for some metabolic diseases. However, the therapeutic effects of glabridin and its derivatives on NASH and their biological functions are unclear. This study demonstrated the role of synthetic glabridin derivatives (SGDs) in alleviating hepatic steatosis and inflammation in a biopsy-confirmed rodent NASH model. SGDs exerted therapeutic effects by activating autophagy and the antioxidant defense system, which mitigate NASH pathogenesis. The cellular target of HSG4112, an SGD, was paraoxonase 2. These findings will enable the development of novel therapeutics for NASH in the future.

category: pharmacology and toxicology


title: Mechanical loading and hyperosmolarity as a daily resetting cue for skeletal circadian clocks

abstract:
Daily rhythms in mammalian behaviour and physiology are generated by a multi-oscillator circadian system entrained through environmental cues (e.g. light). Presence of niche-dependent physiological time cues has been proposed, allowing local tissues flexibility of phase adjustment. However, to date, such stimuli have remained elusive. Here we show that cycles of mechanical loading and osmotic stimuli within physiological range drive rhythmic expression of clock genes and reset clock phase and amplitude in cartilage and intervertebral disc tissues. Hyperosmolarity (not hypo-osmolarity) resets clocks in young and ageing skeletal tissues through mTORC2-AKT-GSK3{beta} pathway, leading to genome-wide induction of rhythmic genes. These results advocate diurnal patterns of mechanical loading and consequent daily surges in osmolarity as a bona fide tissue niche-specific time cue to maintain skeletal circadian rhythms in sync.

One-Sentence SummaryCircadian clocks in aneural skeletal tissues sense the passage of time through rhythmic patterns of loading and osmolarity

category: physiology


title: Functional characterisation of single nucleotide variants of the psychiatric risk gene cacna1c in the zebrafish

abstract: Several genome-wide association studies have associated CACNA1C variants with psychiatric disorders. The molecular mechanisms involved are poorly understood. Taking advantage of the zebrafish larva as a model, we investigated how two different mutations in cacna1c - sa10930 (nonsense mutation) and sa15296 (splice site mutation), affect neuronal function. We characterized changes in cacna1c mRNA, neurotransmitter levels and behaviour, as well as whole-brain activity using single electrode local field potential recordings. Both point mutations resulted in a significant reduction in cacna1c mRNA, as well as social behaviour and prepulse inhibition deficits. Whereas sa15296 mutants displayed abnormal locomotor and open-field behaviour, we observed normal behaviour in the sa10930 mutants. Brain recordings from both mutants had lower spectral power while sa15296 displayed significant seizure-like activity. Finally, sa10930 homozygotes showed increased dopamine and serotonin levels, decreased gamma-aminobutyric acid (GABA) levels, and unchanged glutamate levels while homozygous sa15296 larvae showed increased levels of serotonin and glutamate, and unaffected levels of GABA and dopamine. Our work provides new insights into the functional role of CACNA1C in behavioural, electrophysiological and biochemical traits linked to psychiatric disorders. We show a functional role for the non-coding mutation (sa15296) in the cacna1c in vivo animal model. Consistent with existing hypotheses, our data suggest that disruption of gene expression, neurotransmission, and cortical excitability are involved in CACNA1C-related mechanisms of psychiatric disorders.

category: neuroscience


title: Pharyngeal timing and particle transport defects in Caenorhabditis elegans feeding mutants

abstract: The nematode C. elegans uses rhythmic muscle contractions and relaxations called pumps to filter, transport, and crush food particles. A number of feeding mutants have been identified, including those with slow pharyngeal pumping rate, weak muscle contraction, defective muscle relaxation, and defective grinding of bacteria. Many aspects of these pharyngeal behavioral defects and how they affect pharyngeal function are not well understood. For example, the behavioral deficits underlying inefficient particle transport in  slippery mutants have been unclear. Here we use high speed video microscopy to describe pharyngeal pumping behaviors and particle transport in wild-type animals and in feeding mutants. Different  slippery mutants exhibit distinct defects including weak isthmus contraction, failure to trap particles in the anterior isthmus, and abnormal timing of contraction and relaxation in pharyngeal compartments. Our results show that multiple deficits in pharyngeal timing or contraction can cause defects in particle transport.


title: Topographic connectivity and cellular profiling reveal detailed input pathways and functionally distinct cell types in the subthalamic nucleus

abstract: The subthalamic nucleus (STN) controls psychomotor activity and is an efficient therapeutic deep brain stimulation target in Parkinsons disease patients. Despite evidence indicating position-dependent therapeutic effects and distinct functions within the STN, input circuit and cellular profile in the STN remain largely unclear. Using advanced neuroanatomical techniques, we constructed a comprehensive connectivity map of the indirect and hyperdirect pathways in both the mouse and human STN. Our detailed circuit- and cellular-level connectivity revealed a topographically graded organization with three convergent types of indirect and hyperdirect-pathways. Furthermore, we identified two functional types of glutamatergic STN neurons (parvalbumin, PV +/- neurons) segregated with a topographical distribution. Glutamatergic PV+ STN neurons contribute to burst firing. We confirmed synaptic connectivity from indirect and hyperdirect pathways to both PV+ and PV-. These data suggest a complex interplay of information integration within the basal ganglia underlying coordinated movement control and therapeutic effects.


title: A Biophysical Counting Mechanism for Keeping Time

abstract: The ability to estimate and produce appropriately timed responses is central to many behaviors including speaking, dancing, and playing a musical instrument. A classical framework for estimating or producing a time interval is the pacemaker-accumulator model in which pulses of a pacemaker are counted and compared to a stored representation. However, the neural mechanisms for how these pulses are counted remains an open question. The presence of noise and stochasticity further complicate the picture. We present a biophysical model of how to keep count of a pacemaker in the presence of various forms of stochasticity using a system of bistable Wilson-Cowan units asymmetrically connected in a one-dimensional array; all units receive the same input pulses from a central clock but only one unit is active at any point in time. With each pulse from the clock, the position of the activated unit changes thereby encoding the total number of pulses emitted by the clock. This neural architecture maps the counting problem into the spatial domain, which in turn translates count to a time estimate. We further extend the model to a hierarchical structure to be able to robustly achieve higher counts.


title: Molecular structure and conformation of stereocilia tip-links elucidated by cryo-electron tomography

abstract: Mechanosensory transduction (MT), the conversion of mechanical stimuli into electrical signals, underpins hearing and balance and is carried out within hair cells in the inner ear. Hair cells harbor actin-filled stereocilia, arranged in rows of descending heights, where the tips of stereocilia are connected to their taller neighbors by a filament composed of protocadherin 15 (PCDH15) and cadherin 23 (CDH23), deemed the  tip-link. Tension exerted on the tip-link opens an ion channel at the tip of the shorter stereocilia, thus converting mechanical force into an electrical signal. While biochemical and structural studies have provided insights into the molecular composition and structure of isolated portions of the tip-link, the architecture, location and conformational states of intact tip-links, on stereocilia, remains unknown. Here we report in situ cryo-electron microscopy imaging of the tip-link in mouse stereocilia. We observe individual PCDH15 molecules at the tip and shaft of stereocilia and determine their stoichiometry, conformational heterogeneity, and their complexes with CDH23. The PCDH15/CDH23 complexes occur in clusters, frequently with more than one copy of PCDH15 at the tip of stereocilia, suggesting that tip-links might consist of more than one copy of the PCDH15/CDH23 heterotetramer and by extension, might include multiple MT complexes.


title: Cortical sensorimotor activity in the execution and suppression of discrete and rhythmic movements

abstract: Although the engagement of sensorimotor cortices in movement is well documented, the functional relevance of brain activity patterns remains ambiguous. Especially, the cortical engagement specific to the pre-, within-, and post-movement periods is poorly understood. The present study addressed this issue by examining sensorimotor EEG activity during the performance as well as STOP-signal cued suppression of movements pertaining to two distinct classes, namely, discrete vs. ongoing rhythmic movements. Our findings indicate that the lateralized readiness potential (LRP), which is classically used as a marker of pre-movement processing, indexes multiple pre- and in-movement-related brain dynamics in a movement-class dependent fashion. In- and post-movement event-related (de)synchronization (ERD/ERS) observed in the Mu (8-13 Hz) and Beta (15-30 Hz) frequency ranges were associated with estimated brain sources in both motor and somatosensory cortical areas. Notwithstanding, Beta ERS occurred earlier following cancelled than actually performed movements. In contrast, Mu power did not vary. Whereas Beta power may reflect the evaluation of the sensory predicted outcome, Mu power might engage in linking perception to action. Additionally, the rhythmic movement forced stop (only) showed a post-movement Mu/Beta rebound, which might reflect an active "clearing-out" of the motor plan and its feedback-based online control. Overall, the present study supports the notion that sensorimotor EEG modulations are key markers to investigate control or executive processes, here initiation and inhibition, which are exerted when performing distinct movement classes.


title: An Approach to Measuring Protein Turnover in Human Induced Pluripotent Stem Cell Organoids by Mass Spectrometry

abstract: Patient-derived organoids from induced pluripotent stem cells have emerged as a model for studying human diseases beyond conventional two-dimensional (2D) cell culture. Briefly, these three-dimensional organoids are highly complex, capable of self-organizing, recapitulate cellular architecture, and have the potential to model diseases in complex organs, such as the brain. For example, the hallmark of Parkinsons disease - proteostatic dysfunction leading to the selective death of neurons in the substantia nigra - present a subtle distinction in cell type specificity that is simply lost in 2D cell culture models. As such, the development of robust methods to study global proteostasis and protein turnover in organoids will remain a critical need as organoid models evolve. To solve this problem, we have designed a workflow to extract proteins from organoids and measure global protein turnover using mass spectrometry and stable isotope labeling using amino acids in cell culture (SILAC). This allowed us to measure the turnover rates of 844 proteins and compare protein turnover to previously reported data in primary cell cultures and in vivo models. Taken together, this method will facilitate the study of proteostasis in organoid models of human disease and will provide an analytical and statistical framework to measure protein turnover in organoids of all cell types.


title: Small, correlated changes in synaptic connectivity may facilitate rapid motor learning

abstract: Animals can rapidly adapt their movements to external perturbations. This adaptation is paralleled by changes in single neuron activity in the motor cortices. Behavioural and neural recording studies suggest that when animals learn to counteract a visuomotor perturbation, these changes originate from altered inputs to the motor cortices rather than from changes in local connectivity, as neural covariance is largely preserved during adaptation. Since measuring synaptic changes in vivo remains very challenging, we used a modular recurrent network model to compare the expected neural activity changes following learning through altered inputs (Hinput) and learning through local connectivity changes (Hlocal). Learning under Hinput produced small changes in neural activity and largely preserved the neural covariance, in good agreement with neural recordings in monkeys. Surprisingly given the presumed dependence of stable neural covariance on preserved circuit connectivity, Hlocal led to only slightly larger changes in neural activity and covariance compared to Hinput. This similarity is due to Hlocal only requiring small, correlated connectivity changes to counteract the perturbation, which provided the network with significant robustness against simulated synaptic noise. Simulations of tasks that impose increasingly larger behavioural changes revealed a growing difference between Hinput and Hlocal, which could be exploited when designing future experiments.


title: Prevalence, persistence, and genetics of antibody responses to protein toxins and virulence factors

abstract: Microbial exposures are crucial environmental factors that impact healthspan by sculpting the immune system and microbiota. Antibody profiling via programmable Phage ImmunoPrecipitation Sequencing (PhIP-Seq) provides a high-throughput, costeffective approach for multiplexed detection of exposure and response to thousands of microbial protein products. Here we designed and constructed a library of 95,601 56 amino acid peptide tiles spanning a subset of environmental proteins more likely to be associated with immune responses: those with "toxin" or "virulence factor" keyword annotations. PhIP-Seq was used to profile the circulating antibodies of ~1,000 individuals against this "ToxScan" library of 14,430 toxins and virulence factors from 1,312 genera of organisms. In addition to a detailed analysis of six commonly encountered human commensals and pathogens, we study the age-dependent stability of the ToxScan profile and use a genome-wide association study (GWAS) to find that the MHC-II locus modulates the selection of bacterial epitopes. We detect previously described anti-flagellin antibody responses in a Crohns disease cohort and identify a novel association between anti-flagellin antibodies and juvenile dermatomyositis (JDM). PhIP-Seq with the ToxScan library provides a new window into exposure and immune responses to environmental protein toxins and virulence factors, which can be used to study human health and disease at cohort scale.

category: immunology


title: Long-term analysis of pertussis vaccine immunity uncovers a memory B cell response to whole cell pertussis immunization that is absent from acellular immunized mice

abstract: Over two decades ago acellular pertussis vaccines (aP) replaced whole cell pertussis vaccines (wP) in several countries. Since then, a resurgence in pertussis has been observed, which is hypothesized to be linked to waning immunity. To better understand why waning immunity occurs, we developed a long-term outbred CD1 mouse model to conduct the longest murine pertussis vaccine studies to date, spanning out to 532 days post primary immunization. Vaccine-induced memory results from follicular responses and germinal center formation; therefore, cell populations and cytokines involved with memory were measured alongside protection from challenge. Both aP and wP immunization elicit protection from intranasal challenge and generation of pertussis specific antibody responses in mice. Responses to wP vaccination were characterized by a significant increase in T follicular helper cells in the draining lymph nodes and CXCL13 levels in sera compared to aP mice. In addition, a population of B. pertussis+ memory B cells was found to be unique to wP vaccinated mice. This population peaked post-boost, and was measurable out to day 365 post-vaccination. Anti-B. pertussis and anti-pertussis toxoid antibody secreting cells increased one day after boost and remained high at day 532. The data suggest that follicular responses, and in particular CXCL13 levels in sera, should be monitored in pre-clinical and clinical studies for the development of the next-generation pertussis vaccines.


title: Synthetic sphingolipid analogs and anti-cancer drugs synergistically induced human colon and pancreatic cancer cell death through inhibiting ceramide metabolism

abstract: Ceramide metabolism is a potential target for anti-cancer therapy. Studies show that chemotherapeutic agents can induce apoptosis and it is mediated by ceramide. Synthesized sphingolipid analogs can induce cell death in human lymphocytes and leukemia cells. By screening a group of synthetic sphingolipid analogs, we found that low concentrations of AD2750 and AD2646 induced cell death in human cancer cells by preventing ceramide from converting to sphingomyelin, individually or in combination with commercial cancer drugs. The combination of low concentrations of Taxol and AD2750 or AD2646 significantly increased cell death on human colon cancer cells (HT29). Co-administering low concentrations of Doxorubicin with AD2750 or AD2646 elevated cellular toxicity on human pancreatic cancer cells (CRL1687). This synergistic effect is related to the elevated cellular ceramide. Combining AD2750 or AD2646 with chemotherapy drugs can be used to manipulate ceramide and sphingomyelin metabolism, potentially to affect the growth of human cancer cells and increase the effectiveness of anti-cancer drugs on killing cancer cells.

category: cell biology


title: Single-cell transcriptomics reveals the identity and regulators of human mast cell progenitors

abstract:
Mast cell accumulation is a hallmark of a number of diseases including allergic asthma and systemic mastocytosis. IgE-mediated crosslinking of the Fc{varepsilon}RI receptors causes mast cell activation and contributes to disease pathogenesis. The mast cell lineage is one of the least studied among the hematopoietic cell lineages and there are still controversies about the identity of the mast cell progenitor, i.e., whether Fc{varepsilon}RI expression appears during the hematopoietic progenitor stage or in maturing mast cells. Here, we used single-cell transcriptomics to reveal a temporal association between the appearance of Fc{varepsilon}RI and the mast cell gene signature in CD34+ hematopoietic progenitors. In agreement with these data, the Fc{varepsilon}RI+ hematopoietic progenitors formed morphologically, phenotypically and functionally mature mast cells in long-term culture assays. Single-cell transcriptomics analysis further revealed the expression patterns of prospective cytokine receptors regulating mast cell progenitor development. Culture assays showed that IL-3 and IL-5 promoted disparate effects on progenitor cell proliferation and survival, respectively, whereas IL-33 caused robust Fc{varepsilon}RI downregulation. Taken together, we have demonstrated that Fc{varepsilon}RI appears during the hematopoietic progenitor stage of mast cell differentiation and that external stimuli may regulate the Fc{varepsilon}RI expression. Thus, the results resolve the controversy regarding the appearance of Fc{varepsilon}RI during mast cell development.

One-sentence summarySingle-cell analysis of human hematopoiesis uncovers the stage at which Fc{varepsilon}RI appears during mast cell differentiation and reveals disparate effects of IL-3, IL-5 and IL-33 on mast cell progenitor proliferation, survival, and suppression of Fc{varepsilon}RI expression.


title: JADE family proteins regulate proteasome abundance and activity

abstract: JADE family proteins (JADE1/2/3) have been implicated in diverse cellular functions and signaling pathways ranging from WNT signaling and cell cycle control to cell death and complex transcriptional regulation through histone acetyl-transferase complexes. JADE proteins show a high degree of sequence similarity and share two PHD zinc finger domains. JADE1 interacts with cilia-associated proteins and has been implicated in cilia-related genetic disorders with kidney phenotypes. However, the function of the widely expressed JADE proteins at the molecular level is still elusive. Here we show that JADE proteins regulate proteasome abundance and activity. Using kidney cells as a model, we demonstrate that loss of either JADE protein resulted in increased expression of almost all components of the 26S proteasome. Regulation occurred at the post-translational level and was not the consequence of transcriptional activation. Consistent with a role for JADE proteins in regulating overall proteasomal abundance, proteasomal activity was elevated in Jade-deficient cells, while exogenous expression of JADE1/2/3 decreased the level of proteasome activity. Coimmunoprecipitation experiments confirmed the interaction of proteasomal subunits with Jade1 suggesting a direct role of JADE proteins in regulating turnover, stability and abundance of the 26s proteasome. These data may now explain the plethora of cellular roles that have been attributed to JADE proteins.


title: INF2-mediated actin filament reorganization confers intrinsic resilience to neuronal ischemic injury

abstract: During early stages of ischemic brain injury, glutamate receptor hyperactivation mediates neuronal death via osmotic cell swelling. Here we show that ischemia and excess NMDA receptor activation - conditions that trigger neuronal swelling -- cause actin filaments to undergo a rapid and extensive reorganization within the somatodendritic compartment. Normally, F-actin is concentrated within dendritic spines, with relatively little F-actin in the dendrite shaft. However, beginning <5 min after incubation of neurons with NMDA, F-actin depolymerizes within dendritic spines and polymerizes into long, stable filament bundles within the dendrite shaft and soma. A similar "actinification" of the somatodendritic compartment occurs after oxygen/glucose deprivation in vitro, and in mouse brain after photothrombotic stroke in vivo. Following transient, sub-lethal NMDA exposure these actin changes spontaneously reverse within 1-2 hours. A combination of Na+, Cl-, water, and Ca2+ entry are all necessary, but not individually sufficient, for induction of actinification. Spine F-actin depolymerization is also required. Actinification is driven by activation of the F-actin polymerization factor inverted formin-2 (INF2). Silencing of INF2 renders neurons more vulnerable to NMDA-induced membrane leakage and cell death, and formin inhibition markedly increases ischemic infarct severity in vivo. These results show that ischemia-induced actin filament reorganization within the dendritic compartment is an intrinsic pro-survival response that protects neurons from death induced by swelling.


title: Laminin switches terminal differentiation fate of human trophoblast stem cells under chemically defined culture conditions.

abstract:
Human trophoblast stem cells (hTSCs) have emerged as a powerful tool to model early placental development in vitro. Analogous to the epithelial cytotrophoblast in the placenta, hTSCs can differentiate into cells of the extravillous trophoblast (EVT) lineage or the multinucleate syncytiotrophoblast (STB). Here we present a chemically defined culture system for STB and EVT differentiation of hTSCs. Notably, in contrast to current approaches, we do not utilize transforming growth factor-beta inhibitors or a passage step for EVT differentiation, or forskolin for STB formation. Strikingly, under these conditions, presence of a single additional extracellular cue - lam-inin-1 - switched the terminal differentiation of hTSCs from STB to the EVT lineage. Activation of the sphingosine-1 receptor 3 receptor (S1PR3) using a chemical agonist could drive EVT differentiation of hTSCs in the absence of exogenous laminin, albeit less efficiently. To illustrate the utility of a chemically defined culture system for mechanistic studies, we examined the role of protein kinase C (PKC) signaling during hTSC differentiation to the EVT lineage. Inhibition of PKC/{beta} signaling significantly reduced HLA-G expression and the formation of HLA-G+ mesen-chymal EVTs during hTSC differentiation mediated by laminin exposure; however, it did not prevent commitment to the EVT lineage or STB differentiation. The chemically defined culture system for hTSC differentiation established herein facilitates quantitative analysis of heterogeneity that arises during hTSC differentiation, and will enable mechanistic studies in vitro.

SignificanceDespite its importance to a healthy pregnancy, early human placental development remains poorly understood. Mechanistic studies are impeded by restrictions on research with human embryos and fetal tissues, and significant differences in placentation between humans and commonly used animal models. In this context, human trophoblast stem cells (hTSCs) have emerged as attractive in vitro models for the epithelial cytotrophoblast of the early gestation human placenta. Here we describe chemically defined culture conditions for differentiation of hTSCs to the two major differentiated cell types - extravillous trophoblast and syncytiotrophoblast. These culture conditions enable in vitro studies to reveal molecular mechanisms regulating hTSC differentiation.

category: developmental biology


title: Maintenance of community function through compensation breaks down over time in a desert rodent community

abstract: Understanding the ecological processes that maintain community function in systems experiencing species loss, and how these processes change over time, is key to understanding the relationship between community structure and function and predicting how communities may respond to perturbations in the Anthropocene. Using a 30-year experiment on desert rodents, we show that the impact of species loss on community-level energy use has changed repeatedly and dramatically over time, due to 1) the addition of new species to the community, and 2) a reduction in functional redundancy among the same set of species. Although strong compensation, initially driven by the dispersal of functionally redundant species to the local community, occurred in this system from 1997-2010, since 2010, compensation has broken down due to decreasing functional overlap within the same set of species. Simultaneously, long-term changes in sitewide community composition due to niche complementarity have decoupled the dynamics of compensation from the overall impact of species loss on community-level energy use. Shifting, context-dependent compensatory dynamics, such as those demonstrated here, highlight the importance of explicitly long-term, metacommunity, and eco-evolutionary perspectives on the link between species-level fluctuations and community function in a changing world.

category: ecology


title: Blood variation implicates respiratory limits on elevational ranges of Andean birds

abstract: The extent to which species ranges reflect intrinsic physiological tolerances is a major, unsolved question in evolutionary ecology. To date, consensus has been hindered by the limited tractability of experimental approaches across most of the tree of life. Here, we apply a macrophysiological approach to understand how hematological traits related to oxygen transport shape elevational ranges in a tropical biodiversity hotspot. Along Andean elevational gradients, we measured traits that affect blood oxygen-carrying capacity--total and cellular hemoglobin concentration and hematocrit--for 2,355 individuals of 136 bird species. We used these data to evaluate the influence of hematological traits on elevational ranges. First, we asked whether hematological plasticity is predictive of elevational range breadth. Second, we asked whether variance in hematological traits changed as a function of distance from the midpoint of the elevational range. We found that the correlation between hematological plasticity and elevational range breadth was slightly positive, consistent with a facilitative role for plasticity in elevational range expansion. We further found reduced local variation in hematological traits near elevational range limits and at high elevations, patterns consistent with intensified natural selection, reduced effective population size, or compensatory changes in other cardiohematological traits with increasing distance from species-specific optima for oxygen availability. Our findings suggest that constraints on hematological plasticity and local genetic adaptation to oxygen availability promote the evolution of the narrow elevational ranges that underpin tropical montane biodiversity.


title: Potential of breadfruit cultivation to contribute to climate-resilient low latitude food systems

abstract: The number of people in food crisis around the world is increasing, exacerbated by the challenges of COVID-19 and a rapidly changing climate. Major crop yields are projected to decrease in low-latitude regions due to anthropogenic climate change, making tropical and sub-tropical food systems particularly vulnerable to climate shocks. Increased cultivation of breadfruit (Artocarpus altilis), often categorized as a neglected and underutilized species (NUS), has been suggested as an agricultural adaptation pathway for food insecure tropical and subtropical regions, due to its potential to enhance climate resilience and overall sustainability of low-latitude agricultural systems. To better understand breadfruits cultivation suitability and geographic range in current and future climates, we employ a diverse set of observations and models to delineate the current climatically viable breadfruit range and assess the climatically viable breadfruit range in the future (2061-2080) under stabilization and high emission scenarios. We find that the area of suitable breadfruit range within the tropics and subtropics is projected to decrease ~4.4% in the stabilization scenario and ~4.5% in the high emission scenario. In Southeast Asia and the Pacific Islands, yield quality and consistency show minimal decreases under the high emission scenario, with increases in total suitable area under both scenarios. In contrast, in Latin America and the Caribbean, the current range of breadfruit suitability is projected to contract ~10.1-11.5% (stabilization-high emission). Present and future model suitability outputs suggest that opportunities to successfully expand breadfruit cultivation over the next several decades exist in sub-Saharan Africa, where food insecurity is coincidentally high. However, in all regions, high emission scenario conditions reduce the overall consistency and quality of breadfruit yields compared to the stabilization scenario. Our results have the potential to inform global food security adaptation planning and highlight breadfruit as an ideal NUS to incorporate in food security adaptation strategies in a changing climate.


title: Ecological Stability Emerges at the Level of Strains in the Human Gut Microbiome

abstract: The human gut microbiome is a complex community that harbors substantial ecological diversity at the species level, as well as at the strain level within species. In healthy hosts, species abundance fluctuations in the microbiome community are thought to be stable, and these fluctuations can be described by macroecological laws. However, it is less clear how strain abundances change over time. An open question is whether individual strains behave like species themselves, exhibiting stability and following the macroecological relationships known to hold at the species level, or whether strains have different dynamics, perhaps due to the relatively close phylogenetic relatedness of co-colonizing lineages. In this study, we sought to characterize the typical strain-level dynamics of the healthy human gut microbiome on timescales ranging from days to years. We show that genetic diversity within almost all species is stationary, tending towards a long-term typical value within hosts over time scales of several years, despite fluctuations on shorter timescales. Moreover, the abundance fluctuations of strains can be sufficiently described by a stochastic logistic model (SLM) - a model previously used to describe abundance fluctuations among species around a fixed carrying capacity - in the vast majority of cases, suggesting that strains are dynamically stable. Lastly, we find that strain abundances follow the same macroecological laws known to hold at the species level. Together, our results suggest that macroecological properties of the human gut microbiome, including its stability, emerge at the level of strains.


title: Ruminant-specific retrotransposons shape regulatory evolution of bovine immunity

abstract: Cattle are an important livestock species, and mapping the genomic architecture of agriculturally relevant traits such as disease susceptibility is a major challenge in the bovine research community. Lineage-specific transposable elements (TEs) are increasingly recognized to contribute to gene regulatory evolution and variation, but this possibility has been largely unexplored in ruminant genomes. We conducted epigenomic profiling of the type II interferon (IFN) response in bovine cells, and found thousands of ruminant-specific TEs including MER41_BT and Bov-A2 elements predicted to act as IFN-inducible enhancer elements. CRISPR knockout experiments in bovine cells established that critical immune factors including IFNAR2 and IL2RB are transcriptionally regulated by TE-derived enhancers. Finally, population genomic analysis of 38 individuals revealed that a subset of TE-derived enhancers represent polymorphic insertion sites in modern cattle. Our study reveals that lineage-specific TEs have shaped the evolution of ruminant IFN responses, and potentially continue to contribute to immune gene regulatory differences across modern breeds and individuals. Together with previous work in human cells, our findings demonstrate that lineage-specific TEs have been independently co-opted to regulate IFN-inducible gene expression in multiple species, supporting TE co-option as a recurrent mechanism driving the evolution of IFN-inducible transcriptional networks.

category: evolutionary biology


title: Host phylogeny and ecological associations best explain Wolbachia host shifts in scale insects

abstract: Wolbachia are among the most prevalent and widespread endosymbiotic bacteria on earth. Wolbachia s success in infecting an enormous number of arthropod species is attributed to two features: the range of phenotypes they induce in their hosts, and their ability to switch to new host species. Whilst much progress has been made in elucidating the phenotypes induced by Wolbachia, our understanding of Wolbachia host shifting is still very limited: we lack answers to even fundamental questions concerning Wolbachias routes of transfer and the importance of factors influencing host shifts. Here, we investigate the diversity and host-shift patterns of Wolbachia in scale insects, a group of arthropods with intimate associations with other insects that make them well-suited to studying host shifts. Using Illumina pooled amplicon sequencing of Wolbachia-infected scale insects and their direct associates we determined the identity of all Wolbachia strains, revealing that 32% of samples were multiply infected (with up to five distinct strains per species). We then fitted a Generalised Additive Mixed Model (GAMM) to our data to estimate the influence of factors such as the host phylogeny and the geographic distribution of each species on Wolbachia strain sharing among scale insect species. The model predicts no significant contribution of host geography but strong effects of host phylogeny, with high rates of Wolbachia sharing among closely related species and a sudden drop-off in sharing with increasing phylogenetic distance. We also detected the same Wolbachia strain in scale insects and several intimately associated species (ants, wasps, beetles, and flies). This indicates putative host shifts and potential routes of transfers via these associates and highlights the importance of ecological connectivity in Wolbachia host-shifting.


title: Whale shark rhodopsin adapted to its vertically wide-ranging lifestyle

abstract: Spectral tuning of visual pigments often facilitates adaptation to new environments, and it is intriguing to study the visual ecology of pelagic sharks with expanded habitats. The whale shark, which dives into the deep sea of nearly 2,000 meters besides near-surface filter-feeding, was previously shown to possesses the  blue-shifted rhodopsin (RHO). In this study, our spectroscopy of recombinant whale shark RHO mutants revealed the dominant effect of the novel spectral tuning amino acid site 94, which is implicated in congenital stationary night blindness of humans, accounting for the blue shift. Thermal decay profiling revealed the reduction of the thermal stability of whale shark RHO, as typically observed for cone opsins, which was experimentally shown to be achieved by the site 178, as well as 94. The results suggest that these two sites cooperatively enhance the visual capacity in both the deep sea and the sea surface, enabling exceptionally wide vertical migration of this species.


title: High-throughput RNA isoform sequencing using programmable cDNA concatenation

abstract: Alternative splicing is a core biological process that enables profound and essential diversification of gene function. Short-read RNA sequencing approaches fail to resolve RNA isoforms and therefore primarily enable gene expression measurements - an isoform unaware representation of the transcriptome. Conversely, full-length RNA sequencing using long-read technologies are able to capture complete transcript isoforms, but their utility is deeply constrained due to throughput limitations. Here, we introduce MAS-ISO-seq, a technique for programmably concatenating cDNAs into single molecules optimal for long-read sequencing, boosting the throughput >15 fold to nearly 40 million cDNA reads per run on the Sequel IIe sequencer. We validated unambiguous isoform assignment with MAS-ISO-seq using a synthetic RNA isoform library and applied this approach to single-cell RNA sequencing of tumor-infiltrating T cells. Results demonstrated a >30 fold boosted discovery of differentially spliced genes and robust cell clustering, as well as canonical PTPRC splicing patterns across T cell subpopulations and the concerted expression of the associated hnRNPLL splicing factor. Methods such as MAS-ISO-seq will drive discovery of novel isoforms and the transition from gene expression to transcript isoform expression analyses.

category: genetics


title: The genome and lifestage-specific transcriptomes of a plant-parasitic nematode and its host reveal susceptibility genes involved in trans-kingdom synthesis of vitamin B.

abstract: Plant-parasitic nematodes are a major, and in some cases a dominant, threat to crop production in all agricultural systems. The relative scarcity of classical resistance genes highlights a pressing need to identify new ways to develop nematode-resistant germplasm. Here, we sequence and assemble a high-quality genome of the model cyst nematode Heterodera schachtii to provide a platform for the first system-wide dual analysis of host and parasite gene expression over time, covering all major stages of the interaction. This novel approach enabled the analysis of the hologenome of the infection site, to identify metabolic pathways that were incomplete in the parasite but complemented by the host. Using a combination of bioinformatic, genetic, and biochemical approaches, we show that the highly atypical completion of vitamin B5 biosynthesis by the parasitic animal, putatively enabled by a horizontal gene transfer from a bacterium, is critically important for parasitism. Knockout of either the plant-encoded or the now nematode-encoded steps in the pathway blocks parasitism. Our experiments establish a reference for cyst nematodes, use this platform to further our fundamental understanding of the evolution of plant-parasitism by nematodes, and show that understanding congruent differential expression of metabolic pathways represents a new way to find nematode susceptibility genes, and thereby, targets for future genome editing-mediated generation of nematode-resistant crops.


title: Single-cell transcription profiles in Bloom syndrome patients link BLM deficiency with altered condensin complex expression signatures

abstract: Bloom syndrome (BS) is an autosomal recessive disease clinically characterized by primary microcephaly, growth deficiency, immunodeficiency, and predisposition to cancer. It is mainly caused by biallelic loss-of-function mutations in the BLM gene, which encodes the BLM helicase, acting in DNA replication and repair processes. Here, we describe the gene expression profiles of three BS fibroblast cell lines harboring causative, biallelic truncating mutations obtained by single-cell (sc) transcriptome analysis. We compared the scRNA transcription profiles from three BS patient cell lines to two age-matched wild-type controls and observed specific deregulation of gene sets related to the molecular processes characteristically affected in BS, such as mitosis, chromosome segregation, cell cycle regulation, and genomic instability. We also found specific upregulation of genes of the Fanconi anemia pathway, in particular FANCM, FANCD2, and FANCI, which encode known interaction partners of BLM. The significant deregulation of genes associated with inherited forms of primary microcephaly observed in our study might explain in part the molecular pathogenesis of microcephaly in BS, one of the main clinical characteristics in patients. Finally, our data provide first evidence of a novel link between BLM dysfunction and transcriptional changes in condensin complex I and II genes. Overall, our study provides novel insights into gene expression profiles in BS on a single-cell level, linking specific genes and pathways to BLM dysfunction.


title: Loss of α-actinin-3 confers protection from eccentric contraction damage in fast-twitch EDL muscles from aged mdx dystrophic mice by reducing pathological fibre branching

abstract: The common null polymorphism (R577X) in the ACTN3 gene is present in over 1.5 billion people worldwide and results in the absence of the protein -actinin-3 from the Z-discs of fast-twitch skeletal muscle fibres. We have previously reported that this polymorphism is a modifier of dystrophin deficient Duchenne Muscular Dystrophy. To investigate the mechanism underlying this we use a double knockout (dk)Actn3KO/mdx (dKO) mouse model which lacks both dystrophin and sarcomere -actinin-3. We used dKO mice and mdx dystrophic mice at 12 months (aged) to investigate the correlation between morphological changes to the fast-twitch dKO EDL and the reduction in force deficit produced by an in vitro eccentric contraction protocol. In the aged dKO mouse we found a marked reduction in fibre branching complexity that correlated with protection from eccentric contraction induced force deficit. Complex branches in the aged dKO EDL fibres (28%) were substantially reduced compared to aged mdx EDL fibres (68%) and this correlates with a graded force loss over three eccentric contractions for dKO muscles ([~]35% after first contraction, [~]66% overall) compared to an abrupt drop in mdx upon the first eccentric contraction ([~]73% after first contraction, [~]89% after three contractions). In dKO protection from eccentric contraction damage was linked with a doubling of SERCA1 pump density the EDL. We propose that the increased oxidative metabolism of fast-twitch glycolytic fibres characteristic of the null polymorphism (R577X) and increase in SR Ca2+ pump proteins reduces muscle fibre branching and decreases susceptibility to eccentric injury in the dystrophinopathies.


title: ASIP promoter variants predict sesame coat color in Shiba Inu dogs

abstract: Different patterns of coat color pigmentation in dogs are produced by a sophisticated interaction of several genes. Understanding the mechanisms underlying the diversity of coat colors and their inheritance is important for professional breeders because it helps to predict the phenotypes of the progeny. Although genetics of the main coat colors in dogs is extensively studied, there are types of coat pigmentation that are not explained yet. Recently a new model connected the variants in ASIP gene promoters with different coat colors in dogs. Here we used this model as a framework to investigate the genetics of the rare sesame coat color in Shiba Inu dogs. We determined the combination of two alleles of ASIP gene that determine sesame coat color. This finding can be used by the breeders to produce the dogs with this rare coat color pattern. We also demonstrate the incomplete dominance between the ASIP alleles involved in sesame coat formation. These results are in good agreement with the new model explaining how different levels of ASIP gene expression affect the regulation of pigment synthesis in melanocytes.


title: Genome sequencing of white-blotched river stingray (Potamotrygon leopoldi) provides novel clues for niche-adaptation and skeleton formation

abstract: The white-blotched river stingray (Potamotrygon leopoldi) is a cartilaginous fish native to the Xingu River, a tributary of the Amazon River system. It possesses a lot of unique biological features such as disc-like body shape, bizarre color pattern and living in freshwater habitat while most stingrays and their close relatives are sea dwellers. As a member of the Potamotrygonidae family, P. leopoldi bears evolutionary signification in fish phylogeny, niche adaptation and skeleton formation. In this study, we present its draft genome of 4.11 Gb comprised of 16,227 contigs and 13,238 scaffolds, which has contig N50 of 3,937 kilobases and scaffold N50 of 5,675 kilobases in size. Our analysis shows that P. leopoldi is a slow-evolving fish, diverged from elephant shark about 96 million years ago. We find that two gene families related to immune system, immunoglobulin heavy constant delta genes, and T-cell receptor alpha/delta variable genes, stand out expanded in P. leopoldi only, suggesting robustness in response to freshwater pathogens in adapting novel environments. We also identified the Hox gene clusters in P. leopoldi and discovered that seven Hox genes shared by five representative fishes are missing in P. leopoldi. The RNA-seq data from P. leopoldi and other three fish species demonstrate that fishes have a more diversified tissue expression spectrum as compared to the corresponding mammalian data. Our functional studies suggest that the lack of genes encoding vitamin D-binding protein in cartilaginous (both P. leopoldi and Callorhinchus milii) fishes could partly explain the absence of hard bone in their endoskeleton. Overall, this genome resource provides new insights into the niche-adaptation, body plan and skeleton formation of P. leopoldi as well as the genome evolution in cartilaginous fish.

category: genomics


title: A general framework for identifying rare variant combinations in complex disorders

abstract:
Genetic studies of complex disorders such as autism and intellectual disability (ID) are often based on enrichment of individual rare variants or their aggregate burden in affected individuals compared to controls. However, these studies overlook the influence of combinations of rare variants that may not be deleterious on their own due to statistical challenges resulting from rarity and combinatorial explosion when enumerating variant combinations, limiting our ability to study oligogenic basis for these disorders. We present a framework that combines the apriori algorithm and statistical inference to identify specific combinations of mutated genes associated with complex phenotypes. Our approach overcomes computational barriers and exhaustively evaluates variant combinations to identify non-additive relationships between simultaneously mutated genes. Using this approach, we analyzed 6,189 individuals with autism and identified 718 combinations significantly associated with ID, and carriers of these combinations showed lower IQ than expected in an independent cohort of 1,878 individuals. These combinations were enriched for nervous system genes such as NIN and NGF, showed complex inheritance patterns, and were depleted in unaffected siblings. We found that an affected individual can carry many oligogenic combinations, each contributing to the same phenotype or distinct phenotypes at varying effect sizes. We also used this framework to identify combinations associated with multiple comorbid phenotypes, including mutations of COL28A1 and MFSD2B for ID and schizophrenia and ABCA4, DNAH10 and MC1R for ID and anxiety/depression. Our framework identifies a key component of missing heritability and provides a novel paradigm to untangle the genetic architecture of complex disorders.

SIGNIFICANCEWhile rare mutations in single genes or their collective burden partially explain the genetic basis for complex disorders, the role of specific combinations of rare variants is not completely understood. This is because combinations of rare variants are rarer and evaluating all possible combinations would result in a combinatorial explosion, creating difficulties for statistical and computational analysis. We developed a data mining approach that overcomes these limitations to precisely quantify the influence of combinations of two or more mutated genes on a specific clinical feature or multiple co-occurring features. Our framework provides a new paradigm for dissecting the genetic causes of complex disorders and provides an impetus for its utility in clinical diagnosis.


title: Genetic control of the dynamic transcriptional response to immune stimuli and glucocorticoids at single cell resolution

abstract: Synthetic glucocorticoids, such as dexamethasone, have been used as treatment for many immune conditions, such as asthma and more recently severe COVID-19. Single cell data can capture more fine-grained details on transcriptional variability and dynamics to gain a better understanding of the molecular underpinnings of inter-individual variation in drug response. Here, we used single cell RNA-seq to study the dynamics of the transcriptional response to glucocorticoids in activated Peripheral Blood Mononuclear Cells from 96 African American children. We employed novel statistical approaches to calculate a mean-independent measure of gene expression variability and a measure of transcriptional response pseudotime. Using these approaches, we demonstrated that glucocorticoids reverse the effects of immune stimulation on both gene expression mean and variability. Our novel measure of gene expression response dynamics, based on the diagonal linear discriminant analysis, separated individual cells by response status on the basis of their transcriptional profiles and allowed us to identify different dynamic patterns of gene expression along the response pseudotime. We identified genetic variants regulating gene expression mean and variability, including treatment-specific effects, and demonstrated widespread genetic regulation of the transcriptional dynamics of the gene expression response.


title: Experience resetting in reinforcement learning facilitates exploration-exploitation transitions during a behavioral task for primates

abstract: The exploration-exploitation trade-off is a fundamental problem in re-inforcement learning. To study the neural mechanisms involved in this problem, a target search task in which exploration and exploitation phases appear alternately is useful. Monkeys well trained in this task clearly understand that they have entered the exploratory phase and quickly acquire new experiences by resetting their previous experiences. In this study, we used a simple model to show that experience resetting in the exploratory phase improves performance rather than decreasing the greediness of action selection, and we then present a neural network-type model enabling experience resetting.


title: Responses to Temperatures of Different Drosophila Species

abstract:
Temperature is a critical environmental variable that affects the distribution, survival, and reproduction of most animals. Although temperature receptors have been identified in different animals, how these receptors respond to temperatures is largely unknown. Here we use modified single-fly thermotactic assays to analyze movements and temperature preferences of nine Drosophila species. The ability/inclination to move varies among these species and at different temperatures. Importantly, different species prefer various ranges of temperatures. While wild-type D. melanogaster flies avoid the warm temperature in the warm avoidance assay and the cool temperature in the cool avoidance assay, D. bipectinata and D. yakuba avoid neither warm nor cool temperatures and D. biarmipes and D. mojavensis do not avoid the warm temperature in the warm avoidance assay. These results demonstrate that Drosophila species have different mobilities and temperature preferences, thereby benefiting the research on molecular mechanisms of temperature responsiveness.

Summary statementThe ability to move and the preference for temperatures vary among fly species when flies are exposed to steep temperature gradients.


title: A hymenopteran odorant alerts flies to bury eggs

abstract: Ants are ubiquitous and consume insects at all life stages, presumably creating a strong selective pressure for ant avoidance behaviors across insects. The insect egg stage can be especially defenseless against predation given that eggs are usually immobile and unguarded, suggesting insect mothers may have evolved oviposition strategies to minimize the ant predation risk to their offspring. Given the lack of parental care in most insects, these oviposition strategies would likely be innate rather than learned, since insect mothers are not usually present to assess predation of their eggs. Here, we use the vinegar fly Drosophila melanogaster as a model system for examining parental defensive responses to ant presence. Flies usually lay eggs partially inserted into the food substrate, although some are laid on top of the food and a few are inserted deeply into the food. We found that exposure to ants significantly alters fly oviposition depth: the proportion of eggs on the food surface decreased while the proportion of buried eggs increased. Buried eggs survive ant foraging bouts better than surface eggs, showing that this oviposition depth behavior is adaptive. This induced behavior is conserved across the genus Drosophila and is dependent on the fly olfactory system: anosmic mutant flies fail to bury their eggs in the presence of ants, and ant odor extracts are sufficient to induce egg burying. To further delineate the ant lineages to which flies respond, we exposed flies to the odors from numerous species of ants and other insects. Surprisingly, flies buried their eggs in response to the odors of nearly all hymenopterans tested, including hymenopteran groups that flies rarely interact with in nature like bees and paper wasps. Our data suggest that hymenopterans possess a conserved and ancient odorant, and that drosophilids evolved a mechanism for sensing this odorant early in their evolution as a means of protecting their offspring from ant predation. This study sheds light on the ecology and mechanisms underlying a sscommon biotic interaction in nature, that between insect parents ands the ants that would consume their offspring.


title: Metformin modulates microbiota-derived inosine and ameliorates methamphetamine-induced anxiety and depression-like withdrawal symptoms in mice

abstract:
ObjectiveMetformin exhibits therapeutic potential in behavioural deficits induced by methamphetamine (METH) in rats. Emerging studies suggest gut microbiota may impact psychiatric symptoms, but there is no direct evidence supporting metformins participation in the pathophysiology of withdrawal symptoms via modulation of gut microbiota.

MehodsIn order to define the functional contributions by gut microbiota and metformin to the behavioural deficits during METH withdrawal, we utilized a combination of fecal microbiota transplantation (FMT), high-throughput sequencing, and untargeted metabolomics technologies.

ResultsFirst, METH addicts exhibited higher  diversity and distinct microbial structures compared to heathy controls. In particular, the relative abundance of Rikenellaceae was positively correlated with the severity of anxiety and depression. Second, both human-to-mouse and mouse-to-mouse FMTs confirmed that METH-altered-microbiota transplantation is sufficient to promote anxiety and depression-like behaviours in recipient germ-free mice, and these behavioural disturbances could be ameliorated by metformin. In-depth analysis revealed that METH significantly altered the bacterial composition and structure as well as relative abundance of several bacterial taxa and metabolites, including Rikenellaceae and inosine, respectively, whereas add-on metformin could remodel these alterations. Finally, the inosine complementation successfully restored METH-induced anxiety and depression-like behaviours in mice.

DiscussionThis study demonstrates that METH withdrawal-induced anxiety and depression-like behaviours are convertible and transmissible via gut microbiota in a mouse model. The therapeutic effects of metformin on psychiatric manifestations are associated with microbiota-derived metabolites, highlighting the role of the gut microbiota in substance use disorders and the pathophysiology of withdrawal symptoms.

Study HighlightsO_ST_ABSWhat is known?C_ST_ABSO_LIThere are no targeted therapies for substance withdrawal syndrome, but there is considerable evidence that withdrawal-associated psychiatric manifestations contribute to the poor adherence to rehabilitation treatment as well as the relapse rates.
C_LIO_LIMetformin has shown its therapeutic potential against METH-induced neurobehavioural changes and neurodegeneration in rats through CREB/BDNF and Akt/GSK3 signaling pathways in the anxiety-related brain nuclei.
C_LI

What is new here?O_LIMETH withdrawal-induced anxiety and depression-like behaviours are convertible and transmissible via gut microbiota in a mouse model.
C_LIO_LIThe therapeutic effects of metformin on psychiatric manifestations are associated with microbiota derived metabolites.
C_LIO_LIInosine complementation could restore METH withdrawal-induced anxiety and depression-like behaviours.
C_LI


title: Normalizing automatic spinal cord cross-sectional area measures

abstract:
Spinal cord cross-sectional area (CSA) is a relevant biomarker to assess spinal cord atrophy in various neurodegenerative diseases. However, the considerable inter-subject variability among healthy participants currently limits its usage. Previous studies explored factors contributing to the variability, yet the normalization models were based on a relatively limited number of participants (typically < 300 participants), required manual intervention, and were not implemented in an open-access comprehensive analysis pipeline. Another limitation is related to the imprecise prediction of the spinal levels when using vertebral levels as a reference; a question never addressed before in the search for a normalization method. In this study we implemented a method to measure CSA automatically from a spatial reference based on the central nervous system (the pontomedullary junction, PMJ), we investigated various factors to explain variability, and we developed normalization strategies on a large cohort (N=804).

Cervical spinal cord CSA was computed on T1w MRI scans for 804 participants from the UK Biobank database. In addition to computing cross-sectional at the C2-C3 vertebral disc, it was also measured at 64 mm caudal from the PMJ. The effect of various biological, demographic and anatomical factors was explored by computing Pearsons correlation coefficients. A stepwise linear regression found significant predictors; the coefficients of the best fit model were used to normalize CSA.

The correlation between CSA measured at C2-C3 and using the PMJ was y = 0.98x + 1.78 (R2 = 0.97). The best normalization model included thalamus volume, brain volume, sex and interaction between brain volume and sex. With this model, the coefficient of variation went down from 10.09% (without normalization) to 8.59%, a reduction of 14.85%.

In this study we identified factors explaining inter-subject variability of spinal cord CSA over a large cohort of participants, and developed a normalization model to reduce the variability. We implemented an approach, based on the PMJ, to measure CSA to overcome limitations associated with the vertebral reference. This approach warrants further validation, especially in longitudinal cohorts. The PMJ-based method and normalization models are readily available in the Spinal Cord Toolbox.


title: Fibrillar-Level Strain Gradients Across the Calcified Bone-Cartilage Interface

abstract: The bone-cartilage interface (BCI) and underlying calcified plate is a universal feature in diarthrodial joints. The BCI is an important mechanically-graded interface subjected to shear and compressive strains, and changes at the BCI have been linked to osteoarthritis progression. Here we report the existence of a physiological internal strain gradient (pre-strain) across the BCI at the ultrastructural scale of the extracellular matrix constituents, specifically the collagen fibril. We use X-ray scattering that probes changes in the axial periodicity of fibril-level D-stagger of tropocollagen molecules in the matrix fibrils, as a measure of microscopic pre-strain. We find that mineralized collagen nanofibrils in the calcified BCI are in tension pre-strain relative to the underlying trabecular bone. This behaviour contrasts with the previously accepted notion that fibrillar pre-strain (or D-stagger) in collagenous tissues always reduces with mineralization due to reduced hydration and associated swelling pressure. Within the calcified tissue, a finer-scale gradient in pre-strain over ~50m is likely linked to the tidemark. The increased fibrillar pre-strain at the BCI is linked to prior research reporting large tissue-level residual strains under compression. The findings may have biomechanical adaptative significance: higher in-built molecular level resilience/damage resistance to physiological compression, and the disruption of the molecular-level pre-strains during remodelling of the BCI may be a potential factor in osteoarthritis-based degeneration.


title: ContaTester: Fast cross-contamination estimation and identification for large human sequencing cohorts

abstract:
BackgroundInterest in genomic medicine for human health studies and clinical applications is rapidly increasing. Clinical applications require contamination-free samples to avoid misleading results and provide a sound basis for diagnosis.

ResultsHere we present ContaTester, a tool which requires only allele balance information gathered from a VCF file to detect cross-contamination in germline human DNA samples. Based on a regression model of allele balance distribution, ContaTester allows fast checking of contamination levels for single samples or large cohorts (less than two minutes per sample). We demonstrate the efficiency of ContaTester using experimental validations: ContaTester shows similar results to methods requiring alignment data but with a significantly reduced storage footprint and less computation time. Additionally, for contamination levels above 5%, ContaTester can identify contaminants across a cohort, providing important clues for troubleshooting and quality assessment.

ConclusionsContaTester estimates contamination levels from VCF files generated from whole genome sequencing normal sample and provides reliable contaminant identification for cohorts or experimental batches.


title: Evaluating metagenomic assembly approaches for biome-specific gene catalogues

abstract:
For many environments, biome-specific microbial gene catalogues are being recovered using shotgun metagenomics followed by assembly and gene-calling on the assembled contigs. The assembly can be conducted either by individually assembling each sample or by co-assembling reads from all the samples. The co-assembly approach can potentially recover genes that display too low abundance to be assembled from individual samples. On the other hand, combining samples increases the risk of mixing data from closely related strains, which can hamper the assembly process. In this respect, assembly on individual samples followed by clustering of (near) identical genes is likely preferable. Thus, both approaches have pros and cons and it remains to be evaluated which assembly strategy is most effective. Here, we have evaluated three assembly strategies for generating gene catalogues from metagenomes using a dataset of 124 samples from the Baltic Sea: 1) assembly on individual samples followed by clustering of the resulting genes, 2) co-assembly on all samples, and 3) mix-assembly, combining individual and co-assembly. The mix-assembly approach resulted in a more extensive non-redundant gene set than the other approaches, and with more genes predicted to be complete and that could be functionally annotated. The mix-assembly consists of 67 million genes (Baltic Sea gene set; BAGS) that have been functionally and taxonomically annotated. The majority of the BAGS genes are dissimilar (<95% amino acid identity) to the Tara Oceans gene dataset, and hence BAGS represents a valuable resource for brackish water research.

IMPORTANCESeveral ecosystem types, such as soils and oceans, are studied through metagenomics. It allows the analysis of genetic material of the microbes within a sample without the need for cultivation. When performing the DNA sequencing with an instrument that generates short sequence reads, these reads need to be assembled in order to obtain more complete gene sequences. In this paper, we have evaluated three strategies for assembling metagenome sequences using a large metagenomic dataset from the Baltic Sea. The method that we call mix-assembly generated the greatest number of non-redundant genes and the largest fraction of genes that were predicted to be complete. The resulting gene catalogue will serve as an important resource for brackish water research. We believe this method to be efficient also for generating gene catalogs for other biomes.


title: TranDTA: Prediction Of Drug Target Binding Affinity Using Transformer Representations

abstract: Drug discovery is generally difficult, expensive, and low success rate. One of the essential steps in the early stages of drug discovery and drug repurposing is identifying drug-target interactions. Binding affinity indicates the strength of drug-target pair interactions. In this regard, several computational methods have been developed to predict the drug-target binding affinity, and the input representation of these models has been shown to be very effective in improving accuracy. Although the recent models predict binding affinity more accurate than the first ones, they need the structure of target proteins. Despite the strong interest in protein structure, there is a massive gap between known sequences and experimentally determined structures. Therefore, finding an appropriate presentation for drug and protein sequences is vital for drug-target binding affinity prediction. In this paper, our primary goal is to assess the drug and protein sequence representation for improving drug-target binding affinity prediction.


title: Drug-Target Binding Affinity Prediction Using Transformers


title: A comprehensive atlas of fetal splicing patterns in the brain of adult myotonic dystrophy type 1 patients

abstract: In patients with myotonic dystrophy type 1 (DM1), dysregulation of RNA-binding proteins like MBNL and CELF1 leads to alternative splicing of exons and is thought to induce a return to fetal splicing patterns in adult tissues, including the central nervous system (CNS). To comprehensively evaluate this, we created an atlas of developmentally regulated splicing patterns in the frontal cortex of healthy individuals and DM1 patients by combining RNA-seq data from BrainSpan, GTEx and DM1 patients. Thirty four splice events displayed an inclusion pattern in DM1 patients that is typical for the fetal situation in healthy individuals. The regulation of DM1-relevant splicing patterns could partly be explained by changes in mRNA expression of the splice regulators MBNL1, MBNL2 and CELF1. On the contrary, interindividual differences in splicing patterns between healthy adults could not be explained by differential expression of these splice regulators. Our findings lend transcriptome-wide evidence to the previously noted shift to fetal splicing patterns in the adult DM1 brain as a consequence of an imbalance in antagonistic MBNL and CELF1 activities. Our atlas serves as a solid foundation for further study and understanding of the cognitive phenotype in patients.


title: Towards mouse genetic-specific RNA-sequencing read mapping

abstract: Genetic variations affect behavior and cause disease but understanding how these variants drive complex traits is still an open question. A common approach is to link the genetic variants to intermediate molecular phenotypes such as the transcriptome using RNA-sequencing (RNA-seq). Paradoxically, these variants between the samples are usually ignored at the beginning of RNA-seq analyses of many model organisms. This can skew the transcriptome estimates that are used later for downstream analyses, such as expression quantitative trait locus (eQTL) detection. Here, we assessed the impact of reference-based analysis on the transcriptome and eQTLs in a widely-used mouse genetic population: the BXD panel of recombinant inbred lines. We highlight existing reference bias in the transcriptome data analysis and propose practical solutions which combine available genetic variants, genotypes, and genome reference sequence. The use of custom BXD line references improved downstream analysis compared to classical genome reference. These insights would likely benefit genetic studies with a transcriptomic component and demonstrate that genome references might need to be reassessed and improved.