title: Inversion of asymmetric histone deposition upon replication stress

abstract: Following DNA replication, equal amounts of histones are distributed over sister chromatids by re-deposition of parental histones and deposition of newly synthesized histones. Molecular mechanisms balancing the allocation of new and old histones remain largely unknown. Here, we studied the genome-wide distribution of new histones relative to parental DNA template strands and replication initiation zones using double-click-seq. In control conditions, new histones were preferentially found on DNA replicated by the lagging strand machinery. Strikingly, replication stress induced by hydroxyurea or curaxin treatment, and inhibition of ATR or p53 inactivation, inverted the observed histone deposition bias to the strand replicated by the leading strand polymerase in line with previously reported effects on RPA occupancy. We propose that asymmetric deposition of newly synthesized histones onto sister chromatids reflects differences in the processivity of leading and lagging strand synthesis.


title: Beech tree masting explains the inter-annual variation in the fall and spring peaks of Ixodes ricinus ticks with different time lags

abstract:
BackgroundThe tick Ixodes ricinus is an important vector of tick-borne diseases including Lyme borreliosis. In continental Europe, the nymphal stage of I. ricinus often has a bimodal phenology with a large spring/early summer peak and a smaller fall peak. While there is consensus about the origin of the spring nymphal peak, there are two alternative hypotheses for the fall nymphal peak, direct development versus delayed diapause. These two hypotheses make different predictions about the time lags of the correlations between the spring peak, the fall peak, and seed production (masting) by deciduous trees.

MethodsTo determine which hypothesis is most important for explaining the fall peak, we used data from a long-term surveillance study (15 years) on the density of I. ricinus nymphal ticks at 4 different elevation sites in an area of Switzerland that is endemic for Lyme borreliosis, and long-term data on the mast of the European beech tree from the literature.

ResultsI. ricinus nymphs had a bimodal phenology at the three lower elevation sites, but a unimodal phenology at the top elevation site. At the lower elevation sites, the density of nymphs (DON) in the fall was strongly correlated with the DON in the spring of the following year. The inter-annual variation in the densities of I. ricinus nymphs in the fall and spring were best explained by a 1-year versus a 2-year time lag with the beech tree masting index. Fall nymphs had higher fat content and are younger than spring nymphs. All of these observations are consistent with the direct development hypothesis for the fall peak of I. ricinus nymphs at our study site. Our study provides new insight into the complex bimodal phenology of this important disease vector.

ConclusionsPublic health officials in Europe should be aware that following a strong mast year, the DON will increase 1 year later in the fall and 2 years later in the spring and summer. Population ecology studies of I. ricinus should consider that the spring and fall peak in the same calendar year represent different generations of ticks.

category: ecology


title: Single-cell analysis reveals MHCII expressing keratinocytes in pressure ulcers with worse healing outcomes

abstract: Pressure ulcer (PU) is a chronic wound often seen in spinal cord injury patients and other bed-bound individuals, particularly in the elderly population. Despite its association with high mortality, the pathophysiology of PU remains poorly understood. Here, we compared single-cell transcriptomic profiles of human epidermal cells from PU wound edges with those from uninjured skin and acute wounds (AWs) in healthy donors. We identified significant shifts in the cell composition and gene expression patterns in PU. In particular, we found that major histocompatibility complex class II (MHCII) expressing keratinocytes were enriched in patients with worse healing outcomes. Furthermore, we showed that the IFN{gamma} in PU-derived wound fluid could induce MHCII expression in keratinocytes and that these wound fluid-treated keratinocytes inhibited autologous T cell activation. In line with this observation, we found that T cells from PUs enriched with MHCII+ keratinocytes produced fewer inflammatory cytokines. Overall, our study provides a high-resolution molecular map of human PU compared to AW and intact skin, providing new insights into PU pathology and the future development of tailored wound therapy.

category: genomics


title: Genome annotation with long RNA reads reveals new patterns of gene expression in an ant brain

abstract: Functional genomic analyses rely on high-quality genome assemblies and annotations. Highly contiguous genome assemblies have become available for a variety of species, but accurate and complete annotation of gene models, inclusive of alternative splice isoforms and transcription start and termination sites remains difficult with traditional approaches. Here, we utilized full-length isoform sequencing (Iso-Seq), a long-read RNA sequencing technology, to obtain a comprehensive annotation of the transcriptome of the ant Harpegnathos saltator. The improved genome annotations include additional splice isoforms and extended 3 untranslated regions for more than 4,000 genes. Reanalysis of RNA-seq experiments using these annotations revealed several genes with caste-specific differential expression and tissue-or caste-specific splicing patterns that were missed in previous analyses. The extended 3 untranslated regions afforded great improvements in the analysis of existing single-cell RNA-seq data, resulting in the recovery of the transcriptomes of 18% more cells. The deeper single-cell transcriptomes obtained with these new annotations allowed us to identify additional markers for several cell types in the ant brain, as well as genes differentially expressed across castes in specific cell types. Our results demonstrate that Iso-Seq is an efficient and effective approach to improve genome annotations and maximize the amount of information that can be obtained from existing and future genomic datasets in Harpegnathos and other organisms.


title: Allosteric linkages that emulate a molecular motor enzyme

abstract: Allosteric mechanisms are fundamental to the operation of biomolecular motors. Recreating the molecular phenomena associated with allostery, from first principles, would help advance the design and construction of synthetic molecular motors, which remain quite simple compared natural motors. In this study, I present a model for generating allosteric interactions using mechanical linkages, which are devices in which flexible nodes are connected by rigid rods. I describe how allosteric information can be communicated between multivalent binding sites on an enzyme when linkages bind or dissociate in a stepwise fashion, which takes place stochastically according to assigned binding rates and partitioned binding energies. This design allows geometric competitions to autonomously push a linkage enzyme through a desired sequence of states, driven by consumption of a fuel. I use the model to emulate the chemical and conformational cycle of a myosin monomer, which is demonstrated by simulating chemical reaction networks of the linkage structures, and by describing how two linkage monomers can be connected together to construct a motor that walks on a track. This work shows how the complex behavior of biomolecular motors can be recapitulated with simple geometric and chemical principles that encode allosteric mechanisms. Because the concepts are material-agnostic, they can potentially be used to design and construct allosteric machines using various chemistries.

category: bioengineering


title: Stitching and registering highly multiplexed whole slide images of tissues and tumors using ASHLAR software

abstract:
MotivationStitching microscope images into a mosaic is an essential step in the analysis and visualization of large biological specimens, particularly human and animal tissues. Recent approaches to highly-multiplexed imaging generate high-plex data from sequential rounds of lower-plex imaging. These multiplexed imaging methods promise to yield precise molecular single-cell data and information on cellular neighborhoods and tissue architecture. However, attaining mosaic images with single-cell accuracy requires robust image stitching and image registration capabilities that are not met by existing methods.

ResultsWe describe the development and testing of ASHLAR, a Python tool for coordinated stitching and registration of 103 or more individual multiplexed images to generate accurate whole-slide mosaics. ASHLAR reads image formats from most commercial microscopes and slide scanners, and we show that it performs better than existing open source and commercial software. ASHLAR outputs standard OME-TIFF images that are ready for analysis by other open-source tools and recently developed image analysis pipelines.

Availability and implementationASHLAR is written in Python and available under an MIT license at https://github.com/labsyspharm/ashlar. An informational website with user guides and test data is available at https://labsyspharm.github.io/ashlar/.

category: bioinformatics


title: Stitching and registering highly multiplexed whole slide images of tissues and tumors using ASHLAR


title: Humans postural sways: non-Gauss, variability, and aging aspects of a data set mining

abstract:
This report presents the results of data mining of a sample with two focus-groups. They have the same sizes: seven people into each one, random-wise picked up from the same data set. The trials of groups were idem. The postural swings, that is, the move of the center-of-pressure (COP), were recorded. Maple, a computer math system, has allowed us to apply the Principal Components Analysis, Statistical analysis, Kernel Density Estimations (KDE) for the probabilities. Poincare and Recurrence Plots were other tools for modern data mining. The non-Gauss features of the real distributions are not that to be neglect. They exist not only as outliers but as sharp kurtosis and skewness. Still, they are so far not enow to grave doubts to the Fractional Brownian model. We found some subtle aging hallmarks for focus-groups. First, the trend of variability descriptors to be bi-modal is sheerer for the older group. Second, the coefficients of correlations of the short-time variability index with other descriptors are clear age-related.

2012 ACM Subject ClassificationApplied computing [->] Life and medical sciences [->] Health informatics


title: Direct fast estimator for recombination frequency in the F2 cross

abstract: In this short note, a new unbiased maximum-likelihood estimator is proposed for the recombination frequency in the F2 cross. The estimator is much faster to calculate than its EM algorithm equivalent, yet as efficient. Simulation studies are carried to illustrate the gain over another simple estimate proposed by Benito & Gallego (2004).


title: Host EPAC1 modulates rickettsial adhesion to vascular endothelial cells via regulation of ANXA2 Y23 phosphorylation

abstract:
Recently we have identified that endothelial surface annexin A2 (ANAX2) functions as a receptor for spotted fever group rickettsial adhesin outer membrane protein B (OmpB), which binds to the endothelial cell (EC) surface. Moreover, we reported that intracellular cAMP receptor EPAC1 modulates ANXA2 tyrosine (Y) 23 phosphorylation, and inactivation of EPAC1 suppresses ANXA2 expression on the EC luminal surface by downregulating Y23 phosphorylation. Since we reported that EPAC1 plays a critical role in the initial step to successfully establish rickettsial infection of ECs, this work aims to answer the following: (a) What is the mechanism underlying the regulatory role of EPAC1 in ECs during the initial step of bacterial infection? (b) Is the EPAC1-ANXA2 signaling pathway involved in the regulation of rickettsial adhesion to ECs?

In the present study, an established system that is anatomically-based and quantifies bacterial adhesion to ECs in vivo was combined with novel fluidic force microscopy (FluidFM) to dissect the functional role of the EPAC1-ANXA2 signaling pathway in rickettsiae-EC adhesion. We reveal that the deletion of the EPAC1 gene impedes rickettsial binding to endothelium in vivo. In addition, single living brain microvascular EC study that employs FluidFM and site-directed mutagenesis provides evidence that supports our finding that EPAC1 governs rickettsial adhesion to EC surfaces via regulation of ANXA2 Y23 phosphorylation.

category: pathology


title: Non-structural carbohydrate concentrations in woody organs, but not leaves, of temperate and tropical tree angiosperms are independent of the "fast-slow" plant economic spectrum

abstract:
Background and AimsCarbohydrate reserves play a vital role in plant survival during periods of negative carbon balance. Considering active storage of reserves, there is a trade-off between carbon allocation to growth and to reserves and defense. A resulting hypothesis is that allocation to reserves exhibits a coordinated variation with functional traits associated with the  fast-slow plant economics spectrum.

MethodsWe tested the relationship between non-structural carbohydrates (NSC) of tree organs and functional traits using 61 angiosperm tree species from temperate and tropical forests with phylogenetic hierarchical Bayesian models.

Key ResultsOur results provide evidence that NSC concentrations in woody organs and plant functional traits are largely decoupled, meaning that species resilience is unrelated to their position on the  fast-slow plant economics spectrum. In contrast, we found that variation between NSC concentrations in leaves and the fast-slow continuum was coordinated, as species with higher leaf NSC had traits values associated with resource conservative species such as lower SLA, lower Amax, and high wood density. We did not detect an influence of leaf habit on the variation of NSC concentrations in tree organs.

ConclusionsEfforts to predict the response of ecosystems to global change will need to integrate a suite of plant traits, such as NSC concentrations in woody organs, that are independent of the  fast-slow spectrum and that capture how species respond to a broad range of global change factors.

category: plant biology


title: Exploring Extreme Signaling Failures in Intracellular Molecular Networks

abstract: Developing novel methods for the analysis of intracellular signaling networks is essential for understanding interconnected biological processes that underlie complex human disorders. A fundamental goal of this research is to quantify the vulnerability of a signaling network to the dysfunction of one or multiple molecules, when the dysfunction is defined as an incorrect response to the input signals. In this study, we propose an efficient algorithm to identify the extreme signaling failures that can induce the most detrimental impact on the physiological function of a molecular network. The algorithm basically finds the molecules, or groups of molecules, with the maximum vulnerability, i.e., the highest probability of causing the network failure, when they are dysfunctional. We propose another algorithm that efficiently accounts for signaling feedbacks in this analysis. The algorithms are tested on two experimentally verified ERBB and T cell signaling networks. Surprisingly, results reveal that as the number of concurrently dysfunctional molecules increases, the maximum vulnerability values quickly reach to a plateau following an initial increase. This suggests the specificity of vulnerable molecule (s) involved, as a specific number of faulty molecules cause the most detrimental damage to the function of the network. Increasing a random number of simultaneously faulty molecules does not further deteriorate the function of the network. Such a group of specific molecules whose dysfunction causes the extreme signaling failures can better elucidate the molecular mechanisms underlying the pathogenesis of complex trait disorders, and can offer new insights for the development of novel therapeutics.

category: systems biology


title: Three bacterium-plasmid golden combinations facilitate the spread of ST11/CG258 carbapenemase-producing Klebsiella pneumoniae in China

abstract: Carbapenemase-producing Klebsiella pneumoniae (cpKP) poses serious threats to public health. Previous studies showed that only ST11/CG258-cpKP successfully disseminated in China, however, the underlying genetic bases are still unknown. We conducted a comprehensive genomic-epidemiology analysis on 420 cpKP isolates from 70 hospitals in 24 Chinese provinces during 2009-2017 based on short-/long-reads sequencing. Three  golden combinations of host--blaKPC-carrying plasmids (Clade 3.1+3.2--IncFIIpHN7A8, Clade 3.1+3.2--IncFIIpHN7A8:IncR, Clade 3.3--IncFIIpHN7A8:IncpA1763-KPC) endowed cpKP with advantages both in genotypes (strong-correlation/co-evolution) and phenotypes (resistance/growth/competition), thereby facilitating nationwide spread of ST11/CG258-cpKP. Intriguingly, Bayesian skyline illustrated that the three  golden combinations might directly lead to the strong population expansion during 2007-2008 and subsequent maintenance of the dissemination of ST11/CG258-cpKP after 2008. We tested drug-resistance profiles and proposed combination treatment regimens for CG258/non-CG258 cpKP. Our findings systematically revealed the molecular-epidemiology and genetic-basis for dissemination of Chinese ST11/CG258 cpKP and reminded us to monitor the  golden combinations of cpKP- plasmid closely.

category: microbiology


title: Isolation and characterization of novel temperate virus Aeropyrum globular virus 1 infecting hyperthermophilic archaeon Aeropyrum

abstract:
We isolate a novel archaeal temperate virus named Aeropyrum globular virus 1 (AGV1) from the host Aeropyrum culture. Reproduction of AGV1 was induced by adding 20 mM tris-acetate buffer to exponentially growing host cells. Negatively stained virions showed spherical morphology (60 {+/-}2 nm in diameter) similar to Globuloviridae viruses. The double-stranded circular DNA genome of AGV1 contains 18,222 bp encoding 34 open-reading frames. No ORFs showed significant similarity with Globuloviridae viruses. AGV1 shares three genes, including an integrase gene, with reported spindle-shaped temperate viruses. However we couldnt detect its integration site in the host genome. Moreover AGV1 seemed not to replicate autonomously because there are no origin recognition boxes in the genome. qPCR results showed that the genome copy number of AGV1 was lower than that of the host genome (10-3 copies per host genome). Upon the addition of tris-acetate buffer, a steep increase in the AGV1 genome copy number (9.5-26 copies per host genome at 2 days post-treatment) was observed although clustered regularly interspaced short palindromic repeat (CRISPR) elements of the host genome showed significant matches with AGV1 protospacers. Our findings suggest that AGV1 is a novel globular virus exhibiting an unstable carrier state in the growing host and in that way AGV1 can escape from the host defense system and propagate under stressful host conditions.

ImportanceStudying archaeal viruses yields novel insights into the roles of virospheres and viruses in the evolutionary process of their hosts. Here, we isolated a novel spherical virus named Aeropyrum globular virus 1. AGV1 has integrase gene but its genome is not integrated into the host genome. AGV1 could not replicate autonomously due to the lack of origin recognition boxes and thus its copy number was too low (10-3 copies per host genome) without any inducing stimulus. However, upon the addition of tris-acetate buffer, the AGV1 genome copy number steeply increased instead of a perfect sequence match between the spacer of the host CRISPR/Cas system and the protospacer. Our findings suggest that AGV1 can escape from the host defense system and propagate under stressful conditions for the host by establishing an unstable carrier state. These results reveals a novel aspect of host-virus interactions in extreme environments.


title: Hydrogen sulfide blocks HIV rebound by maintaining mitochondrial bioenergetics and redox homeostasis

abstract: A fundamental challenge in HIV eradication is to understand how the virus establishes latency, maintains stable cellular reservoirs, and promotes rebound upon interruption of antiretroviral treatment (ART). Here, we discovered an unexpected role of the ubiquitous gasotransmitter hydrogen sulfide (H2S) in HIV latency and reactivation. We show that reactivation of HIV-1 is associated with down-regulation of the key H2S producing enzyme cystathionine-{gamma}-lyase (CTH) and reduction in endogenous H2S. Genetic silencing of CTH disrupts redox homeostasis, impairs mitochondrial function, and remodels the transcriptome of latent cells to trigger HIV reactivation. Chemical complementation of CTH activity using a slow-releasing H2S donor, GYY4137, suppressed HIV reactivation and diminished virus replication. Mechanistically, GYY4137 blocked HIV reactivation by inducing the Keap1-Nrf2 pathway, inhibiting NF-{kappa}B, and recruiting the epigenetic silencer, YY1, to the HIV promoter. In latently infected CD4+ T cells from ART-suppressed human subjects, GYY4137 in combination with ART prevented viral rebound and improved mitochondrial bioenergetics. Moreover, prolonged exposure to GYY4137 exhibited no adverse influence on proviral content or CD4+ T cell subsets, indicating that diminished viral rebound is due to a loss of transcription rather than a selective loss of infected cells. In summary, this work provides mechanistic insight into H2S-mediated suppression of viral rebound and suggests the inclusion of an H2S donor in the current ART regimen to achieve a functional HIV-1 cure.


title: HIV-1 provirus transcription and translation in macrophages differs from macrophages containing pre-integrated cDNA complexes and requires E2F transcriptional programs

abstract:
HIV-1 cDNA pre-integration complexes have been shown to persist for weeks in macrophages and to be transcriptionally active. Early and late gene transcripts are produced, along with some viral proteins, yet whole virus is not. While previous work has focused on the transcription and translation of HIV-1 genes; our understanding of cellular milieu that accompanies viral production is incomplete. We have used an in vitro system to model HIV-1 infection of macrophages, and single cell RNA sequencing (scRNA-seq) to compare the transcriptomes of uninfected cells, cells harboring pre-integration HIV-1 complexes (PIC) and those containing integrated provirus and actively making late HIV proteins. These are also compared to control cells, not exposed to virus.

Several observations provide new perspective on the effects of HIV-1 transcription from pre-integrated cDNA versus from integrated provirus. First, HIV-1 transcript levels do not necessarily correlate with virus production, cells harboring PIC cDNA have transcript loads comparable to cells transcribing from provirus and making p24, mCherry, and vpu proteins. Second, all HIV-1 transcripts are easily detectable in abundance from PIC cDNA transcription, as is the case with cells transcribing from provirus, although the frequency of PIC cells with detectable gag-pol, tat, env, and nef transcripts is higher than the corresponding frequencies observed for "Provirus cells". Third, the background transcriptomes of cells harboring pre- integrated HIV-1 cDNA are not otherwise detectably altered from cells not containing any HIV- 1 transcript. Fourth, integration and production of p24, mCherry, and Vpu proteins is accompanied by a switch from transcriptomes characterized by NFkB and AP-1 promoted transcription to a transcriptome characterized by E2F family transcription products. While some of these observations may seem heretical, single cell analysis provides a more nuanced understanding of PIC cDNA transcription and the transcriptomic changes that support HIV-1 protein production from integrated provirus.

Author SummarySingle cell analysis is able to distinguish between HIV-1 infected macrophage cells that are transcribing pre-integrated HIV-1 cDNA and those transcribing HIV-1 provirus. Only cells transcribing HIV-1 provirus are making p24, marker mCherry and Vpu proteins, which corresponds with a change in the host cells background transcriptome from one expressing viral restriction and immunological response genes to one that is expressing genes associated with cell replication and oxidative phosphorylation.


title: MARCH8 targets cytoplasmic lysine residues of various viral envelope glycoproteins

abstract: The host transmembrane protein MARCH8 is a RING finger E3 ubiquitin ligase that downregulates various host transmembrane proteins, such as MHC-II. We have recently reported that MARCH8 expression in virus-producing cells impairs viral infectivity by reducing virion incorporation of not only HIV-1 envelope glycoproteins but also vesicular stomatitis virus G-glycoprotein through two different pathways. However, the MARCH8 inhibition spectrum remains largely unknown. Here, we investigate the antiviral spectrum of MARCH8 using HIV-1 pseudotyped with a variety of viral envelope glycoproteins. Pseudotyping experiments revealed that viral envelopes derived from the rhabdovirus, arenavirus, coronavirus, and togavirus (alphavirus) families were sensitive to MARCH8-mediated inhibition. Lysine mutations at the cytoplasmic tails of rabies virus-G, lymphocytic choriomeningitis virus glycoproteins, SARS-CoV and SARS-CoV-2 spike proteins, and Chikungunya virus and Ross River virus E2 proteins conferred resistance to MARCH8. Immunofluorescence showed impaired downregulation of the mutants of these viral envelopes by MARCH8, followed by lysosomal degradation, suggesting that MARCH8-mediated ubiquitination leads to intracellular degradation of these envelopes. Indeed, rabies virus-G and Chikungunya virus E2 proteins proved to be clearly ubiquitinated. We conclude that MARCH8 has inhibitory activity on a variety of viral envelope glycoproteins whose cytoplasmic lysine residues are targeted by this antiviral factor.


title: Functional and structural characterization of Serratia marcescens ExbB: determinants of the interaction with HasB/TonB

abstract: ExbB and ExbD are cytoplasmic membrane proteins that associate with TonB to convey the energy of the proton-motive force to outer membrane receptors in Gram-negative bacteria for iron uptake. The opportunistic pathogen Serratia marcescens (Sm) possesses both TonB and a heme-specific TonB paralog, HasB. ExbBSm has a long periplasmic extension absent in other bacteria such as E. coli (Ec). Long ExbBs are found in several genera of Alphaproteobacteria, most often in correlation with a hasB gene. We investigated specificity determinants of ExbBSm and HasB. We determined the cryo-EM structures of ExbBSm and of the ExbB-ExbDSm complex from S. marcescens. ExbBSm alone is a stable pentamer, and its complex includes two ExbD monomers. We showed that ExbBSm extension interacts with HasB and is involved in heme acquisition and we identified key residues in the membrane domain of ExbBSm and ExbBEc, essential for function and likely involved in the interaction with TonB/HasB. Our results shed light on the new class of inner membrane energy machinery formed by ExbB,ExbD and HasB.


title: Structural and molecular determinants for the interaction of ExbB from Serratia marcescens and HasB, a TonB paralog


title: Caerin 1.1 and 1.9 peptides from Australian tree frog inhibit antibiotic-resistant bacteria growth in a murine skin infection model

abstract: Host-defence caerin 1.9 peptide was originally isolated from skin secretion of Australian tree frog, and inhibits the growth of a wide range of bacteria in vitro. In this study, we demonstrated that caerin 1.9 shows high bioactivity against several bacteria strains, such as Staphylococcus aureus, Acinetobacter Baumannii, methicillin-resistant Staphylococcus aureus (MRSA), and Streptococcus hemolyiicus in vitro. Importantly, unlike antibiotic Tazocin, caerin 1.9 does not induce bacterial resistance after 30 rounds of in vitro culture. Moreover, caerin 1.1, another peptide of caerin family, has additive antibacterial effect when used together with caerin 1.9. Furthermore, caerin 1.1 and 1.9 prepared in the form of a temperature sensitive gel inhibit MRSA growth in skin bacterial infection model of two murine strains. These results indicate that caerin 1.1 and 1.9 may have the advantage than conventional antibiotics against bacterial infection of skin.


title: Evolution under low antibiotic concentrations: a risk for the selection of Pseudomonas aeruginosa multidrug resistant mutants in nature

abstract:
BACKGROUNDAntibiotic pollution of non-clinical environments might have a relevant impact on human health if resistant pathogens are selected. However, this potential risk is often overlooked, since drug concentrations in nature are usually below their minimal inhibitory concentrations (MICs). Albeit, antibiotic resistant bacteria can be selected even at sub-MIC concentrations, in a range that is dubbed the sub-MIC selective window, which depends on both the antibiotic and the pathogen.

OBJECTIVESDetermine the sub-MIC selective windows of seven antibiotics of clinical relevance in the opportunistic pathogen Pseudomonas aeruginosa and evaluate the risk for selecting resistant mutants in nature, based on published data about the amount of antimicrobials detected in natural environments.

METHODSWe conducted evolution experiments of P. aeruginosa PA14 in presence of sub-MIC concentrations of ceftazidime, amikacin, levofloxacin, ciprofloxacin, tetracycline, polymyxin B or imipenem, and measured drug susceptibility of the evolved populations.

RESULTSSub-MIC selective window of quinolones was the largest, and the ones of polymyxin B and imipenem, the narrowest. Clinically relevant multidrug resistant (MDR) mutants (presenting MICs above EUCAST clinical breakpoints) arose within the sub-MIC selective windows of the majority of antibiotics tested, being these phenotypes probably mediated by efflux pumps' activity.

DISCUSSIONOur data show that the concentration of antibiotics reported in aquatic ecosystems -colonizable by P. aeruginosa- are, in occasions, higher than the ones able to select MDR mutants. This finding has implications for understanding the role of different ecosystems and conditions in the emergence of antibiotic resistance from a One-Health point of view. Further, it highlights the importance of delineating the sub-MIC selective windows for drugs of clinical value in pathogens with environmental niches, in order to evaluate the health risks due to antibiotic pollution of natural ecosystems and ultimately tackle antibiotic resistance.


title: Rapid and Efficient Inactivation of SARS-CoV-2 from Surfaces using UVC Light Emitting Diode Device

abstract: Efforts are underway to develop countermeasures to prevent the environmental spread of COVID-19 pandemic caused by SARS-CoV-2. Physical decontamination methods like Ultraviolet radiation has shown to be promising. Here, we describe a novel device emitting ultraviolet C radiation (UVC), called NuvaWave, to rapidly and efficiently inactivate SARS-CoV-2. SARS-CoV-2 was dried on a chambered glass slides and introduced in a NuvaWave robotic testing unit. The robot simulated waving NuvaWave over the virus at a pre-determined UVC radiation dose of 1, 2, 4 and 8 seconds. Post-UVC exposure, virus was recovered and titered by plaque assay in Vero E6 cells. We observed that relative control (no UVC exposure), exposure of the virus to UVC for one or two seconds resulted in a >2.9 and 3.8 log10 reduction in viral titers, respectively. Exposure of the virus to UVC for four or eight seconds resulted in a reduction of greater than 4.7-log10 reduction in viral titers. The NuvaWave device inactivates SARS-CoV-2 on surfaces to below the limit of detection within one to four seconds of UVC irradiation. This device can be deployed to rapidly disinfect surfaces from SARS-CoV-2, and to assist in mitigating its spread in a variety of settings.


title: Eicosanoid signaling as a therapeutic target in middle-aged mice with severe COVID-19

abstract: Coronavirus disease 2019 (COVID-19) is especially severe in aged populations1. Resolution of the COVID-19 pandemic has been advanced by the recent development of SARS-CoV-2 vaccines, but vaccine efficacy is partly compromised by the recent emergence of SARS-CoV-2 variants with enhanced transmissibility2. The emergence of these variants emphasizes the need for further development of anti-SARS-CoV-2 therapies, especially in aged populations. Here, we describe the isolation of a new set of highly virulent mouse-adapted viruses and use them to test a novel therapeutic drug useful in infections of aged animals. Initially, we show that many of the mutations observed in SARS-CoV-2 during mouse adaptation (at positions 417, 484, 501 of the spike protein) also arise in humans in variants of concern (VOC)2. Their appearance during mouse adaptation indicates that immune pressure is not required for their selection. Similar to the human infection, aged mice infected with mouse-adapted SARS-CoV-2 develop more severe disease than young mice. In murine SARS, in which severity is also age-dependent, we showed that elevated levels of an eicosanoid, prostaglandin D2 (PGD2) and of a phospholipase, PLA2G2D, contributed to poor outcomes in aged mice3,4. Using our virulent mouse-adapted SARS-CoV-2, we show that infection of middle-aged mice lacking expression of DP1, a PGD2 receptor, or PLA2G2D are protected from severe disease. Further, treatment with a DP1 antagonist, asapiprant, protected aged mice from a lethal infection. DP1 antagonism is one of the first interventions in SARS-CoV-2-infected animals that specifically protects aged animals, and demonstrates that the PLA2G2D-PGD2/DP1 pathway is a useful target for therapeutic interventions. (Words: 254)


title: FACT subunit SUPT16H associates with BRD4 and contributes to silencing of antiviral interferon signaling

abstract: Summary/AbstractFACT (FAcilitates Chromatin Transcription) is a heterodimeric protein complex composed of SUPT16H and SSRP1, and a histone chaperone participating in chromatin remodeling during gene transcription. FACT complex is profoundly regulated, and contributes to both gene activation and suppression. Here we reported that SUPT16H, a subunit of FACT, is acetylated at lysine 674 (K674) of middle domain (MD), which involves TIP60 histone acetyltransferase. Such acetylation of SUPT16H is recognized by bromodomain protein BRD4, which promotes protein stability of SUPT16H. We further demonstrated that SUPT16H-BRD4 associates with histone modification enzymes (EZH2, HDAC1) and affects histone marks (H3K9me3, H3K27me3 and H3ac). BRD4 is known to profoundly regulate interferon (IFN) signaling, while such function of SUPT16H has never been explored. Surprisingly, our results revealed that SUPT16H genetic knockdown via RNAi or pharmacological inhibition by using its inhibitor, curaxin 137 (CBL0137), results in the induction of IFNs and interferon-stimulated genes (ISGs). Through this mechanism, CBL0137 is shown to efficiently inhibit infection of multiple viruses, including Zika, influenza, and SARS-CoV-2. Furthermore, we demonstrated that CBL0137 also causes the remarkable activation of IFN signaling in natural killer (NK) cells, which promotes the NK-mediated killing of virus-infected cells in a co-culture system using human primary NK cells. Overall, our studies unraveled the previously un-appreciated role of FACT complex in regulating IFN signaling in both epithelial and NK cells, and also proposed the novel application of CBL0137 to treat viral infections.

category: molecular biology


title: Single-molecule imaging of chromatin remodelers reveals role of ATPase in promoting fast kinetics of target search and dissociation from chromatin

abstract: Conserved ATP-dependent chromatin remodelers establish and maintain genome-wide chromatin architectures of regulatory DNA during cellular lifespan, but the temporal interactions between remodelers and chromatin targets have been obscure. We performed live-cell single-molecule tracking for RSC, SWI/SNF, CHD1, ISW1, ISW2, and INO80 remodeling complexes in budding yeast and detected hyperkinetic behaviors for chromatin-bound molecules that frequently transition to the free state for all complexes. Chromatin-bound remodelers display notably higher diffusion than nucleosomal histones, and strikingly fast dissociation kinetics with 4-7 s mean residence times. These enhanced dynamics require ATP binding or hydrolysis by the catalytic ATPase, uncovering an additional function to its established role in nucleosome remodeling. Kinetic simulations show that multiple remodelers can repeatedly occupy the same promoter region on a timescale of minutes, implicating an unending  tug-of-war that controls a temporally shifting window of accessibility for the transcription initiation machinery.


title: Cas9-mediated genome editing in Giardia intestinalis

abstract: CRISPR/Cas9 system is an extremely powerful technique that is extensively used for various genome modifications in different organisms including parasitic protists. Giardia intestinalis, a protist parasite infecting about 280 million people around the world each year, has been eluding the routine use of CRISPR/Cas9 for generating knock-out cell lines due to the presence of four copies of each gene in its two nuclei. Apart from single exception employing rather laborious Cre/loxP system, no full knock-out cell line has been established yet. In this work, we show the ability of in-vitro assembled CRISPR/Cas9 components to successfully edit the genome of G. intestinalis. We further established a cell line stably expressing Cas9 in both G. intestinalis nuclei. Subsequent introduction of a template for homologous recombination containing the transcription units for the resistance marker and gRNA resulted in the removal of all gene copies at once for three independent experimental genes, mem, cwp1 and mlf1. The method was also applicable for the incomplete disruption of an essential gene, as documented by markedly decreased expression of tom40. Finally, testing the efficiency of Cas9-induced recombination revealed that homologous arms as short as 150 bp can be sufficient to establish a full knock-out cell line in G. intestinalis.

category: cell biology


title: Cas9-mediated gene disruption in tetraploid Giardia intestinalis


title: Estimating parameters of a stochastic cell invasion model with fluorescent cell cycle labelling using Approximate Bayesian Computation

abstract: We develop a parameter estimation method based on approximate Bayesian computation (ABC) for a stochastic cell invasion model using fluorescent cell cycle labeling with proliferation, migration, and crowding effects. Previously, inference has been performed on a deterministic version of the model fitted to cell density data, and not all the parameters were identifiable. Considering the stochastic model allows us to harness more features of experimental data, including cell trajectories and cell count data, which we show overcomes the parameter identifiability problem. We demonstrate that, whilst difficult to collect, cell trajectory data can provide more information about the parameters of the cell invasion model. To handle the intractability of the likelihood function of the stochastic model, we use an efficient ABC algorithm based on sequential Monte Carlo. Rcpp and MATLAB implementations of the simulation model and ABC algorithm used in this study are available at https://github.com/michaelcarr-stats/FUCCI.


title: Surrogate production of genome edited sperm from a different subfamily by spermatogonial stem cell transplantation

abstract: The surrogate reproduction technique provides a powerful tool for production of allogenic or xenogeneic gametes derived from endangered species or those with valuable genetic traits. Production of functional donor-derived gametes through intra- or inter-specific spermatogonial stem cell transplantation (SSCT) has been achieved in many species. However, generation of functional gametes from a phylogenetically distant species such as from a different subfamily by SSCT has never been successful. Here, using two small cyprinid fishes, Chinese rare minnow (gobiocypris rarus, for brief: Gr) and zebrafish (danio rerio), which belong to different subfamilies, as donors and recipients for SSCT, we optimized the SSCT technique and successfully obtained Gr-derived sperm carrying targeted genome modifications in zebrafish. We revealed that the transplanted Gr spermatogonia supported the host gonadal development and underwent normal spermatogenesis, resulting in a reconstructed fertile testis containing Gr spermatids and zebrafish testicular somatic cells. Interestingly, the surrogate spermatozoa resembled those of host zebrafish but not donor Gr in morphology and swimming behavior. Finally, we showed that Gr-derived genome edited sperm was successfully produced in zebrafish by cross-subfamily SSCT, when the pou5f3 and chd gene knockout Gr SSCs were used as surrogate donors. This is the first report demonstrating the surrogate production of genome edited sperm from a phylogenetically distant species, and this method is feasible to be applied to future breeding of commercial fishes.

category: developmental biology


title: Modelling effects of water regulation on the population viability of threatened amphibians

abstract:
The regulation of river systems alters hydrodynamics and often reduces lateral connectivity between river channels and floodplains. For taxa such as frogs that rely on floodplain wetlands to complete their lifecycle, decreasing inundation frequency can reduce recruitment and increase the probability of local extinction. We virtually reconstructed the inundation patterns of wetlands under natural and regulated flow conditions and built stochastic population models to quantify the probability of local extinction under different inundation scenarios. Specifically, we explored the interplay of habitat size, inundation frequency, and successive dry years on the local extinction probability of the threatened southern bell frog Litoria raniformis in the Murray River floodplains of South Australia. We hypothesised that the changes to wetland inundation resulting from river regulation are a principal driver of L. raniformis declines in this semi-arid system.

Regulation has reduced the inundation frequency of essential habitats below critical thresholds for the persistence of many fresh water-dependent species. Successive dry years raise the probability of local extinction, and these effects are strongest in smaller wetlands. Larger wetlands and those with more frequent average inundation are less susceptible to these effects.

Elucidating these trends informs the prioritisation of treatment sites and the frequency of conservation interventions. Environmental water provision (through pumping or the operation of flow-regulating structures) is a promising tool to reduce the probability of breeding failure and local extinction. Our modelling approach can be used to prioritise the delivery of environmental water to L. raniformis and potentially many other frog species.


title: Forecasting the effects of water regulation on the population viability of a threatened amphibian


title: Virulent but not temperate bacteriophages display hallmarks of rapid translation initiation

abstract: Bacteriophages rely almost exclusively on host-cell machinery to produce their proteins, and their mRNAs must therefore compete with host mRNAs for valuable translational resources. In many bacterial species, highly translated mRNAs are characterized by the presence of a Shine-Dalgarno sequence motif upstream of the start codon and weak secondary structure within the start codon region. However, the general constraints and principles underlying the translation of phage mRNAs are largely unknown. Here, we show that phage mRNAs are highly enriched in strong Shine-Dalgarno sequences and have comparatively weaker secondary structures in the start codon region than host-cell mRNAs. Phage mRNAs appear statistically similar to the most highly expressed host genes in E. coli according to both features, strongly suggesting that they initiate translation at particularly high rates. Interestingly, we find that these observations are driven largely by virulent phages and that temperate phages encode mRNAs with similar start codon features to their host genes. These findings apply broadly across a wide-diversity of host-species and phage genomes. Further study of phage translational regulation--with a particular emphasis on virulent phages--may provide new strategies for engineering phage genomes and recombinant expression systems more generally.


title: GC-biased gene conversion in X-Chromosome palindromes conserved in human, chimpanzee, and rhesus macaque

abstract: Gene conversion is GC-biased across a wide range of taxa. Large palindromes on mammalian sex chromosomes undergo frequent gene conversion that maintains arm-to-arm sequence identity greater than 99%, which may increase their susceptibility to the effects of GC-biased gene conversion. Here, we demonstrate a striking history of GC-biased gene conversion in 12 palindromes conserved on the X chromosomes of human, chimpanzee, and rhesus macaque. Primate X-chromosome palindrome arms have significantly higher GC content than flanking single-copy sequences. Nucleotide replacements that occurred in human and chimpanzee palindrome arms over the past 7 million years are one-and-a-half times as GC-rich than the ancestral bases they replaced. Using simulations, we show that our observed pattern of nucleotide replacements is consistent with GC-biased gene conversion with a magnitude of 70%, similar to previously reported values based on analyses of human meioses. However, GC-biased gene conversion explains only a fraction of the observed difference in GC content between palindrome arms and flanking sequence, suggesting that additional factors are required to explain elevated GC content in palindrome arms. This work supports a greater than 2:1 preference for GC bases over AT bases during gene conversion, and demonstrates that the evolution and composition of mammalian sex chromosome palindromes is strongly influenced by GC-biased gene conversion.


title: Transcriptomic Changes During Stage Progression of Mycosis Fungoides

abstract:
BackgroundMycosis fungoides (MF) is the most common cutaneous T cell lymphoma, which in the early patch/plaque stages runs an indolent course. However, ~25% of MF patients develop skin tumors, a hallmark of progression to the advanced stage and associated with high mortality. The mechanisms involved in stage progression are poorly elucidated.

MethodsWe performed whole-transcriptome and whole-exome sequencing of malignant MF cells from skin biopsies obtained by laser-capture microdissection. We compared three types of MF lesions: early-stage plaques (ESP, n=12), and plaques and tumors from patients in late-stage disease (late-stage plaques, LSP, n=10, and tumors, TMR, n=15). Gene Ontology (GO) and KEGG analysis were used to determine pathway changes specific for different lesions which we linked to the recurrent somatic mutations overrepresented in MF tumors.

ResultsThe key upregulated pathways during stage progression were those related to cell proliferation and survival (MEK/ERK, Akt-mTOR), Th2/Th9 signaling (IL4, STAT3, STAT5, STAT6), meiomitosis (CT45A1, CT45A3, STAG3, GTSF1, and REC8) and DNA repair (PARP1, MYCN, OGG1). Principal coordinate clustering of the transcriptome revealed extensive gene expression differences between early (ESP) and advanced-stage lesions (LSP and TMR). LSP and TMR showed remarkable similarities at the level of the transcriptome, which we interpreted as evidence of cell percolation between lesions via hematogenous self-seeding.

ConclusionStage progression in MF is associated with Th2/Th9 polarization of malignant cells, activation of proliferation, survival, as well as increased genomic instability. Global transcriptomic changes in multiple lesions are probably caused by hematogenous cell percolation between discrete skin lesions.


title: Dual-axis Volta phase plate cryo-electron tomography of Ebola virus-like particles reveals actin-VP40 interactions

abstract:
Cryo-electron tomography (cryo-ET) is a pivotal imaging technique for studying the structure of pleomorphic enveloped viruses and their interactions with the host at native conditions. Owing to the limited tilting range of samples with a slab geometry, electron tomograms suffer from so-called missing wedge information in Fourier space. In dual-axis cryo-ET, two tomograms reconstructed from orthogonally oriented tilt series are combined into a tomogram with improved resolution as the missing wedge information is reduced to a pyramid. Volta phase plate (VPP) allows to perform in-focus cryo-ET with high contrast transfer at low-resolution frequencies and thus its application may improve the quality of dual-axis tomograms. Here, we compare dual-axis cryo-ET with and without VPP on Ebola virus-like particles to visualize and segment viral and host cell proteins within the membrane-enveloped filamentous particles. Dual-axis VPP cryo-ET reduces the missing wedge information and ray artifacts arising from the weighted back-projection during tomogram reconstruction, thereby minimizing ambiguity in the analysis of crowded environments and facilitating 3D segmentation. We show that dual-axis VPP tomograms provide a comprehensive description of macromolecular organizations such as nucleocapsid assembly states, the distribution of glycoproteins on the viral envelope and asymmetric arrangements of the VP40 layer in non-filamentous regions of virus-like particles. Our data reveal actin filaments within virus-like particles in close proximity to the viral VP40 scaffold, suggesting a direct interaction between VP40 and actin filaments. Dual-axis VPP cryo-ET provides more complete 3D information at high contrast and allows for better interpretation of macromolecule interactions and pleomorphic organizations.

HighlightsO_LIVolta phase plate dual-axis cryo-electron tomography provides high contrast tomography data with reduced back-projection ray artifacts and missing wedge information in Fourier space
C_LIO_LIVolta phase plate dual-axis cryo-electron tomography facilitates interpretation of protein-membrane interactions
C_LIO_LIVolta phase plate dual-axis cryo-electron tomography reduces ambiguity in manual 3D rendering and markedly improves 3D isosurface modeling
C_LIO_LIEbola virus-like particles contain actin filaments in close proximity to the VP40 layer
C_LI

category: biophysics


title: Integrin-dependent migratory switches regulate the translocation of Toxoplasma-infected dendritic cells across brain endothelial monolayers

abstract: Multiple cellular processes, such as immune responses and cancer cell metastasis, crucially depend on interconvertible migration modes. However, knowledge is scarce on how infectious agents impact the processes of cell adhesion and migration at restrictive biological barriers. In extracellular matrix, dendritic cells (DCs) infected by the obligate intracellular protozoan Toxoplasma gondii undergo mesenchymal-to-amoeboid transition (MAT) for rapid integrin-independent migration. Here, in a cellular model of the blood-brain barrier, we report that parasitised DCs adhere to polarised endothelium and shift to integrin-dependent motility, accompanied by elevated transendothelial migration (TEM). Upon contact with endothelium, parasitised DCs dramatically reduced velocities and adhered under both static and shear stress conditions, thereby obliterating the infection-induced amoeboid motility displayed in collagen matrix. The motility of adherent parasitised DCs on endothelial monolayers was restored by blockade of {beta}1 and {beta}2 integrins or ICAM-1, which conversely reduced motility on collagen-coated surfaces. Moreover, parasitised DCs exhibited enhanced translocation across highly polarised primary murine brain endothelial cell monolayers. Blockade of {beta}1, {beta}2 integrins, ICAM-1 and PECAM-1 reduced TEM frequencies. Finally, gene silencing of the pan-integrin-cytoskeleton linker talin (Tln1) or of {beta}1 integrin (Itgb1) in primary DCs resulted in increased motility on endothelium and decreased TEM. Adding to the paradigms of leukocyte diapedesis, the findings provide novel insights in how an intracellular pathogen impacts the migratory plasticity of leukocytes in response to the cellular environment, to promote infection-related dissemination.

category: immunology


title: OXSR1 inhibits inflammasome activation by limiting potassium efflux during mycobacterial infection

abstract: Pathogenic mycobacteria inhibit inflammasome activation as part of their pathogenesis. While it is known that potassium efflux is a trigger for inflammasome activation, the interaction between mycobacterial infection, potassium efflux and inflammasome activation has not been investigated. Here we use Mycobacterium marinum infection of zebrafish embryos and Mycobacterium tuberculosis of human THP-1 cells to demonstrate that pathogenic mycobacteria upregulate the host WNK signalling pathway kinases SPAK and OXSR1 which control intracellular potassium balance. We show that genetic depletion or inhibition of OXSR1 decreases bacterial burden and intracellular potassium levels. The protective effects of OXSR1 depletion are mediated by NLRP3 inflammasome activation and are dependent on caspase-mediated release of IL-1{beta} and the downstream activation of protective TNF-. The elucidation of this druggable pathway to potentiate inflammasome activation provides a new avenue for the development of host-directed therapies against intracellular infections.


title: The Magnaporthe oryzae MAP kinase Pmk1 regulates polycomb repressive complex 2 to reprogram genes expression for biotrophic growth

abstract:
Biotrophic and hemibiotrophic fungi have evolved the ability to colonize living plant cells, but how they establish biotrophic growth by remodeling gene expression is poorly understood. By using in planta invasive hyphae (IH) of Magnaporthe oryzae to perform an integrated Chromatin immunoprecipitation sequencing (ChIPseq) and RNA-seq analysis, combining with biological and cellular analyses, we found Polycomb repressive complex 2 (PRC2)-mediated epigenetic repression plays a key role in regulating biotrophic growth. ChIPseq for biotrophic IH samples identified 1701 PRC2 target genes. RNA-seq analysis showed that expression of 462 PRC2 target genes were up-regulated in the {Delta}suz12 mutant, while 82 were down-regulated, indicating a major role of PRC2 in gene repression of IH. During biotrophic growth, PRC2 repressed fungal cell wall synthesis genes and extracellular enzyme genes required for penetration, and secondary metabolites biosynthesis genes required for necrotrophic growth. A great number of effector-encoding genes were repressed by PRC2, which were highly expressed during penetration stage, suggesting PRC2 coordinates biotrophic growth by regulating effector suppression for immune evasion. This regulation was finely coordinated by Pmk1, through regulating phosphorylation, nuclear localization and protein abundance of Suz12. Our results indicate that the Pmk1-PRC2 regulatory module is required for gene remodeling to facilitate biotrophic growth in M. oryzae.

IMPORTANCEBiotrophic and hemibiotrophic fungi establish a biotrophic stage for infection in host cells. For example, M. oryzae forms appressoria to penetrate host cell and establish a biotrophic growth stage for infection. How gene expression patterns are elaborately controlled for fungal biotrophic growth is largely unknown. In this study, we found that, the PRC2-mediated H3K27me3 repressed fungal penetration-required cell wall synthesis genes and extracellular enzyme genes, and necrotrophic growth-required secondary metabolites biosynthesis genes for biotrophic growth. Interestingly, a great number of effector-encoding genes were also repressed by PRC2 at biotrophic stage, which were highly expressed at penetration stage, suggesting PRC2 coordinates biotrophic growth by regulating effector suppression for immune evasion. The PRC2-mediated epigenetic repression is therefore required for the gene expression remodeling during fungal infection. This regulation was finely coordinated by Pmk1, through regulating nuclear localization and protein abundance of the PRC2 component Suz12.


title: Homologs of circadian clock proteins impact the metabolic switch between light and dark growth in the cyanobacterium Synechocystis sp. PCC 6803

abstract: The putative circadian clock system of the facultative heterotrophic cyanobacterial strain Synechocystis sp. PCC 6803 comprises the following three Kai-based systems: a KaiABC-based potential oscillator that is linked to the SasA-RpaA two-component output pathway and two additional KaiBC systems without a cognate KaiA component. Mutants lacking the genes encoding the KaiAB1C1 components or the response regulator RpaA show reduced growth in light/dark cycles and do not show heterotrophic growth in the dark. In the present study, the effect of these mutations on central metabolism was analyzed by targeted and nontargeted metabolite profiling. The strongest metabolic changes were observed in the dark in {Delta}rpaA and, to a lesser extent, in the {Delta}kaiAB1C1 mutant. These observations included the overaccumulation of 2-phosphoglycolate, which correlated with the overaccumulation of the RbcL subunit in the mutants, and taken together, these data suggest enhanced RubisCO activity in the dark. The imbalanced carbon metabolism in the {Delta}rpaA mutant extended to the pyruvate family of amino acids, which showed increased accumulation in the dark. Hence, the deletion of the response regulator rpaA had a more pronounced effect on metabolism than the deletion of the kai genes. The larger impact of the rpaA mutation is in agreement with previous transcriptomic analyses and likely relates to a KaiAB1C1-independent function as a transcription factor. Collectively, our data demonstrate an important role of homologs of clock proteins in Synechocystis for balanced carbon and nitrogen metabolism during light-to-dark transitions.


title: Inference of Morphogen Gradient Precision from Molecular Noise Data

abstract: During development, morphogen gradients provide spatial information for tissue patterning. Gradients and readout mechanisms are inevitably variable, yet the resulting patterns are strikingly precise. Measurement limitations currently preclude precise detection of morphogen gradients over long distances. Here, we develop a new formalism to estimate gradient precision along the entire patterning axis from measurements close to the source. Using numerical simulations, we infer gradient variability from measured molecular noise levels in morphogen production, decay, and diffusion. The predicted precision is much higher than previously measured--precise enough to allow even single gradients to define the central progenitor boundaries during neural tube development. Finally, we show that the patterning mechanism is optimized for precise progenitor cell numbers, rather than precise boundary positions, as the progenitor domain size is particularly robust to gradient alterations. We conclude that single gradients can yield the observed developmental precision, which provides new prospects for tissue engineering.


title: Species coexistence and temporal environmental fluctuations: a quantitative comparison between stochastic and seasonal variations.

abstract: Temporal environmental variations may promote diversity in communities of competing populations. Here we compare the effect of environmental stochasticity with the effect of periodic (e.g., seasonal) cycles, using analytic solutions and individual-based Monte-Carlo simulations. Even when stochasticity facilitates coexistence it still allows for rare sequences of bad years that may drive a population to extinction, therefore the stabilizing effect of periodic variations is stronger. Correspondingly, the mean time to extinction grows exponentially with community size in periodic environment and switch to power-law dependence under stochastic fluctuations. On the other hand, the number of temporal niches in periodic environment is typically lower, so as diversity increases stochastic temporal variations may support higher species richness.


title: Evolutionary rates of testes-expressed genes differ between monogamous and promiscuous Peromyscus species

abstract: Reproductive proteins, including those expressed in the testes, are among the fastest evolving proteins across the tree of life. Sexual selection on traits involved in sperm competition is thought to be a primary driver of testes gene evolution and is expected to differ between promiscuous and monogamous species due to intense competition between males to fertilize females in promiscuous lineages and lack thereof in monogamous ones. Here, we employ the rodent genus Peromyscus as a model to explore differences in evolutionary rates between testes-expressed genes in monogamous and promiscuous species. We find candidate genes that may be associated with increased sperm production in promiscuous species and gene ontology categories that show patterns of molecular convergence associated with phenotypic convergence in independently evolved monogamous species. Overall, our results highlight the possible molecular consequences of differences in mating system, likely due to differences in selective pressures.

category: evolutionary biology


title: Evolutionary rates of testes-expressed genes differ in monogamous and promiscuous Peromyscus species


title: Novel neuroanatomical integration and scaling define avian brain shape evolution and development

abstract:
How do large and unique brains evolve? Historically, comparative neuroanatomical studies have attributed the evolutionary genesis of highly encephalized brains to deviations along, as well as from, conserved scaling relationships among brain regions. However, the relative contributions of these concerted (integrated) and mosaic (modular) processes as drivers of brain evolution remain unclear, especially in non-mammalian groups. While proportional brain sizes have been the predominant metric used to characterize brain morphology to date, we perform a high-density geometric morphometric analysis on the encephalized brains of crown birds (Neornithes or Aves) compared to their stem taxa--the non-avialan coelurosaurian dinosaurs. When analyzed together with developmental neuroanatomical data of model archosaurs (Gallus, Alligator), crown birds exhibit a distinct allometric relationship that dictates their brain evolution and development. Furthermore, analyses by neuroanatomical regions reveal that the acquisition of this derived shape-to-size scaling relationship occurred in a mosaic pattern, where the  avian-grade optic lobe and cerebellum evolved first among non-avialan dinosaurs, followed by major changes to the evolutionary and developmental dynamics of cerebrum shape after the origin of Avialae.

Notably, the brain of crown birds is a more integrated structure than non-avialan archosaurs, implying that diversification of brain morphologies within Neornithes proceeded in a more coordinated manner, perhaps due to spatial constraints and abbreviated growth period. Collectively, these patterns demonstrate a plurality in evolutionary processes that generate encephalized brains in archosaurs and across vertebrates.


title: FMRP bridges R-loops and DHX9 through direct interactions

abstract: Fragile X Syndrome (FXS) occurs when mutations in the FMR1 gene cause the absence or dysfunction of FMRP, known mainly as a translation repressor. We recently showed that FXS cells suffer genome-wide DNA double-strand breaks near R-loops under replication stress. The expression of FMRP, and not an FMRP-I304N mutant of the K-homology 2 RNA-binding domain, suppresses the R-loop-induced DNA breakage. These observations led us to hypothesize that FMRP safeguards the genome through promotion of R-loop detection and/or resolution. Here, we demonstrate that FMRP directly binds R-loops through multivalent interactions between the carboxy-terminal intrinsically disordered region and the R-loop sub-structures. We also show that the amino-terminal folded domain of FMRP directly binds DHX9, an R-loop resolvase, in a KH2-dependent manner. The FMRP-DHX9 interaction is recapitulated by co-immunoprecipitation in human cells. Our findings are consistent with a model in which FMRP recruits DHX9 to R-loop forming sites by bridging their interaction through its amino- and carboxy-termini, respectively.

category: biochemistry


title: IL-7 is essential for accumulation of antigen-specific CD8 T cells and to generate clonotype-specific effector responses during airway influenza/A infection.

abstract:
Airborne diseases are the leading cause of infectious disease-related deaths in the world. In particular, the influenza virus activates a network of immune cells that leads to clearance or an overzealous response that can be fatal. Tight regulation of the cytokines that enable proper activation and function of immune cells is necessary to clear infections efficiently while minimizing damage to the host. Interleukin-7 (IL-7) is a cytokine known for its importance in T cell development and survival. How IL-7 shapes CD8 T cell responses during an acute viral infection is less understood. We had previously shown that IL-7 signaling deficient mice have reduced accumulation of influenza-specific CD8 T cells following infection. We sought to determine whether IL-7 affects early CD8 T cell expansion in the mediastinal lymph node and effector function in the lungs. Using IL-7R signaling deficient mice, we show that IL-7 is required for a normal sized mediastinal lymph node and the early clonal expansion of antigen-specific CD8 T cells therein. Bone marrow chimeric models and adoptive transfer of transgenic TCR CD8 T cells reveal a cell-intrinsic role for IL-7 in the accumulation of NP366-374 and PA224-233-specific CD8 T cells. We also found that IL-7 dictates terminal differentiation, degranulation and cytokine production in PA224-233-specific but not NP366-374-specific CD8 T cells. We further demonstrate that IL-7 is induced in the lung tissue by viral infection and we characterize multiple cellular sources that contribute to IL-7 production. Drugs that manipulate IL-7 signaling are currently under clinical trial for multiple conditions. Our findings on IL-7 and its effects on lower respiratory diseases will be important for expanding the utility of these therapeutics.

Author SummaryInterleukin-7 plays an important role in development of immune cells such as lymphocytes. In recent years, its role in the immune system has been expanded beyond the development of immune cells to include revitalizing of lymphocytes during tumor and chronic viral response. We show here that IL-7 is required for accumulation and function of specialized lymphocytes in the lungs during an acute influenza infection.


title: Suppression of Fatty Acid Oxidation by Thioesterase Superfamily Member 2 in Skeletal Muscle Promotes Hepatic Steatosis and Insulin Resistance

abstract: Thioesterase superfamily member 2 (Them2) is highly expressed in oxidative tissues where it hydrolyzes long chain fatty acyl-CoA esters to free fatty acids and CoA. Although mice globally lacking Them2 (Them2-/-) are protected against diet-induced obesity, insulin resistance and hepatic steatosis, liver-specific Them2-/- mice remain susceptible. To explore the contribution of Them2 in extrahepatic tissues, we created mice with Them2 deleted in skeletal muscle (S-Them2-/-), cardiac muscle (C-Them2-/-) or adipose tissue (A-Them2-/-). When fed a high-fat diet, S-Them2-/- but not C-Them2-/- or A-Them2-/- mice exhibited reduced weight gain. Only S-Them2-/- mice exhibited improved glucose homeostasis together with improved insulin sensitivity in skeletal muscle. Increased rates of fatty acid oxidation in skeletal muscle of S-Them2-/- mice were reflected in alterations in skeletal muscle metabolites, including short chain fatty acids, branched chain amino acids and the pentose phosphate pathway. Protection from diet-induced hepatic steatosis in S-Them2-/- mice was attributable to increased VLDL triglyceride secretion rates in support of demands of increased muscle fatty acid utilization. These results reveal a key role for skeletal muscle Them2 in the pathogenesis of diet-induced obesity, insulin resistance and hepatic steatosis.


title: LIN37-DREAM Prevents DNA End Resection and Homologous Recombination at DNA Double Strand Breaks in Quiescent Cells

abstract: DNA double strand break (DSB) repair by homologous recombination (HR) is thought to be restricted to the S- and G2- phases of the cell cycle in part due to 53BP1 antagonizing DNA end resection in G1-phase and non-cycling quiescent (G0) cells. Here, we show that LIN37, a component of the DREAM transcriptional repressor, functions in a 53BP1-independent manner to prevent DNA end resection and HR in G0 cells. Loss of LIN37 leads to expression of HR proteins, including BRCA1, BRCA2, PALB2 and RAD51, and DNA end resection in G0 cells even in the presence of 53BP1. In contrast to 53BP1-deficiency, DNA end resection in LIN37-deficient G0 cells depends on BRCA1 and leads to RAD51 filament formation and HR. LIN37 is not required to protect DNA ends in cycling cells at G1-phase. Thus, LIN37 regulates a novel 53BP1-independent cell phase-specific DNA end protection pathway that functions uniquely in quiescent cells.


title: Assessment of the pollution incident performance of water and sewerage companies in England

abstract: Comparison of water and sewerage company pollution performance in relation to severity, frequency and self-reporting of pollution incidents is made difficult by differences in environmental and operational conditions. In England, the deterioration in pollution incident performance across the water industry, makes it important to investigate common trends that could be addressed at national and regional levels. Yet, to date there has been no external peer-reviewed analysis of national pollution incident data in England. This project aimed to analyse available pollution incident data to assess and compare the performance of water and sewerage companies in England. Results indicated that there was significant variation in pollution incident numbers and the severity of the impact on the water environment for different asset types (operational property of the water and sewerage company such as a sewage treatment works). Increasing numbers of pollution incidents from pumping stations and sewage treatment works, were largely responsible for overall increases in pollution incidents. The highest increase in pollution incidents in 2019, was observed from pumping stations. Variation was evident in company self-reporting percentages across asset types. There were significant positive relationships between the self-reporting percentages of pollution incidents and total numbers of reported pollution incidents per 10,000 km sewer length for pumping stations and sewage treatment works. These results indicate that in at least these asset types, an estimated 5% of pollution incidents could go unreported, if not self-reported by the company. Pollution events not reported quickly by companies, can lead to more severe impacts to the water environment so rapid and consistent reporting of incidents is crucial for limiting damage. The results have significance for the water industry internationally, because the issues presented here are not restricted to England. Whilst this research highlighted a number of key areas for more detailed analysis, in the short-term, research should focus on investigating best practice for reporting pollution incidents. It is important to get an accurate baseline of the number of pollution incidents and whether a proportion are currently going unreported. This research should seek to aid the standardisation of reporting practice across the water industry.


title: Tempo and mode of gene expression evolution in the brain across Primates

abstract: Primate evolution has led to a remarkable diversity of behavioral specializations and pronounced brain size variation among species 1,2. Gene expression provides a promising opportunity for studying the molecular basis of brain evolution, but it has been explored in very few primate species to date e.g. 3,4. To understand the landscape of gene expression evolution across the primate lineage, we generated and analyzed RNA-Seq data from four brain regions in an unprecedented eighteen species. Here we show a remarkable level of variation in gene expression among hominid species, including humans and chimpanzees, despite their relatively recent divergence time from other primates. We found that individual genes display a wide range of expression dynamics across evolutionary time reflective of the diverse selection pressures acting on genes within primate brain tissue. Using our sample that represents an unprecedented 190-fold difference in primate brain size, we identified genes with variation in expression most correlated with brain size and found several with signals of positive selection in their regulatory regions. Our study extensively broadens the context of what is known about the molecular evolution of the brain across primates and identifies novel candidate genes for study of genetic regulation of brain development and evolution.


title: Metabotropic group II glutamate receptors in the basolateral amygdala mediate cue-triggered increases in incentive motivation

abstract:
RationaleReward-associated cues can trigger incentive motivation for reward and invigorate reward-seeking behaviour via Pavlovian-to-instrumental transfer (PIT). Glutamate signaling within the basolateral amygdala (BLA) modulates cue-triggered increases in incentive motivation. However, the role of BLA metabotropic group II glutamate (mGlu2/3) receptors is largely unknown.

ObjectivesIn Experiment 1, we characterized cue-triggered increases in incentive motivation for water reward using the PIT paradigm. In Experiment 2, we assessed the influence of intra-BLA microinjections of the mGlu2/3 receptor agonist LY379268 on this effect.

MethodsWater-restricted male Sprague-Dawley rats learned to press a lever for water. Separately, they learned to associate one of two auditory cues with free water. On test days, rats could lever press under extinction conditions (no water), with intermittent, non-contingent CS+ and CS- presentations. In Experiment 1, rats were tested under baseline conditions. In Experiment 2, rats received intra-BLA microinjections of LY379268 (0, 3 and 6 g/hemisphere) before testing.

ResultsAcross experiments, CS+, but not CS- presentations increased water-associated lever pressing during testing, even though responding was reinforced neither by water nor the CS+. Intra-BLA LY379268 abolished both CS+ potentiated pressing on the water-associated lever and CS+ evoked conditioned approach to the site of water delivery. LY379268 did not influence locomotion or instrumental and Pavlovian response rates during intervals between CS presentations or during the CS-, indicating no motor effects.

ConclusionsmGlu2/3 receptor activity in the BLA mediates CS-triggered potentiation of incentive motivation for reward, suppressing both CS-induced increases in instrumental pursuit of the reward and anticipatory approach behaviour.

category: neuroscience


title: The non-redundant nature of the Axin2 regulatory network in the canonical Wnt signaling pathway

abstract:
Axin is one of two essential scaffolds in the canonical Wnt pathway that converts signals at the plasma membrane to signals inhibiting the degradation of {beta}-catenin, leading to its accumulation and specific gene activation. In vertebrates there are two forms of Axin, Axin1 and Axin2, which are similar at the protein level and genetically redundant. We show here that differential regulation of the two genes on the transcriptional and proteostatic level confers robustness and differential responsiveness that can be used in tissue specific regulation. Such subtle features may distinguish other redundant gene pairs that are commonly found in vertebrates through gene knockout experiments.

Significance StatementThe mystery of two functionally redundant Axin genes in all vertebrates are can now be explained by the demonstration that they form a nested proteostatic and transcriptional feedback system that confers regulatory options in different developmental settings, a form of dynamic versatility that may explain the widespread occurrence of closely related seemingly redundant genes with similar functions.


title: Translational Proteomics for Transfusion Medicine: Resolution of the IVIG Proteomes of Different Geographically Sourced and Prepared IVIG Immunotherapies.

abstract: Intravenous Immunoglobulins (IVIGs) are prepared from thousands of donor plasmas and as a result comprise an extreme broad mix and depth of Antibody (Ab) specificities. IVIG formulations available in Australia are from both local and overseas donor sources and extracted using a variety of purification methods and immunoglobulin purities, with the Australia-derived and prepared IVIG listed at >98% and the overseas-derived preparations at [≥] 95%. Because of these differences it was predicted that the formulations might individually vary in composition and that together with obvious genetic and geographic antigenic (Ag) environment differences may result in notable variability between formulations. Hence a focussed comparative proteomic profiling of IVIG formulations was undertaken to identify notable similarities and differences across products. Comparisons between formulations did reveal marked qualitative differences in 2D-gel Antibody profiling that included parameters of isoelectric charge (pI), as well as immunoglobulin (Ig) monomer to dimer ratio variability between products, including high molecular weight (MW) immunoglobulin multimers for some. These notable differences were in part quite likely a product of the respective purification methods used, and capacity to select (or de-select) for antibodies of such different properties. Furthermore, for identification of non-Ig proteins carried over from plasma through purifications Mass spectrometry was performed. This identified a few such ancillary proteins, and their identities, in general, differed between formulations. Proteins detected included the most abundant protein of plasma, albumin, as well as other mostly large and abundant proteins; RAG1 - V(D)J recombination activating protein1, gelsolin, complement Factor-B, serotransferrin, tetranectin, NADH ubiquinone oxidoreductase, caspase 3 and VEGFR1. An alternate strategy used commercial Multiplex xMAP assay to detect cytokines, which are small and present in plasma at trace but highly active quantities. This revealed various different cytokine profiles across the formulations studied. The identification of additional proteins, and especially cytokines in IVIGs, is particularly notable, and the positive, negative or null biological relevance for clinical use, needs resolution. Collectively these findings reveal marked differences between Australian and overseas-derived (non-Australian) IVIGs in immunoglobulin composition and biochemical characteristics, and presence of additional carry-over proteins from plasma. These findings prompt the need for further evaluation of the micro-compositions of individual formulations. Perhaps detailed mining and improved comparative understanding of each IVIG formulation, may enable highly tailored and strategic clinical use of certain formulations that are personalised best-fit treatments for particular conditions. Such as in treatment of a neuropathy, as compared to another formulation, more suited for treating a particular infectious disease. The most salient and overarching study conclusion is need for caution in attributing equivalence across IVIGs.


title: Direct observation of aggregate-triggered selective autophagy

abstract: Degradation of aggregates by selective autophagy is important as damaged proteins may impose a threat to cellular homeostasis. Although the core components of the autophagy machinery are well-characterized, the spatiotemporal regulation of many selective autophagy processes, including aggrephagy, remains largely unexplored. Furthermore, because most live-cell imaging studies have so far focused on starvation-induced autophagy, little is known about the dynamics of aggrephagy. Here, we describe the development and application of the mKeima-PIM assay, which enables live-cell observation of autophagic turnover and degradation of inducible protein aggregates in conjunction with key autophagy players. This allowed us to quantify the relative timing and duration of different steps of aggrephagy and revealed the short-lived nature of the autophagosome. The assay furthermore showed the spatial distribution of omegasome formation, highlighting that autophagy initiation is directly instructed by the cargo. Moreover, we found that nascent autophagosomes mostly remain immobile until acidification occurs. Thus, our assay provides new insights into the spatiotemporal regulation and dynamics of aggrephagy.


title: Single Cell Chemical Proteomics (SCCP) Interrogates the Timing and Heterogeneity of Cancer Cell Commitment to Death

abstract: Chemical proteomics studies the effects of drugs upon cellular proteome. Due to the complexity and diversity of tumors, the response of cancer cells to drugs is also heterogeneous, and thus proteome analysis at single cell level is needed. Here we demonstrate that single cell proteomics techniques have become quantitative enough to tackle the drug effects on the target proteins, enabling single cell chemical proteomics (SCCP). Using SCCP, we studied here the time-resolved response of individual adenocarcinoma A549 cells to anticancer drugs methotrexate, camptothecin and tomudex, revealing the early emergence of cellular subpopulations committed and uncommitted to death. As a novel and useful approach to exploring the heterogeneous response to drugs of cancer cells, SCCP may prove to be a "killer application" for single cell proteomics.

category: cancer biology


title: Physical Characterization of Triolein and Implications for Its Role in Lipid Droplet Biogenesis

abstract: Lipid droplets (LDs) are neutral lipid storing organelles surrounded by a phospholipid (PL) monolayer. At present, how LDs are formed in the endoplasmic reticulum (ER) bilayer is poorly understood. In this study, we present a revised all-atom (AA) triolein (TG) model, the main constituent of the LD core, and characterize its properties in a bilayer membrane to demonstrate the implications of its behavior in LD biogenesis. In bilayer simulations, TG resides at the surface, adopting PL-like conformations (denoted in this work as SURF-TG). Free energy sampling simulation results estimate the barrier for TG relocating from the bilayer surface to the bilayer center to be [~]2 kcal/mol in the absence of an oil lens. SURF-TG is able to modulate membrane properties by increasing PL ordering, decreasing bending modulus, and creating local negative curvature. The other neutral lipid, dioleoyl-glycerol (DAG), also reduces the membrane bending modulus and populates the negative curvature regions. A phenomenological coarse-grained (CG) model is also developed to observe larger scale SURF-TG-mediated membrane deformation. The CG simulations confirm that TG nucleates between the bilayer leaflets at a critical concentration when SURF-TG is evenly distributed. However, when one monolayer contains more SURF-TG, the membrane bends toward the other leaflet, followed by TG nucleation if a concentration is higher than the critical threshold. The central conclusion of this study is that SURF-TG is a negative curvature inducer, as well as a membrane modulator. To this end, a model is proposed in which the accumulation of SURF-TG in the luminal leaflet bends the ER bilayer toward the cytosolic side, followed by TG nucleation.


title: Unmethylated Regions Encompass The Functional Space Within The Maize Genome

abstract: Delineating the functional space within genomes has been a long-standing goal shared among geneticists, molecular biologists, and genome scientists. The genome of Zea mays (maize) has served as a model for locating functional elements within the gene-distal intergenic space. A recent development has been the discovery and use of accessible chromatin as a proxy for functional regulatory elements. However, the idea has recently arisen that DNA methylation data could supplement the use of accessible chromatin data for homing in on regulatory regions. Here, I test the robustness of using DNA methylation as a proxy for functional space. I find that CHG methylation can be non-arbitrarily partitioned into hypo-methylated and hyper-methylated regions. Hypo-methylated CHG regions are stable across development and contain nearly all accessible chromatin. Note: changes that will be made in version 2: expand introduction; expand discussion; add additional analyses; expand methods; link to github scripts.


title: Inhibition is the hallmark of CA3 intracellular dynamics around awake ripples

abstract: Hippocampal ripples are transient population bursts that structure cortico-hippocampal communication and play a central role in memory processing. However, the mechanisms controlling ripple initiation in behaving animals remain poorly understood. Here we combine multisite extracellular and whole cell recordings in awake mice to contrast the brain state and ripple modulation of subthreshold dynamics across hippocampal subfields. We find that entorhinal input to DG exhibits UP and DOWN dynamics with ripples occurring exclusively in UP states. While elevated cortical input in UP states generates depolarization in DG and CA1, it produces persistent hyperpolarization in CA3 neurons. Furthermore, growing inhibition is evident in CA3 throughout the course of the ripple buildup, while DG and CA1 neurons exhibit depolarization transients 100 ms before and during ripples. These observations highlight the importance of CA3 inhibition for ripple generation, while pre-ripple responses indicate a long and orchestrated ripple initiation process in the awake state.


title: Object motion kinematics influence both feedforward and feedback motor responses during virtual catching

abstract: An important window into sensorimotor function is how we catch moving objects. Studies that examined catching of free-falling objects report that the timing of the motor response is independent of the momentum of the projectile, whereas the motor response amplitude scales with projectile momentum. However, this pattern may not be a general strategy of catching since objects accelerate under gravity in a characteristic manner (unlike object motion in the horizontal plane) and the human visual motion-processing system is not adept at encoding acceleration. Accordingly, we developed a new experimental paradigm using a robotic manipulandum and augmented reality where participants stabilized against the impact of a virtual object moving at constant velocity in the horizontal plane. Participants needed to apply an impulse that mirrored the object momentum to bring it to rest and received explicit feedback on their performance. In different blocks, object momentum was varied by an increase in its speed or mass. In contrast to previous reports on free falling objects, we observed that increasing object speed caused earlier onset of arm muscle activity and limb force relative to the impending time to contact. Also, arm force increased as a function of target momentum linked to changes in speed or mass. Our results demonstrate velocity-dependent timing to catch objects and a complex pattern of scaling to momentum.


title: Computational mechanisms of osmoregulation: a reinforcement learning model for sodium appetite

abstract:
Homeostatic control with oral nutrient intake is a vital complex system involving the orderly interactions between the external and internal senses, behavioral control, and reward learning. Sodium appetite is a representative system and has been intensively investigated in animal models of homeostatic systems and oral nutrient intake. However, the system-level mechanisms for regulating sodium intake behavior and homeostatic control remain unclear.

In the current study, we attempted to provide a mechanistic understanding of sodium appetite behavior by using a computational model, the homeostatic reinforcement learning model, in which homeostatic behaviors are interpreted as reinforcement learning processes. Through simulation experiments, we confirmed that our homeostatic reinforcement learning model successfully reproduced homeostatic behaviors by regulating sodium appetite. These behaviors include the approach and avoidance behaviors to sodium according to the internal states of individuals. In addition, based on the assumption that the sense of taste is a predictor of changes in the internal state, the homeostatic reinforcement learning model successfully reproduced the previous paradoxical observations of the intragastric infusion test, which cannot be explained by the classical drive reduction theory. Moreover, we extended the homeostatic reinforcement learning model to multi-modal data, and successfully reproduced the behavioral tests in which water and sodium appetite were mediated by each other. Finally, through an experimental simulation of chemical manipulation in a specific neural population in the brain stem, we proposed a testable hypothesis for the function of neural circuits involving sodium appetite behavior.

The study results support the idea that osmoregulation via sodium appetitive behavior can be understood as a reinforcement learning process and provide a mechanistic explanation for the underlying neural mechanisms of sodium appetite and homeostatic behavior.

Author SummaryThe taste of high-concentration saltwater is rewarding during sodium depletion, while it is aversive in a sodium sufficient state. This "sodium appetite" is a clear manifestation of homeostasis maintenance and proper action selection in animals, reflecting the internal environment. To reveal the computational mechanism of this property, we applied a machine learning model, in which homeostatic stability is a reward and the goal is to maximize the sum of the reward, and simulated animal behavioral experiments. The results suggest that the mechanism of sodium-appetite behavior is based on the machine learning model. Specifically, by replicating the results of neural circuit manipulation, which controls sodium appetite, an algorithm in which the function of a neural population affects sodium appetite behaviors was proposed. Our results provide a fundamental computational model for a mechanism by a function of a neural cell type to regulate animal behavior. More generally, this study can be fundamental to understanding the computational process of decision making reflecting the internal environment.


title: The Backbone Network of Dynamic FunctionalConnectivity

abstract: 1.Temporal networks have become increasingly pervasive in many real-world applications, including the functional connectivity analysis of spatially separated regions of the brain. A major challenge in analysis of such networks is the identification of noise confounds, which introduce temporal ties that are non-essential, or links that are formed by chance due to local properties of the nodes. Several approaches have been suggested in the past for static networks or temporal networks with binary weights for extracting significant ties whose likelihood cannot be reduced to the local properties of the nodes. In this work, we propose a data-driven procedure to reveal the irreducible ties in dynamic functional connectivity of resting state fRMI data with continuous weights. This framework includes a null model that estimates the latent characteristics of the distributions of temporal links through optimization, followed by a statistical test to filter the links whose formation can be reduced to the activities and local properties of their interacting nodes. We demonstrate the benefits of this approach by applying it to a resting state fMRI dataset, and provide further discussion on various aspects and advantages of it.


title: The Backbone Network of Dynamic Functional Connectivity


title: Segment number threshold determines juvenile onset of germline cluster proliferation in Platynereis dumerilii

abstract: Many animals rely on sexual reproduction to propagate by using gametes (oocytes and sperm). Development of sexual characters and generation of gametes are tightly coupled with the growth of an organism. Platynereis dumerilii is a marine segmented worm which has been used to study germline development and gametogenesis. Platynereis has 4 Primordial Germ Cells (PGCs) that arise early in development and these cells are thought to give rise to germ cell clusters found across the body in the juvenile worms. The germ cell clusters eventually form the gametes. The stages of germline development and how the 4 PGCs become the numerous germ cell clusters are not well-documented in the juvenile stages. Platynereis, like other segmented worms, grows by adding new segments at its posterior end. The number of segments generally reflect the growth state of the worms and therefore is a useful and easily measurable growth state metric. To understand how growth correlates with development and gametogenesis, we investigated germline development across several developmental stages using germline/multipotency markers. We found that segment number predicted the state of germline development and the abundance of germline clusters. Additionally, we found that keeping worms short in segment number via changing external conditions or via amputations supported segment number threshold requirement for germline development. Finally, we asked if these clusters in Platynereis play a role in regeneration (as similar free-roaming cells are observed in Hydra and planarian regeneration) and found that the clusters were not required for regeneration in Platynereis, suggesting a strictly germline nature. Overall, these molecular analyses suggest a previously unidentified developmental transition dependent on the growth state in juvenile Platynereis when germline proliferation is substantially increased.


title: Developmental Circadian Disruption Alters Placental Signaling in Mice

abstract: Circadian disruption has been largely overlooked as a developmental exposure. The placenta, a conduit between the maternal and fetal environments, may relay circadian cues to the fetus. We have previously shown that developmental chronodisruption causes visual impairment and increased retinal microglial and macrophage marker expression. Here, we investigated the impacts of environmental circadian disruption on fetal and placental outcomes in a C57BL/6J mouse (Mus musculus) model. Developmental chronodisruption had no effect on embryo count, placental weight, or fetal sex ratio. When measured with RNAseq, mice exposed to developmental circadian disruption (CD) had differential placental expression of several transcripts including Serpinf1, which encodes pigment-epithelium derived factor (PEDF). Immunofluorescence of microglia/macrophage markers, Iba1 and CD11b, also revealed significant upregulation of immune cell markers in CD-exposed placenta. Our results suggest that in utero circadian disruption enhances placental immune cell expression, potentially programming a pro-inflammatory tissue environment that increases the risk of chronic disease in adulthood.


title: Strengthened mutualistic adaptation between teosinte and its rhizosphere biota in cold climates

abstract:
O_LIWhile abiotic environments consistently shape local adaptation, the strength of local adaptation to biotic interactions may vary more. One theory, COCO (CO-evolutionary Outcomes across Conditionality), predicts it may be strongest where species experience greater stress, because stress increases fitness impacts of species interactions. For example, in plant interactions with rhizosphere biota, positive outcomes increase with stress from low soil fertility, drought and cold.
C_LIO_LITo investigate the influence of abiotic stress gradients on adaptation between plants and rhizosphere biota, we used a greenhouse common garden experiment recombining teosinte, Zea mays ssp. mexicana (wild relative of maize), and rhizosphere biota, collected across a stress gradient (elevational variation in temperature, precipitation, and nutrients).
C_LIO_LIWe found stronger local adaptation between teosinte and rhizosphere biota from colder, more stressful sites, as expected by COCO. However, biota from less stressful, warmer sites provided greater average benefits across teosinte populations. Links between plant traits and 20-element profiles of plant leaves explained fitness variation, persisted in the field, were influenced by both plants and biota, and largely reflected patterns of local adaptation.
C_LIO_LIIn sum, we uncovered greater local adaptation to biotic interactions in colder sites, and that both plants and rhizosphere biota affect the expression of plant phenotypes.
C_LI