title: Homologous or Heterologous Booster of Inactivated Vaccine Reduces SARS-CoV-2 Omicron Variant Escape from Neutralizing Antibodies

abstract: The massive and rapid transmission of SARS-CoV-2 has led to the emergence of several viral variants of concern (VOCs), with the most recent one, B.1.1.529 (Omicron), which accumulated a large number of spike mutations, raising the specter that this newly identified variant may escape from the currently available vaccines and therapeutic antibodies. Using VSV-based pseudovirus, we found that Omicron variant is markedly resistant to neutralization of sera form convalescents or individuals vaccinated by two doses of inactivated whole-virion vaccines (BBIBP-CorV). However, a homologous inactivated vaccine booster or a heterologous booster with protein subunit vaccine (ZF2001) significantly increased neutralization titers to both WT and Omicron variant. Moreover, at day 14 post the third dose, neutralizing antibody titer reduction for Omicron was less than that for convalescents or individuals who had only two doses of the vaccine, indicating that a homologous or heterologous booster can reduce the Omicron escape from neutralizing. In addition, we tested a panel of 17 SARS-CoV-2 monoclonal antibodies (mAbs). Omicron resists 7 of 8 authorized/approved mAbs, as well as most of the other mAbs targeting distinct epitopes on RBD and NTD. Taken together, our results suggest the urgency to push forward the booster vaccination to combat the emerging SARS-CoV-2 variants.


title: Assessment of chimeric antigen receptor T (CAR-T) cytotoxicity by droplet microfluidics in vitro

abstract: Chimeric antigen receptor T (CAR-T) cells are cytotoxic T cells engineered to specifically kill cancer cells expressing specific target receptor(s). Prior CAR-T efficacy tests include CAR expression analysis by qPCR or ELISA, in vitro measurement of interferon-{gamma} (IFN{gamma}) or interleukin-2 (IL-2), and xenograft models. However, the in vitro measurements did not reflect CAR-T cytotoxicity, whereas xenograft models are low throughput and costly. Here we presented a robust in vitro droplet microfluidic assay for CAR-T cytotoxicity assessment. This method not only enabled assessment of CAR-T cytotoxic activity under different fluid viscosity conditions, but also facilitated measurement of CAR-T expansion and dissection of mechanism of action via phenotype analysis in vitro. Furthermore, our data suggested that label-free cytotoxicity analysis is feasible by acquiring data before and after treatment. Hence, this study presented a novel in vitro method for assessment of cellular cytotoxicity that could potentially be applied to any cell-kill-cell experiment with varying solvent composition.

category: cancer biology


title: Excitatory effect of biphasic kHz field stimulation on CA1 pyramidal neurons in slices

abstract:
BackgroundElectrical stimulation in the kilohertz-frequency range has been successfully used for treatment of various neurological disorders. Nevertheless, the mechanisms underlying this stimulation are poorly understood.

ObjectiveTo study the effect of kilohertz-frequency electric fields on neuronal membrane biophysics we developed a reliable experimental method to measure responses of single neurons to kilohertz field stimulation in brain slice preparations.

MethodsIn the submerged brain slice pyramidal neurons of the CA1 subfield were recorded in the whole-cell configuration before, during and after stimulation with an external electric field at 2kHz, 5kHz or 10 kHz.

ResultsReproducible excitatory changes in rheobase and spontaneous firing were elicited during kHz-field application at all stimulating frequencies. The rheobase only decreased and spontaneous firing either was initiated in silent neurons or became more intense in previously spontaneously active neurons. Response thresholds were higher at higher frequencies. Blockade of glutamatergic synaptic transmission did not alter the magnitude of responses. Inhibitory synaptic input was not changed by kilohertz field stimulation.

ConclusionkHz-frequency current applied in brain tissue has an excitatory effect on pyramidal neurons during stimulation. This effect is more prominent and occurs at a lower stimulus intensity at a frequency of 2kHz as compared to 5kHz and 10kHz.

category: neuroscience


title: Reduced infectivity but increased immune escape of the new SARS-CoV-2 variant of concern Omicron

abstract: A new detected SARS-CoV-2 variant Omicron (B.1.1.529) had reported from more than 80 countries. In the past few weeks, a new wave of infection driven by Omicron is in progress. Omicron Spike (S) protein pseudotyped virus was used to determine the effect of S mutations on its capacity of infectivity and immune evasion. Our results showed the lower entry efficiency and less cleavage ability of Omicron than D614G variant. Pseudotype-based neutralizing assay was performed to analyze neutralizing antibodies elicited by previously infection or the RBD-based protein subunit vaccine ZF2001 against the Omicron variant. Sera sampled at around one month after symptom onset from 12 convalescents who were previously infected by SARS-CoV-2 original strain shows a more than 20-fold decrease of neutralizing activity against Omicron variant, when compared to D614G variant. Among 12 individuals vaccinated by RBD subunit vaccine, 58.3% (7/12) sera sampled at 15-60 days after 3rd-dose vaccination did not neutralize Omicron. Geometric mean titers (GMTs, 50% inhibitory dose [ID50]) of these sera against Omicron were 9.4-fold lower than against D614G. These results suggested a higher risk of Omicron breakthrough infections and reduced efficiency of the protective immunity elicited by existing vaccines. There are important implications about the modification and optimization of the current epidemic prevention and control including vaccine strategies and therapeutic antibodies against Omicron variant.

category: microbiology


title: Perception and propagation of activity through the cortical hierarchy is determined by neural variability

abstract: The brains of higher organisms are composed of anatomically and functionally distinct regions performing specialised tasks; but regions do not operate in isolation. Orchestration of complex behaviours requires communication between brain regions, but how neural activity dynamics are organised to facilitate reliable transmission is not well understood. We studied this process directly by generating neural activity that propagates between brain regions and drives behaviour, allowing us to assess how populations of neurons in sensory cortex cooperate to transmit information. We achieved this by imaging two hierarchically organised and densely interconnected regions, the primary and secondary somatosensory cortex (S1 and S2) in mice while performing two-photon photostimulation of S1 neurons and assigning behavioural salience to the photostimulation. We found that the probability of perception is determined not only by the strength of the photostimulation signal, but also by the variability of S1 neural activity. Therefore, maximising the signal-to-noise ratio of the stimulus representation in cortex is critical to its continued propagation downstream. Further, we show that propagated, behaviourally salient activity elicits balanced, persistent, and generalised activation of the downstream region. Hence, our work adds to existing understanding of cortical function by identifying how population activity is formatted to ensure robust transmission of information, allowing specialised brain regions to communicate and coordinate behaviour.


title: SoyFGB v2.0: a unique access to variations of Chinese Soybean Gene Bank (CNSGB) germplasm

abstract:
In Chinese National Soybean GeneBank (CNSGB), we have collected more than 30,000 soybean accessions. However, data sharing for soybean remains an especially  sensitive question, and how to share the genome variations within rule frame has been bothering the soybean germplasm workers for a long time.

Here we release a big data source named Soybean Functional Genomics & Breeding database (SoyFGB v2.0) (https://sfgb.rmbreeding.cn/), which embed a core collection of 2,214 soybean resequencing genome (2K-SG) from the CNSGB germplasm. This source presents a unique example which may help elucidating the following three major functions for multiple genome data mining with general interests for plant researchers. 1) On-line analysis tools are provided by the  Analysis module for haplotype mining in high-throughput genotyped germplasms with different methods. 2) Variations for 2K-SG are provided in SoyFGB v2.0 by Browse module which contains two functions of  SNP and  InDel. Together with the  Gene (SNP & InDel) function embedded in Search module, the genotypic information of 2K-SG for targeting gene / region is accessible. 3) Scaled phenotype data of 42 traits, including 9 quality and 33 quantitative traits are provided by SoyFGB v2.0. With the scaled-phenotype data search and seed request tools under a control list, the germplasm information could be shared without direct downloading the unpublished phenotypic data or information of sensitive germplasms.

In a word, the mode of data mining and sharing underlies SoyFGB v2.0 may inspire more ideas for works on genome resources of not only soybean but also the other plants.

category: genomics


title: Differences in running performance of single- and group-housed mice

abstract:
Mice are one of several common animal models in neuroscience and mouse behavior is becoming increasingly relevant. Mice are housed either in groups or alone in standard cages during which they show a variety of different behaviors. Moreover, housing conditions might alter the behavior of the mice as do social interactions. When given access to running wheels as an environmental enrichment, mice tend to run long distances preferentially during their dark cycle. However, it is currently not well understood whether and how mice utilize running wheels when single-housed or group-housed. Here we developed a low-cost running wheel data acquisition system to study running in adult C57BL/6 mice at high temporal resolution under different social conditions. As expected, adult C57BL/6 mice prefer to run in stretches during the dark cycle and mostly rest during the light cycle. When single-housed, running bouts occur independent from each other as indicated by an exponential decaying autocorrelation. In contrast, mice run [~]50% less when housed in groups of n = 3 and their temporal pattern of running exhibits a power law decay in the autocorrelation indicative of potential social interactions. Our results demonstrate that running wheels are a limited resource for which mice compete for when they are group-housed, thereby reducing their overall running activity.

Significance StatementVoluntary cage wheel running is an important environmental enrichment for mice housed either alone or in groups. We show that this resource is considerably restricted for group-housed mice.

category: animal behavior and cognition


title: Effect of lipopolysaccharide (LPS) on HAEC cells. Does nicotinamide N-methyltranferase sensitize HAEC cells to LPS?

abstract: Treatment of endothelial cells with bacterial lipopolysaccharide (LPS) evokes a number of metabolic and functional consequences which built a multifaceted physiological response of endothelium to bacterial infection. Here effects of LPS on human aortic endothelial cells (HAEC) have been investigated. Among the spectrum of biochemical changes substantially elevated N-nicotinamide methyltransferase (NNMT) protein level was particularly intriguing. This important enzyme may potentially affect cellular metabolism by two means: direct regulation of methylnicotinamide level and availability of nicotinamide, that at least potentially may influence NAD+ synthesis, and regulation of S-adenosylmethionine concentration and therefore controlling methylation of many proteins including chromatin. This may have epigenetic consequences. This paper is focused on NNMT, despite the fact that in the presence of LPS additional effects of this compound mask pure (canonical) consequences of the elevated NNMT protein which are an increased MNA synthesis or reduced NAD+ level. On the other hand, however, it has been shown that silencing of the NNMT-encoding gene prevents several changes which are observed in control HAECs treated with LPS. They include significantly increased calcium response to thapsigargin (store-operated calcium entry), altered energy metabolism which is switched to anaerobic glycolysis and rearrangement of the mitochondrial network. However, a biochemical mechanism behind the protective consequences of the NNMT deficiency in cells treated with LPS remains unexplained.

category: biochemistry


title: A miRNAs Based Exploration of promising Biomarkers in Cervical Cancer using Bioinformatic Methods

abstract: Cervical Cancer (CC) is a gynecologic cancer. In this cancer early detection is incredibly tough because most of the patients are not have any specific symptoms that results in suspending the proper identification. In this work, we selected TCGA CESC datasets and miRNA Seq analysis was done. The expression profiles of miRNAs in cervical cancer datasets were investigated using bioinformatics tools. The expression profiles of miRNA in Normal tissue, primary tumor and metastatic samples were analyzed. Based on p-value, principal component analysis and comparative literature survey, we reported 6 over-expressed (5X) miRNA at metastatic stage namely, hsa-mir-363, hsa-mir-429, hsa-mir-141, hsa-mir-93, hsa-mir-203b and hsa-mir-18a. Expression profiles were compared in heatmap. The target genes for the selected miRNAs were investigated for interaction and pathway details. The identification of two hub proteins (PTEN and MYC) in Protein-Protein Interaction Network was followed by pathway analysis. Our results indicate that hsa-mir-363, hsa-mir-429, hsa-mir-141, hsa-mir-93, hsa-mir-203b and hsa-mir-18a could be a potential diagnostic biomarkers for early-stage CESC and serve as prognostic predictors for patients with CESC.

category: bioinformatics


title: An algorithm and application to efficiently analyze DNA fiber data

abstract: The duplication of genetic information (DNA replication) is central to life. Numerous control mechanisms ensure the exact course of the process during each cell division. Disturbances of DNA replication have severe consequences for the affected cell, and current models link them to cancer development. One of the most accurate methods for studying DNA replication is labeling newly synthesized DNA molecules with halogenated nucleotides, followed by immunofluorescence and microscopy detection, known as DNA fiber labeling. The method allows the registration of the activity of single replication complexes by measuring the length of the "trace" left by each of them. The major difficulty of the method is the labor-intensive analysis, which requires measuring the lengths of a large number of labeled fragments. Recently, the interest in this kind of image analysis has grown rapidly. In this manuscript, we provide a detailed description of an algorithm and a lightweight Java application to automatically analyze single DNA molecule images we call "DNA size finder". DNA size finder significantly simplified the analysis of the experimental data while increasing reliability by the standardized measurement of a greater number of DNA molecules. It is freely available and does not require any paid platforms or services to be used. We hope that the application will facilitate both the study of DNA replication control and the effects of various compounds used in human activity on the process of DNA replication.


title: Design of Epitope Based Peptide Vaccine against Brucella Abortus OmpW Family Protein using Immunoinformatics

abstract: Brucella abortus is a small aerobic, non-spore-forming, non-motile intracellular coccobacilli localized in the reproductive organs of host animals and causes acute or chronic disorders. It infects approximately 200 cases per 100,000 of the population and has become endemic in many countries. OmpW family protein is an outer membrane protein involved in the initial interaction between the pathogen and its host. This study predicts an effective epitope-based vaccine against OmpW family protein of Brucella abortus using immunoinformatics tools. Sequences were obtained from NCBI and prediction tests were accomplished to analyze possible epitopes for B and T cells. Seven B cell epitopes passed the antigenicity, accessibility and hydrophilicity tests. Forty-three MHC I epitopes were the most promising, while 438 from MHC II. For the population coverage, the epitopes covered 99.97% of the alleles worldwide excluding certain MHC II alleles. We recommend invivo and invitro studies to prove its effectiveness.


title: Narcolepsy and the Dissociation of REM Sleep and Cataplexy through Ambient Temperature Manipulation

abstract:
Narcolepsy is characterized by increased REM sleep propensity and cataplexy. Although narcolepsy is caused by the selective loss or dysfunction of hypocretin (Hcrt) neurons within the lateral hypothalamus (LH), mechanisms underlying REM sleep propensity and cataplexy remain to be elucidated. We have recently shown that wild type (WT) mice increase REM sleep expression when exposed to thermoneutral ambient temperature (Ta) warming during the light (inactive) phase. We hypothesized that the loss of Hcrt may lead to exaggerated responses with respect to increased REM sleep and cataplexy during Ta warming. To test this hypothesis, Hcrt-KO mice were implanted for chronic sleep recordings and housed in a temperature-controlled cabinet. Sleep-wake expression and both spontaneous cataplexy and food-elicited cataplexy were evaluated at constant Ta and during a Ta manipulation protocol. Here we show several unexpected findings. First, Hcrt-KO mice show opposite circadian patterns with respect to REM sleep responsiveness to thermoneutral Ta warming compared to WT mice. As previous demonstrated, WT mice increased REM sleep when Ta warming is presented during the inactive (light) phase, whereas Hcrt-KO showed a significant decrease in REM sleep expression. In contrast, Hcrt-KO mice increased REM sleep expression upon exposure to Ta warming when presented during the active (dark) phase, a circadian time when WT mice showed no significant changes in REM sleep as a function of Ta. Second, we found that REM sleep and cataplexy can be dissociated through Ta manipulation. Specifically, although Ta warming significantly increased REM sleep expression in Hcrt-KO mice during the active phase, cataplexy bout number and total cataplexy duration significantly decreased. In contrast, cataplexy expression was favoured during Ta cooling when REM sleep expression significantly decreased. Finally, video actigraphy and sleep-wake recordings in Hcrt-KO mice demonstrated that Ta manipulation did not significantly alter waking motor activity patterns or waking or NREM sleep durations. These data suggest that neural circuits gating REM sleep and cataplexy expression can be dissociated with Ta manipulation.

Statement of SignificanceCataplexy and the loss of muscle tone has historically been viewed as a component of REM sleep that inappropriately intrudes into wakefulness. In addition to fragmented sleep, the intrusion of REM-like events into wakefulness has led to the hypothesis that narcolepsy represents a dysregulation of boundary state control. We show that REM sleep and cataplexy can be dissociated during the dark (active) phase through Ta manipulation. Such dissociation may provide clues regarding the cause of boundary state instability in Hcrt-KO mice, as well as provide a new method to interrogate mechanisms of REM sleep and cataplexy.


title: A meta-analysis of task-based differences in bilingual L1 and L2 language networks

abstract: The functional organization of first (L1) and second (L2) language processing in bilinguals remains a topic of great interest to the neurolinguistics community. Functional magnetic resonance imaging (fMRI) studies report meaningful differences in the location and extent of hemodynamic changes between tasks performed in the L1 and L2, yet there is no consensus on whether these networks can be considered truly distinct. In part, this may be due to the multiplicity of task designs implemented in such studies, which complicates the interpretation of their findings. This paper compares the results of previous bilingual meta-analyses to a new ALE meta-analysis that categorizes neuroimaging studies by task design. Factors such as the age of L2 acquisition (AoA) and the L2 language proficiency level of participants are also considered. The findings support previous accounts of the effect of participant characteristics on linguistic processing, while at the same time revealing dissociable differences in fMRI activation for L1 and L2 networks within and across tasks that appear independent of these external factors.


title: Natural history of nonhuman primates after conjunctival exposure to Ebola virus

abstract: Transmission of Ebola virus (EBOV) primarily occurs via contact exposure of mucosal surfaces with infected body fluids. Historically, nonhuman primate (NHP) challenge studies have employed intramuscular or small particle aerosol exposure, which are uniformly lethal routes of infection, but mimic worst-case scenarios such as a needlestick. When exposed by more likely routes of natural infection, limited NHP studies have shown delayed onset of disease and reduced mortality. Here we performed a series of systematic natural history studies in cynomolgus macaques with a range of conjunctival exposure doses. Challenge with 10,000 plaque forming units (PFU) of EBOV was uniformly lethal, whereas 5/6 subjects survived low and moderate dose challenges (100 or 500 PFU). Conjunctival challenge resulted in a protracted time-to death. Asymptomatic disease was observed in survivors with limited detection of EBOV replication. Inconsistent seropositivity in survivors may suggest physical or natural immunological barriers are sufficient to prevent widespread viral dissemination.


title: Thermal sensitivity and seasonal change in the gut microbiome of a desert ant, Cephalotes rohweri

abstract: Microorganisms within ectotherms must withstand the variable body temperatures of their hosts. Shifts in host body temperature resulting from climate change have the potential to shape ectotherm microbiome composition. Microbiome compositional changes occurring in response to temperature in nature have not been frequently examined, restricting our ability to predict microbe-mediated ectotherm responses to climate change. In a set of field-based observations, we characterized gut bacterial communities and thermal exposure across a population of desert arboreal ants (Cephalotes rohweri). In a paired growth chamber experiment, we exposed ant colonies to variable temperature regimes differing by 5 {degrees}C for three months. We found that the abundance and composition of ant-associated bacteria were sensitive to elevated temperatures in both field and laboratory experiments. We observed a subset of taxa that responded similarly to temperature in the experimental and observational study, suggesting a role of seasonal temperature and local temperature differences amongst nests in shaping microbiomes within the ant population. Bacterial mutualists in the genus Cephalotococcus (Opitutales: Opitutaceae) were especially sensitive to change in temperature--decreasing in abundance in naturally warm summer nests and warm growth chambers. We also report the discovery of a member of the Candidate Phlya Radiation (Phylum: Gracilibacteria), a suspected epibiont, found in low abundance within the guts of this ant species.


title: Microtubule plus-end regulation by centriolar cap proteins

abstract: Centrioles are microtubule-based organelles required for the formation of centrosomes and cilia. Centriolar microtubules, unlike their cytosolic counterparts, grow very slowly and are very stable. The complex of centriolar proteins CP110 and CEP97 forms a cap that stabilizes the distal centriole end and prevents its over-elongation. Here, we used in vitro reconstitution assays to show that whereas CEP97 does not interact with microtubules directly, CP110 specifically binds microtubule plus ends, potently blocks their growth and induces microtubule pausing. Cryo-electron tomography indicated that CP110 binds to the luminal side of microtubule plus ends and reduces protofilament peeling. Furthermore, CP110 directly interacts with another centriole biogenesis factor, CPAP/SAS- 4, which tracks growing microtubule plus ends, slows down their growth and prevents catastrophes. CP110 and CPAP synergize in inhibiting plus-end growth, and this synergy depends on their direct binding. Together, our data reveal a molecular mechanism controlling centriolar microtubule plus- end dynamics and centriole biogenesis.

category: cell biology


title: Identification of ER/SR resident proteins as biomarkers for ER/SR calcium depletion in skeletal muscle cells

abstract: Aberrations to endoplasmic/sarcoplasmic reticulum (ER/SR) calcium concentration can result in the departure of endogenous proteins in a phenomenon termed exodosis. Redistribution of the ER/SR proteome can have deleterious effects to cell function and cell viability, often contributing to disease pathogenesis. Many proteins prone to exodosis reside in the ER/SR via an ER retention/retrieval sequence (ERS) and are involved in protein folding, protein modification, and protein trafficking. While the consequences of their extracellular presence have yet to be fully delineated, the proteins that have undergone exodosis may be useful for biomarker development. Skeletal muscle cells rely upon tightly coordinated ER/SR calcium release for muscle contractions, and perturbations to calcium homeostasis can result in myopathies. Ryanodine receptor type-1 (RYR1) is a calcium release channel located in the SR. Mutations to the RYR1 gene can compromise calcium homeostasis leading to a vast range of clinical phenotypes that include hypotonia, myalgia, respiratory insufficiency, ophthalmoplegia, fatigue and malignant hyperthermia (MH). There are currently no FDA approved treatments for RYR1-related myopathies (RYR1-RM). Here we examine the exodosis profile of skeletal muscle cells following ER/SR calcium depletion. Proteomic analysis identified 4,465 extracellular proteins following ER/SR calcium depletion with 1,280 proteins significantly different than vehicle. A total of 54 ERS proteins were identified and 33 ERS proteins significantly increased following ER/SR calcium depletion. Specifically, ERS protein, mesencephalic astrocyte-derived neurotrophic factor (MANF), was elevated following calcium depletion, making it a potential biomarker candidate for human samples. Despite no significant elevation of MANF in plasma levels among healthy volunteers and RYR1-RM individuals, MANF plasma levels positively correlated with age in RYR1-RM individuals, presenting a potential biomarker of disease progression. Selenoprotein N (SEPN1) was also detected only in extracellular samples following ER/SR calcium depletion. This protein is integral to calcium handling and SEPN1 variants have a causal role in SEPN1-related myopathies (SEPN1-RM). Extracellular presence of ER/SR membrane proteins may provide new insight into proteomic alterations extending beyond ERS proteins. Pre-treatment of skeletal muscle cells with bromocriptine, an FDA approved drug recently found to have anti-exodosis effects, curbed exodosis of ER/SR resident proteins. Changes to the extracellular content caused by intracellular calcium dysregulation presents an opportunity for biomarker development and drug discovery.


title: Testing for genetic assimilation with phylogenetic comparative analysis: Conceptual, methodological, and statistical considerations

abstract: Genetic assimilation is a process that leads to reduced phenotypic plasticity during adaptation to novel conditions, a potentially important phenomenon under global environmental change. Null expectations when testing for genetic assimilation, however, are not always clear. For instance, the statistical artifact of regression to the mean could bias us towards detecting genetic assimilation when it has not occurred. Likewise, the specific mechanism underlying plasticity expression may affect null expectations under neutral evolution. We used macroevolutionary numerical simulations to examine both of these important issues and their interaction, varying whether or not plasticity evolves, the evolutionary mechanism, trait measurement error, and experimental design. We also modified an existing reaction norm correction method to account for phylogenetic non-independence. We found: 1) regression to the mean is pervasive and can generate spurious support for genetic assimilation; 2) experimental design and post-hoc correction can minimize this spurious effect; and 3) neutral evolution can produce patterns consistent with genetic assimilation without constraint or selection, depending on the mechanism of plasticity expression. Additionally, we re-analyzed published macroevolutionary data supporting genetic assimilation, and found that support was lost after proper correction. Considerable caution is thus required whenever investigating genetic assimilation and reaction norm evolution at macroevolutionary scales.

category: evolutionary biology


title: Evidence of learning walks related to scorpion home burrow navigation

abstract: The Navigation by Chemotextural Familiarity Hypothesis (NCFH) suggests that scorpions use their midventral pectines to gather chemical and textural information near their burrows and use this information as they subsequently return home. For NCFH to be viable, animals must somehow acquire home-directed "tastes" of the substrate, such as through path integration (PI) and/or learning walks. We conducted laboratory behavioral trials using desert grassland scorpions (Paruroctonus utahensis). Animals reliably formed burrows in small mounds of sand we provided in the middle of circular, sandlined behavioral arenas. We processed overnight infrared video recordings with a MATLAB script that tracked animal movements at 1-2 s intervals. In all, we analyzed the movements of 23 animals, representing nearly 1500 hours of video recording. We found that once animals established their home burrows, they immediately made one to several short, looping excursions away from and back to their burrows before walking greater distances. We also observed similar excursions when animals made burrows in level sand in the middle of the arena (i.e., no mound provided). These putative learning walks, together with recently reported PI in scorpions, may provide the crucial home-directed information requisite for NCFH.


title: Identification of critical genes and biological signaling for metformin treated liver cancer

abstract: Liver cancer is a leading source of cancer-related mortality in the world. A number of studies have shown the correlation of metformin treatment with a decrease in cancer risk. However, the relevant molecules and mechanisms are not clear during the treatment. In this study, our aim is to identify the significant molecules and signaling pathways in the treatment of metformin in liver cancer cells by analyzing the RNA sequence. The GSE190076 dataset was created by performing the Illumina NovaSeq 6000 (Homo sapiens). The KEGG and GO analyses indicated that DNA synthesis and cell cycle are the main processes during the treatment of metformin. Moreover, we determined numerous genes including RRM2, CDC6, CDC45, UHRF1, ASF1B, ZWINT, PCNA, ASPM, MYC, and TK1 by using the PPI network. Therefore, our study may guide the clinical work on the treatment of liver cancer by using metformin.


title: Structural and functional characterizations of altered infectivity and immune evasion of SARS-CoV-2 Omicron variant

abstract: The SARS-CoV-2 Omicron with increased fitness is spreading rapidly worldwide. Analysis of cryo-EM structures of the Spike (S) from Omicron reveals amino acid substitutions forging new interactions that stably maintain an "active" conformation for receptor recognition. The relatively more compact domain organization confers improved stability and enhances attachment but compromises the efficiency of viral fusion step. Alterations in local conformation, charge and hydrophobic microenvironments underpin the modulation of the epitopes such that they are not recognized by most NTD- and RBD-antibodies, facilitating viral immune escape. Apart from already existing mutations, we have identified three new immune escape sites: 1) Q493R, 2) G446S and 3) S371L/S373P/S375F that confers greater resistance to five of the six classes of RBD-antibodies. Structure of the Omicron S bound with human ACE2, together with analysis of sequence conservation in ACE2 binding region of 25 sarbecovirus members as well as heatmaps of the immunogenic sites and their corresponding mutational frequencies sheds light on conserved and structurally restrained regions that can be used for the development of broad-spectrum vaccines and therapeutics.

category: biophysics


title: Network architecture determines vein fate during spontaneous reorganization, with a time delay

abstract: Vascular networks continuously reorganize their morphology by growing new or shrinking existing veins to optimize function. Flow shear stress on vein walls has been set forth as the local driver for this continuous adaptation. Yet, shear feedback alone cannot account for the observed diversity of network dynamics - a puzzle made harder by scarce spatio-temporal data. Here, we resolve network-wide vein dynamics and shear during spontaneous reorganization in the prototypical vascular networks of Physarum polycephalum. Our experiments reveal a plethora of vein dynamics (stable, growing, shrinking) that are not directly proportional to local shear. We observe (a) that shear rate sensing on vein walls occurs with a time delay of 1 min to 3 min and (b) that network architecture dependent parameters - such as relative pressure or relative vein resistance - are key to determine vein fate. We derive a model for vascular adaptation, based on force balance at the vein walls. Together with the time delay, our model reproduces the diversity of experimentally observed vein dynamics, and confirms the role of network architecture. Finally, we observe avalanches of network reorganization events which cause entire clusters of veins to vanish. Such avalanches are consistent with architectural feedback as the vein connections perpetually change with reorganization. As these network architecture dependent parameters are intrinsically connected with the laminar fluid flow in the veins, we expect our findings to play a role across flow-based vascular networks.


title: Multi-Wavelength Analytical Ultracentrifugation of Biopolymer Mixtures and Interactions

abstract: Multi-wavelength analytical ultracentrifugation (MW-AUC) is a recent development made possible by new analytical ultracentrifuge optical systems. MW-AUC is suitable for a wide range of applications and biopolymer systems and is poised to become an essential tool to characterize macromolecular interactions. It adds an orthogonal spectral dimension to the traditional hydrodynamic characterization by exploiting unique chromophores in analyte mixtures that may or may not interact. Here we illustrate the utility of MW-AUC for representative classes of challenging biopolymer systems, including interactions between mixtures of different sized proteins with small molecules, mixtures of loaded and empty viral AAV capsids contaminated with free DNA, and mixtures of different proteins, where some have identical hydrodynamic properties, all of which are difficult to resolve with traditional AUC methods. We explain the improvement in resolution and information content obtained by this technique compared to traditional single- or dual-wavelength approaches. We discuss experimental design considerations and limitations of the method, and address the advantages and disadvantages of the two MW optical systems available today, and the differences in data analysis strategies between the two systems.


title: VRK1 is required in VRK2-methylated cancers of the nervous system

abstract:
Collateral lethality occurs when loss of one paralog renders cancer cells dependent on the remaining paralog. Combining genome scale CRISPR/Cas9 screens coupled with RNA-sequencing in over 900 cancer cell lines, we found that cancers of nervous system lineage, including adult and pediatric gliomas and neuroblastomas, required the nuclear kinase Vaccinia-Related Kinase 1 (VRK1) for their survival. VRK1 dependency was inversely correlated with expression of its paralog VRK2. VRK2 knockout (KO) sensitized cells to VRK1 suppression, and conversely, VRK2 overexpression increased cell fitness in the setting of VRK1 suppression. DNA methylation of the VRK2 promoter was associated with low VRK2 expression in human neuroblastomas, and adult and pediatric gliomas. Mechanistically, depletion of VRK1 reduced Barrier-to-Autointegration Factor (BAF) phosphorylation during mitosis, resulting in DNA damage and apoptosis. Together, these studies identify VRK1 as a synthetic lethal target in VRK2 promoter-methylated adult and pediatric gliomas and neuroblastomas.

Statement of SignificanceWe credential VRK1 as a target in adult and pediatric gliomas, and neuroblastomas with VRK2 promoter methylation. This demonstrates the utility of paralog-driven synthetic lethal interactions for biomarker-linked, targeted therapeutics.


title: Broad neutralization of SARS-CoV-2 variants by an inhalable bispecific single-domain antibody

abstract: The effectiveness of SARS-CoV-2 vaccines and therapeutic antibodies has been limited by the continuous emergence of viral variants, and by the restricted diffusion of antibodies from circulation into the sites of respiratory virus infection. Here, we report the identification of two highly conserved regions on Omicron variant RBD recognized by broadly neutralizing antibodies. Based on this finding, we generated a bispecific single-domain antibody that was able to simultaneously and synergistically bind these two regions on a single Omicron variant RBD as revealed by Cryo-EM structures. This inhalable antibody exhibited exquisite neutralization breadth and therapeutic efficacy in mouse models of SARS-CoV-2 infections. The structures also deciphered an uncommon cryptic epitope within the spike trimeric interface that may have implications for the design of broadly protective SARS-CoV-2 vaccines and therapeutics.

category: immunology


title: Pipeline for retrieval of COVID-19 immune signatures

abstract:
ObjectiveThe accelerating pace of biomedical publication has made retrieving papers and extracting specific comprehensive scientific information a key challenge. A timely example of such a challenge is to retrieve the subset of papers that report on immune signatures (coherent sets of biomarkers) to understand the immune response mechanisms which drive differential SARS-CoV-2 infection outcomes. A systematic and scalable approach is needed to identify and extract COVID-19 immune signatures in a structured and machine-readable format.

Materials and MethodsWe used SPECTER embeddings with SVM classifiers to automatically identify papers containing immune signatures. A generic web platform was used to manually screen papers and allow anonymous submission.

ResultsWe demonstrate a classifier that retrieves papers with human COVID-19 immune signatures with a positive predictive value of 86%. Semi-automated queries to the corresponding authors of these publications requesting signature information achieved a 31% response rate. This demonstrates the efficacy of using a SVM classifier with document embeddings of the abstract and title, to retrieve papers with scientifically salient information, even when that information is rarely present in the abstract. Additionally, classification based on the embeddings identified the type of immune signature (e.g., gene expression vs. other types of profiling) with a positive predictive value of 74%.

ConclusionCoupling a classifier based on document embeddings with direct author engagement offers a promising pathway to build a semistructured representation of scientifically relevant information. Through this approach, partially automated literature mining can help rapidly create semistructured knowledge repositories for automatic analysis of emerging health threats.


title: Increased resistance of SARS-CoV-2 Omicron Variant to Neutralization by Vaccine-Elicited and Therapeutic Antibodies

abstract: Currently authorized vaccines for SARS-CoV-2 have been highly successful in preventing infection and lessening disease severity. The vaccines maintain effectiveness against SARS-CoV-2 Variants of Concern but the heavily mutated, highly transmissible Omicron variant poses an obstacle both to vaccine protection and monoclonal antibody therapies. Analysis of the neutralization of Omicron spike protein-pseudotyped lentiviruses showed a 26-fold relative resistance (compared to D614G) to neutralization by convalescent sera and 26-34-fold resistance to Pfizer BNT162b2 and Moderna vaccine-elicited antibodies following two immunizations. A booster immunization increased neutralizing titers against Omicron by 6-8-fold. Previous SARS-CoV-2 infection followed by vaccination resulted in the highest neutralizing titers against Omicron. Regeneron REGN10933 and REGN10987, and Lilly LY-CoV555 and LY-CoV016 monoclonal antibodies were ineffective against Omicron, while Sotrovimab was partially effective. The results highlight the benefit of a booster immunization in providing protection against Omicron but demonstrate the challenge to monoclonal antibody therapies.


title: Viral population genomics reveals host and infectivity impact on SARS-CoV-2 adaptive landscape

abstract: Public health surveillance, drug treatment development, and optimization of immunological interventions all depend on understanding pathogen adaptation, which differ for specific pathogens. SARS-CoV-2 is an exceptionally successful human pathogen, yet complete understanding of the forces driving its evolution is lacking. Here, we leveraged almost four million SARS-CoV-2 sequences originating mostly from non-vaccinated naive patients to investigate the impact of functional constraints and natural immune pressures on the sequence diversity of the SARS-CoV-2 genome. Overall, we showed that the SARS-CoV-2 genome is under strong and intensifying levels of purifying selection with a minority of sites under diversifying pressure. With a particular focus on the spike protein, we showed that sites under selection were critical for protein stability and virus fitness related to increased infectivity and/or reduced neutralization by convalescent sera. We investigated the genetic diversity of SARS-CoV-2 B and T cell epitopes and determined that the currently known T cell epitope sequences were highly conserved. Outside of the spike protein, we observed that mutations under selection in variants of concern can be associated to beneficial outcomes for the virus. Altogether, the results yielded a comprehensive map of all sites under selection across the entirety of SARS-CoV-2 genome, highlighting targets for future studies to better understand the virus spread, evolution and success.


title: Unsupervised genome-wide cluster analysis: nucleotide sequences of the omicron variant of SARS-CoV-2 are similar to sequences from early 2020

abstract: The GISAID database contains more than 100,000 SARS-CoV-2 genomes, including sequences of the recently discovered SARS-CoV-2 omicron variant and of prior SARS-CoV-2 strains that have been collected from patients around the world since the beginning of the pandemic. We applied unsupervised cluster analysis to the SARS-CoV-2 genomes, assessing their similarity at a genome-wide level based on the Jaccard index and principal component analysis. Our analysis results show that the omicron variant sequences are most similar to sequences that have been submitted early in the pandemic around January 2020. Furthermore, the omicron variants in GISAID are spread across the entire range of the first principal component, suggesting that the strain has been in circulation for some time. This observation supports a long-term infection hypothesis as the omicron strain origin.


title: Thresholding Approach for Low-Rank Correlation Matrix based on MM algorithm

abstract:
BackgroundLow-rank approximation is used for interpreting the features of a correlation matrix using visualization tools; however, a low-rank approximation may result in estimation that is far from zero even if the corresponding original value is zero. In such a case, the results lead to a misinterpretation.

MethodsTo overcome this, we propose a novel approach to estimate a sparse low-rank correlation matrix based on threshold values. We introduce a new cross-validation function to tune the corresponding threshold values. To calculate the value of a function, the MM algorithm is used to estimate the sparse low-rank correlation matrix, and a grid search was performed to select the threshold values.

ResultsThrough numerical simulation, we found that the false positive rate (FPR) and average relative error of the proposed method were superior to those of the tandem approach. For the application of microarray gene expression, the FPRs of the proposed approach with d = 2, 3, and 5 were 0.128, 0.139, and 0.197, respectively, whereas the FPR of the tandem approach was 0.285.

ConclusionsWe propose a novel approach to estimate sparse low-rank correlation matrices. The advantage of the proposed method is that it provides results that are interpretable through the use of a heatmap, thereby avoiding result misinterpretations. We demonstrated the superiority of the proposed method through both numerical simulations and real examples.


title: Computational analysis of transition temperatures (Tts) of proteins fused to elastin-like polypeptide (ELP): deep fake evaluation of proteins, linkers, and trailers features

abstract:
The phase transition temperature (Tt) prediction of the Elastin-like polypeptides (ELPs) is not trivial because it is related to complex sets of variables such as composition, sequence length, hydrophobic characterization, hydrophilic characterization, the sequence order in the fused proteins, linkers and trailer constructs. In this paper, two unique quantitative models are presented for the prediction of the Tt of a family of ELPs that could be fused to different proteins, linkers, and trailers. The lack of need to use multiple software, peptide information, such as PDB file, as well as knowing the second and third structures of proteins are the advantages of this model besides its high accuracy and speed. One of our models could predict the Tt values of the fused ELPs by entering the protein, linker, and trailer features with R2=99%. Also, another model is able to predict the Tt value by entering the fused protein feature with R2=96%. For more reliability, our method is enriched by Artificial Intelligence (AI) to generate similar proteins. In this regard, Generative Adversarial Network (GAN) is our AI method to create fake proteins and similar values. The experimental results show that our strategy for prediction of Tt is reliable in large data.

Author SummaryThe application of Elastin-like polypeptides (ELPs) as a protein tag is now developed in a variety of biotechnology aspects especially in proteins purification and drug delivery. ELPs application as a protein tag is owed to retain the phase transition behavior when ELPs fused to other proteins, linkers and tags. ELPs undergo the phase transition behavior by changing from soluble phase to insoluble phase above its inverse transition temperature (Tt) within a short time span. This biophysical behavior is usually reversible at the temperature below the Tt. There are few reports for evaluation of the Tt of ELPs types by using the dissimilar equations and algorithms. Our current predictions are the most accurate calculations presented so far by using the protein, linker and trailer effects and the results were evaluated in accordance with the available experimental data. Furthermore, our results also show that our strategy for prediction of Tt is reliable in large data.


title: Symphonizing pileup and full-alignment for deep learning-based long-read variant calling

abstract: Deep learning-based variant callers are becoming the standard and have achieved superior SNP calling performance using long reads. In this paper, we present Clair3, which makes the best of two major method categories: pile-up calling handles most variant candidates with speed, and full-alignment tackles complicated candidates to maximize precision and recall. Clair3 ran faster than any of the other state-of-the-art variant callers and performed the best, especially at lower coverage.


title: Cortical glutamatergic projection neuron types contribute to distinct functional subnetworks

abstract: The cellular basis of cerebral cortex functional architecture remains not well understood. A major challenge is to monitor and decipher neural network dynamics across broad cortical areas yet with projection neuron (PN)-type resolution in real time during behavior. Combining genetic targeting and wide-field imaging, we monitored activity dynamics of subcortical-projecting (PTFezf2) and intratelencephalic-projecting (ITPlxnD1) types across dorsal cortex of mice during different brain states and behaviors. ITPlxnD1 and PTFezf2 neurons showed distinct activation patterns during wakeful resting, spontaneous movements, and upon sensory stimulation. Distinct ITPlxnD1 and PTFezf2 subnetworks were dynamically tuned to different sensorimotor components of a naturalistic feeding behavior, and optogenetic inhibition of subnetwork nodes disrupted specific components of this behavior. Lastly, ITPlxnD1 and PTFezf2 projection patterns are consistent with their subnetwork activation patterns. Our results show that, in addition to the concept of columnar organization, dynamic areal and PN type-specific subnetworks are a key feature of cortical functional architecture linking microcircuit components with global brain networks.


title: Prolactin action is necessary for parental behavior in male mice

abstract: Parental care is critical for successful reproduction in mammals. In comparison to maternal care, the neuroendocrine mechanisms supporting paternal care are less well-studied. Laboratory mice show a mating-induced suppression of infanticide (normally observed in virgins) and onset of paternal behavior. Using this model, we sought to investigate whether the hormone prolactin plays a role in paternal behavior, as it does for maternal behavior. First, using c-fos immunoreactivity in Prlr-IRES-Cre-tdtomato reporter mouse sires, we show that the circuitry activated during paternal interactions contains prolactin-responsive neurons, including the medial preoptic area, bed nucleus of the stria terminalis, and medial amygdala. To evaluate whether prolactin action is required for the establishment and display of paternal behavior, we conditionally deleted the prolactin receptor (Prlr) from 3 distinct cell types: glutamatergic, GABAergic, and CaMKII-expressing forebrain neurons. Prlr-deletion from CaMKII-expressing forebrain neurons, but not from glutamatergic or GABAergic cells, resulted in a profound effect on paternal behavior, as none of these males completed the pup retrieval task. Finally, although sires do not show an acute increase in circulating prolactin levels in response to pups, pharmacological blockade of prolactin-release at the time of pup exposure resulted in failure to retrieve pups, similar to when the Prlr was deleted from CaMKII neurons, with prolactin administration rescuing this behavior. Taken together, our data show that paternal behavior in sires is dependent on basal levels of circulating prolactin acting at the Prlr on CaMKII-expressing neurons. These new data in male mice demonstrate that prolactin has a similar action in both sexes to promote parental care.


title: Preserved T cell reactivity to the SARS-CoV-2 Omicron variant indicates continued protection in vaccinated individuals.

abstract:
ImportanceThe emergence of the highly contagious Omicron variant of SARS-CoV-2 and the findings of a significantly reduced neutralizing potency of sera from convalescent or vaccinated individuals imposes the study of cellular immunity to predict the degree of immune protection to the yet again new coronavirus.

DesignProspective monocentric observational study.

SettingConducted between December 20-21 at the Santa Lucia Foundation IRCCS.

Participants61 volunteers (Mean age 41.62, range 21-62; 38F/23M) with different vaccination and SARS-CoV-2 infection backgrounds donated 15 ml of blood. Of these donors, one had recently completed chemotherapy, and one was undergoing treatment with monoclonal antibodies; the others reported no known health issue.

Main Outcome(s) and Measure(s)The outcomes were the measurement of T cell reactivity to the mutated regions of the Spike protein of the Omicron SARS-CoV-2 variant and the assessment of remaining T cell immunity to the spike protein by stimulation with peptide libraries.

ResultsLymphocytes from freshly drawn blood samples were isolated and immediately tested for reactivity to the Spike protein of SARS-CoV-2. T cell responses to peptides covering the mutated regions in the Omicron variant were decreased by over 47% compared to the same regions of the ancestral vaccine strain. However, overall reactivity to the peptide library of the full-length protein was largely maintained (estimated 83%). No significant differences in loss of immune recognition were identified between groups of donors with different vaccination and/or infection histories.

Conclusions and RelevanceWe conclude that despite the mutations in the Spike protein, the SARS-CoV-2 Omicron variant is nonetheless recognized by the cellular component of the immune system. It is reasonable to assume that protection from hospitalization and severe disease is maintained.

Key PointsO_ST_ABSQuestionC_ST_ABSDoes the Omicron variant of SARS-CoV-2 escape cellular immunity?

FindingsThis observational study was performed on 61 vaccinated donors with established immunity to SARS-CoV-2. Cellular responses to the mutated regions of the Omicron Spike protein were detected in 80% of donors. The mutations reduced T cell recognition by 47% compared to the vaccine strain. Reactivity to the whole Spike protein, however, was present in 100% of donors, and the fraction of remaining immunity to SARS-CoV-2 was estimated to be 83%.

MeaningCellular immunity to the Omicron variant is maintained despite the mutations in its Spike protein, and may thus confer protection from severe COVID-19 in vaccinated individuals.


title: Molecular probes of spike ectodomain and its subdomains for SARS-CoV-2 variants, Alpha through Omicron

abstract: Since the outbreak of the COVID-19 pandemic, widespread infections have allowed SARS-CoV-2 to evolve in human, leading to the emergence of multiple circulating variants. Some of these variants show increased resistance to vaccines, convalescent plasma, or monoclonal antibodies. In particular, mutations in the SARS-CoV-2 spike have drawn attention. To facilitate the isolation of neutralizing antibodies and the monitoring the vaccine effectiveness against these variants, we designed and produced biotin-labeled molecular probes of variant SARS-CoV-2 spikes and their subdomains, using a structure-based construct design that incorporated an N-terminal purification tag, a specific amino acid sequence for protease cleavage, the variant spike-based region of interest, and a C-terminal sequence targeted by biotin ligase. These probes could be produced by a single step using in-process biotinylation and purification. We characterized the physical properties and antigenicity of these probes, comprising the N-terminal domain (NTD), the receptor-binding domain (RBD), the RBD and subdomain 1 (RBD-SD1), and the prefusion-stabilized spike ectodomain (S2P) with sequences from SARS-CoV-2 variants of concern or of interest, including variants Alpha, Beta, Gamma, Epsilon, Iota, Kappa, Delta, Lambda, Mu, and Omicron. We functionally validated probes by using yeast expressing a panel of nine SARS-CoV-2 spike-binding antibodies and confirmed sorting capabilities of variant probes using yeast displaying libraries of plasma antibodies from COVID-19 convalescent donors. We deposited these constructs to Addgene to enable their dissemination. Overall, this study describes a matrix of SARS-CoV-2 variant molecular probes that allow for assessment of immune responses, identification of serum antibody specificity, and isolation and characterization of neutralizing antibodies.


title: Subgenomic SARS-CoV-2 replicon and reporter replicon cell lines enable ultrahigh throughput antiviral screening and mechanistic studies with antivirals, viral mutations or host factors that affect COVID-19 replication

abstract: Replicon-based technologies were used to develop reagents and assays for advanced drug discovery efforts against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and for examining all facets of the SARS-CoV-2 replication cycle at reduced biocontainment level. Specifically: a) 21 replicons were cloned in bacterial artificial chromosomes (BACs) and delivered as transfectable plasmid DNA or transcribed RNA in various cell types. Replicons carrying mutations that affect the activity or antiviral susceptibility of SARS-CoV-2 enzymes were used to establish utility for mechanistic studies while reducing the community risks associated with gain-of-function studies in fully infectious virus. b) A BHK-21 stable cell line harboring SARS-CoV-2 replicon was generated and characterized in robust high/ultra-high throughput assays of antiviral efficacy with orthogonal SARS-CoV-2 replication reporter genes (Nano luciferase and enhanced green fluorescent protein-eGFP); the estimated antiviral potencies in the fully infectious SARS-CoV-2 system and in the transient or stable replicon systems were similar. HEK293 and Calu1 stable cell lines expressing SARS-CoV-2 replicon have also been prepared. Finally, c) we generated trans-encapsidated replicons by co-expression with SARS-CoV-2 structural proteins, thus producing single-round infectious SARS-CoV-2 virus-like particles that are able to transduce susceptible cell types and have expanded utility to enable study of virion assembly and entry into target cells. Hence, these SARS-CoV-2 replicon-based reagents include a novel approach to replicon-harboring cell line generation and are valuable tools that can be used at lower biosafety level (BSL2) for drug discovery efforts, characterization of SARS-CoV-2 and variant evolution in the COVID-19 pandemic, mechanisms of inhibition and resistance, and studies on the role of SARS-CoV-2 genes and host dependency factors.


title: SARS-CoV-2 diverges from other betacoronaviruses in only partially activating the IRE1α/XBP1 ER stress pathway in human lung-derived cells

abstract:
Despite the efficacy of vaccines, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has killed over 5 million individuals worldwide and continues to spread in countries where the vaccines are not yet widely available or its citizens are hesitant to become vaccinated. Therefore, it is critical to unravel the molecular mechanisms that allow SARS-CoV-2 and other coronaviruses to infect and overtake the host machinery of human cells. Coronavirus replication triggers endoplasmic reticulum (ER) stress and activation of the unfolded protein response (UPR), a key host cell pathway widely believed essential for viral replication. We examined the activation status and requirement of the master UPR sensor IRE1 kinase/RNase and its downstream transcription factor effector XBP1s, which is processed through an IRE1-mediated mRNA splicing event, in human lung-derived cells infected with betacoronaviruses. We found human respiratory coronavirus OC43 (HCoV-OC43), Middle East respiratory syndrome coronavirus (MERS-CoV), and the murine coronavirus (MHV) all induce ER stress and strongly trigger the kinase and RNase activities of IRE1 as well as XBP1 splicing. In contrast, SARS-CoV-2 only partially activates IRE1 whereby it autophosphorylates, but its RNase fails to splice XBP1. Moreover, IRE1 was dispensable for optimal replication in human cells for all coronaviruses tested. Our findings demonstrate that IRE1 activation status differs upon infection with distinct betacoronaviruses and is not essential for efficient replication of any of them. Our data suggest that SARS-CoV-2 actively inhibits the RNase of autophosphorylated IRE1 through an unknown mechanism, perhaps as a strategy to eliminate detection by the host immune system.

ImportanceSARS-CoV-2 is the third lethal respiratory coronaviruses after MERS-CoV and SARS-CoV to emerge this century, causing millions of deaths world-wide. Other common coronaviruses such as OC43 cause less severe respiratory disease. Thus, it is imperative to understand the similarities and differences among these viruses in how each interacts with host cells. We focused here on the inositol-requiring enzyme 1 (IRE1) pathway, part of the host unfolded protein response to virus-induced stress. We found that while MERS-CoV and OC43 fully activate the IRE1 kinase and RNase activites, SARS-CoV-2 only partially activates IRE1, promoting its kinase activity but not RNase activity. We propose that SARS-CoV-2 prevents IRE1 RNase activation as a strategy to eliminate detection by the host immune system.


title: Single-cell monitoring of dry mass and dry density reveals exocytosis of cellular dry contents in mitosis

abstract: Cell mass and composition change with cell cycle progression. Our previous work characterized buoyant mass accumulation dynamics in mitosis (Miettinen et al., 2019), but how dry mass and cell composition change in mitosis has remained unclear. To better understand mitotic cell growth and compositional changes, we develop a single-cell approach for monitoring dry mass and the density of that dry mass every [~]75 seconds with 1.3% and 0.3% measurement precision, respectively. We find that suspension grown mammalian cells lose dry mass and increase dry density following mitotic entry. These changes display large, non-genetic cell-to-cell variability, and the changes are reversed at metaphase-anaphase transition, after which dry mass continues accumulating. The change in dry density causes buoyant and dry mass to differ specifically in early mitosis, thus reconciling existing literature on mitotic cell growth. Mechanistically, the dry composition changes do not require mitotic cell swelling or elongation. Instead, cells in early mitosis increase lysosomal exocytosis, and inhibition of exocytosis prevents the dry composition from changing. Overall, our work provides a new approach for monitoring single-cell dry mass and composition and reveals that mitosis is coupled to extensive exocytosis-mediated secretion of cellular contents.


title: Ecological interactions: Patterns of host utilization by tropical butterflies

abstract: Structural complexity of ecological networks facilitate the functional robustness of natural ecosystems. Threatened by the human actions such as habitat destruction and climate change, species may be more or less prone to ecological perturbations depending on the nature of their interactions. We examined the host network of tropical butterflies from the Indian region to see their level of interconnectedness. We manually curated larval host utilization data for 1053 butterflies of India. About 98.8% of species that occur pan-India and 90.6% of species exclusive to the Western Ghats had known hosts whereas it was only 25.9% for species exclusive to north-east India. There were 2589 unique butterfly-host interactions comprising 519 butterfly species and their 1091 known hosts. However, nearly 30% of the species had only single hosts. The Fabaceae and Poaceae were the key host families that accounted for 32.8% of the interactions. There were clear host preferences and monocots hosted disproportionately more butterfly species and interactions. Vanessa cardui had at least 39 known hosts while Ochlandra travancorica supported 19 butterfly species. There were 2693 species-pairs and 4226 interactions among 469 butterflies due to shared hosts. Many butterfly species that have relatively few/unique hosts might be vulnerable in the context of habitat destruction and climate change. This work has great relevance to the ecology and conservation of butterflies in India.

category: ecology


title: Information flow, Gating, and Energetics in dimeric molecular motors

abstract: Molecular motors belonging to the kinesin and myosin super family hydrolyze ATP by cycling through a sequence of chemical states. These cytoplasmic motors are dimers made up of two linked identical monomeric globular proteins. Fueled by the free energy generated by ATP hydrolysis, the motors walk on polar tracks (microtubule or filamentous actin) processively, which means that only one head detaches and executes a mechanical step while the other stays bound to the track. Thus, the one motor head must regulate chemical state of the other, referred to as "gating", a concept that is not fully understood. Inspired by experiments, showing that only a fraction of the energy from ATP hydrolysis is used to advance the kinesin motors against load, we demonstrate that additional energy is used for coordinating the chemical cycles of the two heads in the dimer - a feature that characterizes gating. To this end, we develop a general framework based on information theory and stochastic thermodynamics, and establish that gating could be quantified in terms of information flow between the motor heads. Applications of the theory to kinesin-1 and Myosin V show that information flow occurs, with positive cooperativity, at external resistive loads that are less than a critical value, Fc. When force exceeds Fc, effective information flow ceases. Interestingly, Fc, which is independent of the input energy generated through ATP hydrolysis, coincides with force at which the probability of backward steps starts to increase. Our findings suggest that transport efficiency is optimal only at forces less than Fc, which implies that these motors must operate at low loads under in vivo conditions.


title: A NEUROGENIC GENE EXPRESSION SIGNATURE SUPPORTS HUMAN THERMOGENIC ADIPOSE TISSUE DEVELOPMENT IN VIVO.

abstract: Mechanisms that control "beige/brite" thermogenic adipose tissue development may be harnessed to improve human metabolic health. To define these mechanisms, we developed a species-hybrid model in which human mesenchymal progenitor cells were used to develop white or thermogenic/beige adipose tissue in mice. The hybrid adipose tissue developed distinctive features of human adipose tissue, such as larger adipocyte size, despite its neurovascular architecture being entirely of murine origin. Thermogenic adipose tissue recruited a denser, qualitatively distinct vascular network, differing in genes mapping to circadian rhythm pathways, and denser sympathetic innervation. The enhanced thermogenic neurovascular network was associated with human adipocyte expression of THBS4, TNC, NTRK3 and SPARCL1, which enhance neurogenesis, and decreased expression of MAOA and ACHE, which control neurotransmitter tone. Systemic inhibition of MAOA, which is present in human but absent in mouse adipocytes, induced browning of human but not mouse adipose tissue, revealing the physiological relevance of this pathway. Our results reveal species-specific cell type dependencies controlling the development of thermogenic adipose tissue and point to human adipocyte MAOA as a potential target for metabolic disease therapy.

category: developmental biology


title: A Neurogenic Signature Involving Monoamine Oxidase-A controls Human Thermogenic Adipose Tissue Development


title: Nerve-mediated amputation-induced stem cell activation primes distant appendages for future regeneration events in axolotl

abstract: Animals exhibit extreme diversity in regenerative ability. This likely reflects different, lineage-specific selective pressures in their evolutionary histories, but how specific molecular features of regenerative programs help solve species-specific challenges has not been examined in detail. Here we discover that, in the highly-regenerative axolotl salamander, a conserved, body-wide stem cell activation response triggered in response to limb removal primes undisturbed limbs for regeneration upon subsequent amputation. This response should be particularly useful to salamanders, which frequently lose limbs in response to cannibalism. We further demonstrate the body-wide response requires both peripheral nervous system input at these distant sites and mTOR signaling. We defined gene expression changes within the nerves and nearby tissues, harboring responsive stem cells, leading to identification of candidate genetic pathways influencing distant stem cell activation following amputation. Functional experimentation confirmed a requirement for adrenergic signaling in amputation-induced activation of distant stem cells. These findings reveal a direct link between systemic cellular activation responses to local tissue damage and overall regenerative ability. Similar systemic activation responses to tissue removal have been observed in animals with widely differing regenerative abilities (e.g., planaria to mice), suggesting that it is the responses downstream of these signals, likely sculpted by differing evolutionary pressures, that ultimately distinguish regenerators from non-regenerators.


title: Nipah virus detection at bat roosts following spillover events in Bangladesh, 2012-2019

abstract: Knowledge of the dynamics and genetic diversity of Nipah virus circulating in bats and at the human-animal interface is limited by current sampling efforts, which produce few detections of viral RNA. We report on a series of investigations at bat roosts identified near human Nipah cases in Bangladesh between 2012 and 2019. Pooled bat urine samples were collected from 23 roosts; seven roosts (30%) had at least one sample with Nipah RNA detected from the first visit. In subsequent visits to these seven roosts, RNA was detected in bat urine up to 52 days after the presumed exposure of the human case, although the probability of detection declined rapidly with time. These results suggest that rapidly deployed investigations of Nipah virus shedding from bat roosts near human cases could increase the success of viral sequencing compared to background surveillance and enhance our understanding of Nipah virus ecology and evolution.


title: Effects of periodic bottlenecks on the dynamics of adaptive evolution in microbial populations

abstract:
Population bottlenecks are common in nature, and they can impact the rate of adaptation in evolving populations. On the one hand, each bottleneck reduces the genetic variation that fuels adaptation. On the other hand, fewer founders can undergo more generations and leave more descendants in a resource-limited environment, which allows surviving beneficial mutations to spread more quickly. Here we investigate the impact of repeated bottlenecks on the dynamics of adaptation in experimental populations of Escherichia coli. We propagated 48 populations under four dilution regimes (2-, 8-, 100-, and 1000-fold), all reaching the same final size each day, for 150 days. A simple model in which adaptation is limited by the supply rate of beneficial mutations predicts that fitness gains should be maximized with [~]8-fold dilutions. The model also assumes that selection acts only on the overall growth rate and is otherwise identical across dilution regimes. However, we found that selection in the 2-fold regime was qualitatively different from the other treatments. Moreover, we observed earlier and greater fitness gains in the populations subjected to 100- and 1000-fold dilutions than in those that evolved in the 8-fold regime. We also ran simulations using parameters estimated independently from a long-term experiment using the same ancestral strain and environment. The simulations produced dynamics similar to our empirical results under these regimes, and they indicate that the simple model fails owing to the assumption that the supply of beneficial mutations limits adaptation.

SignificanceMany microorganisms experience population "bottlenecks" during transmission between hosts or when propagated in the laboratory. These bottlenecks reduce genetic variation, potentially impeding natural selection. However, bottlenecks can also increase the number of generations over which selection acts, potentially accelerating adaptation. We explored this tension by propagating 48 bacterial populations for 150 days under regimes in which they were diluted 2-, 8-, 100-, or 1000-fold each day. On balance, the bacteria evolved earlier and adapted faster at the higher dilutions. These results contradict theory that accounts for the loss of genetic variation at higher dilutions, but not for the increased number of generations. We also performed simulations that reflect these opposing factors, and the simulations agree well with our experimental results.


title: Retrovirus-derived acquired genes, RTL5 and RTL6, are novel constituents of the innate immune system in the eutherian brain.

abstract: Retrotransposon Gag-like 5 (RTL5, also known as sushi-ichi-related retrotransposon homolog 8 (SIRH8)) and RTL6 (aka SIRH3) are eutherian-specific genes presumably derived from a retrovirus and phylogenetically related to each other. RTL5 encodes a strongly acidic protein while RTL6 encodes an extremely basic protein, and the former is well conserved and the latter extremely well conserved among the eutherians, indicating their unique and critically important roles as acquired genes. Here we report that RTL5 and RTL6 are microglial genes playing roles in the front line of brain innate immune responses against distinct pathogens. Venus and mCherry knock-in mice exhibited expression of RTL5-mCherry and RTL6-Venus fusion proteins in microglia and as extracellular granules in the central nervus system (CNS), and displayed a rapid response to pathogens such as lipopolysaccharide (LPS), double-stranded (ds) RNA analog and non-methylated CpG DNA. These proteins trapped pathogens in microglia in a variety of RTL-pathogen complexes depending on the pathogens. These results demonstrate that RTL5 and RTL6 exert functional effects against different hazardous substances cooperatively and/or independently to protect the developing and/or mature brain. This provides the first evidence that retrovirus-derived genes play a role in the innate immune system of the eutherian brain.


title: Gut melange a trois: fluctuating selection modulated by microbiota, host immune system, and antibiotics

abstract: Iron is critical in host-microbe interactions, and its availability is tightly regulated in the mammalian gut. Antibiotics and inflammation can perturb iron availability in the gut, which could alter host-microbe interactions. Here, we show that an adaptive allele of iscR, a major regulator of iron homeostasis of Escherichia coli, is under fluctuating selection in the mouse gut. In vivo competitions in immune-competent, immune-compromised, and germ-free mice reveal that the selective pressure on an iscR mutant E. coli is modulated by the presence of antibiotics, the microbiota, and the immune system. In vitro assays show that iron availability is an important mediator of the iscR allele fitness benefits or costs. We identify Lipocalin-2, a hosts immune protein that prevents bacterial iron acquisition, as a major host mechanism underlying fluctuating selection of iscR. Our results provide a remarkable example of strong fluctuating selection acting on bacterial iron regulation in the mammalian gut.


title: Ablation of the FACIT collagen XII disturbs musculoskeletal ECM organization and causes patella dislocation and myopathy

abstract: Collagen XII, belonging to the fibril-associated collagens with interrupted triple helix (FACIT) family, assembles from three identical -chains encoded by the COL12A1 gene. The trimeric molecule consists of three N-terminal noncollagenous NC3 domains joined by disulfide bonds followed by a short interrupted collagen triple helix at the C-terminus. Collagen XII is expressed widely in the musculoskeletal system and mutations in the COL12A1 gene cause an Ehlers-Danlos/myopathy overlap syndrome, which is associated with skeletal abnormalities and muscle weakness. Our study defines the role of collagen XII in patella development using the Col12a1-/- mouse model. Deficiency in Col12a1 expression causes malformed facies patellaris femoris grooves at an early stage, which leads to patella subluxation and growth retardation. Due to the patella subluxation, more muscle fibers with centralized nuclei occur in the quadriceps than in the gastrocnemius muscles indicating a local effect. To further understand the role of collagen XII in the skeletal tissues single cell RNAseq (scRNA-seq) was performed. Comparison of the gene expression in the tenocyte cell sub-population of wild type and Col12a1-/- mice showed that several matrix genes are altered. Finally, we reinvestigated collagen XII deficient patients and observed a patella instability.

category: genetics


title: Single-nuclei isoform RNA sequencing reveals combination patterns of transcript elements across human brain cell types

abstract: Single-nuclei RNA-Seq is being widely employed to investigate cell types, especially of human brain and other frozen samples. In contrast to single-cell approaches, however, the majority of single-nuclei RNA counts originate from partially processed RNA leading to intronic cDNAs, thus hindering the investigation of complete isoforms. Here, using microfluidics, PCR-based artifact removal, target enrichment, and long-read sequencing, we developed single-nuclei isoform RNA-sequencing ( SnISOr-Seq), and applied it to the analysis of human adult frontal cortex samples. We found that exons associated with autism exhibit coordinated and more cell-type specific inclusion than exons associated with schizophrenia or ALS. We discovered two distinct modes of combination patterns: first, those distinguishing cell types in the human brain. These are enriched in combinations of TSS-exon, exon-polyA site, and distant (non-adjacent) exon pairs. Second, those with all isoform combinations found within one neural cell type, which are enriched in adjacent exon pairs. Furthermore, adjacent exon pairs are predominantly mutually associated, while distant pairs are frequently mutually exclusive. Finally, we observed that human-specific exons are as tightly coordinated as conserved exons, pointing to an efficient evolutionary mechanism underpinning coordination. SnISOr-Seq opens the door to single-nuclei long-read isoform analysis in the human brain, and in any frozen, archived or hard-to-dissociate sample.


title: First high-quality genome assembly data of sago palm (Metroxylon sagu Rottboll)

abstract:
The sago palm (Metroxylon sagu Rottboll) is a all-rounder palm, it is both a tropical halophytic starch-producing palm as well as an ornamental plant. Recently, a genome survey was conducted on this palm using Illumina sequencing platform but the BUSCO genome completeness is very low (21.5%) and most of them ([~]78%) are either fragmented or missing. Thus, in this study, the sago palm genome completeness was further improved with the utilization of the Nanopore sequencing platform that produced longer reads. A hybrid genome assembly was conducted and the outcome was a much complete sago palm genome with BUSCO completeness achieved at as high as 97.9% with only [~]2% of them either fragmented or missing. The estimated genome size of the sago palm is 509,812,790 bp in this study. A sum of 33,242 protein-coding genes were revealed from the sago palm genome and around 96.39% of them had been functionally annotated. An investigation on the carbohydrate metabolism KEGG pathways also unearthed that starch synthesis was one of the major sago palm activities. These data are indispensable for future molecular evolutionary and genome-wide association studies.

Specifications Table

O_TBL View this table:
org.highwire.dtl.DTLVardef@1a2dc6aorg.highwire.dtl.DTLVardef@134e433org.highwire.dtl.DTLVardef@171efa9org.highwire.dtl.DTLVardef@12d6f26org.highwire.dtl.DTLVardef@1dc32a6_HPS_FORMAT_FIGEXP  M_TBL C_TBL Value of the DataO_LIFirst complete genome dataset for the eco-economic important sago palm (Metroxylon sagu Rottboll).
C_LIO_LIHigh completeness of sago palm genomic dataset will facilitate future researches, such as genome-wide association studies.
C_LIO_LIThe data is useful in pioneering sago palm genetic landscape investigations which in turn unmask the mystery behind its high starch yield, salinity tolerance and disease resistance.
C_LI


title: Rheology of crossbridge ensembles

abstract: How skeletal muscle responds to mechanical perturbations, its rheology, is important for animal movement control. The molecular machinery of myosin II-actin crossbridge cycling is a crucial part of muscles rheological properties, and multiple models have been proposed for this mechanochemical process. But current understanding of the scale-connection from individual molecular motors to ensemble rheology is limited. Here we present computational and mathematical analyses of several different hypotheses of crossbridge dynamics, from 2-state to 5-state myosin II motor models, and show that an ensemble of actomyosin crossbridges exhibits surprisingly simple rheological behavior in all cases. The ensemble rheology is captured by a sum of at most three linear viscoelastic sub-processes, and as few as one for some crossbridge models. This simplification lends itself to computationally efficient phenomenological muscle models with experimentally measurable parameters, while still remaining grounded in crossbridge theory. However, the collapse of the ensemble behavior to few linear sub-processes identifies major limitations of crossbridge models that cannot be resolved by adding complexity to the crossbridge cycle and point to the roles of inter-crossbridge interactions and non-crossbridge elements.


title: Depletion of CD206+ Tumour Macrophages via a Peptide-Targeted Star-Shaped Polyglutamate Inhibits Tumourigenesis and Metastatic Dissemination in Mouse Breast Cancer Models

abstract: Although many studies have explored the depletion of tumour-associated macrophages (TAMs) as a therapeutic strategy for solid tumours, currently available compounds suffer from poor efficacy and dose-limiting side effects. Here, we developed a novel TAM-depleting agent ("OximUNO") that specifically targets CD206+ TAMs and demonstrated efficacy in triple negative breast cancer (TNBC) mouse models. OximUNO comprises a star-shaped polyglutamate (St-PGA) decorated with the CD206-targeting peptide mUNO that carries the chemotherapeutic drug doxorubicin (DOX). In TNBC models, a fluorescently labelled mUNO-decorated St-PGA homed to CD206+ TAMs within primary lesions and metastases. OximUNO exhibited no acute liver or kidney toxicity in vivo. Treatment with OximUNO reduced the progression of primary tumour lesions and pulmonary metastases, significantly diminished the number of CD206+ TAMs and increased the CD8/FOXP3 expression ratio (demonstrating immunostimulation). Our findings suggest the potential benefit of OximUNO as a TAM-depleting agent for TNBC treatment. Importantly, our studies also represent the first report of a peptide-targeted St-PGA as a targeted therapeutic nanoconjugate.


title: Depletion of CD206+ Tumour Macrophages via a Peptide-Targeted Star-Shaped Polyglutamate Inhibits Tumourigenesis and Metastatic Dissemination in Breast Cancer Models


title: Comparative proteomic analysis of Fusarium oxysporum f. sp. cubense strains (Foc R1 and Foc TR4) provides better insights into mechanisms of their virulence, habitat adaptation and pathogenesis

abstract: Fusarium oxysporum f. sp. cubense (Foc), a devastative soil-borne fungal pathogen causing vascular wilt (i.e. Panama disease) which leads to severe crop losses in most of the banana-growing regions of the world. As there is no single source of effective management practices available so far, understand the pathogenicity of the organism may help in designing effective control measures through molecular approaches. The study aims to compare the proteome of the two pathogenic Foc virulent strains, Race 1 (Foc R1) and tropical race 4 (Foc TR4) that are capable of infecting the Cavendish group of bananas using 2-dimensional (2-D) gel electrophoresis, MALDI-TOF/MS and MS/MS analysis. The results of the study revealed that the proteins, peroxiredoxins, NAD-aldehyde dehydrogenase (NAD-ALDH), MAPK protein, pH-response regulator protein palA/rim-20 and isotrichodermin C15 hydroxylase have shared homology with the fungal proteins, which regulate the osmotic stress response, signal transduction, root colonization and toxin biosynthesis. These are the important functions for the pathogen survival in an unfavourable environment, and successful establishment and infection of the banana host. The present study also identified several putative pathogenicity related proteins in both Foc R1 and Foc TR4. Specifically, certain Foc TR4 specific putative pathogenicity related proteins, phytotoxins biosynthesis gene, fructose 1,6-bisphosphate aldolase class II, Synembryn-like proteins found to contribute strong virulence. Overexpression or knockout of the elective genes could help in devising better control measures for the devastative pathogens in the future. To the best of our knowledge, this is the first report on the proteomics of Foc R1 and Foc TR4 strains of Indian origin that infect Cavendish bananas.

category: pathology


title: BDNF-induced BDNF release mediates presynaptic LTP and is regulated by cannabinoids

abstract:
Although brain-derived neurotrophic factor (BDNF) and its effector, Tropomyosin receptor kinase B (TrkB), are implicated in activity-dependent synaptic plasticity, the precise underlying mechanisms remain unclear. In the dentate gyrus, a hippocampal input region that expresses uniquely high levels of BDNF, repetitive activation of mossy cells (MCs) induces a presynaptic, BDNF/TrkB-dependent form of LTP at MC to granule cell (GC) synapses. Here, we report that activity-induced BDNF release from MC axons in mice elicits postsynaptic BDNF release in a TrkB- and calcium-dependent manner, and that BDNF-induced BDNF release is essential for LTP induction. Suppression of BDNF release by tonic and phasic activity of presynaptic type-1 cannabinoid receptors dampened LTP, revealing an unprecedented role of these receptors in controlling neuropeptide release. Lastly, BDNF-mediated MC-GC LTP can be elicited in vivo. Thus, BDNF-induced BDNF release emerges as a mechanism for activity-dependent LTP that may contribute to dentate gyrus-dependent learning, epilepsy, and neuropsychiatric disorders.

HighlightsO_LIPre- and postsynaptic BDNF are both required for presynaptic LTP in the dentate gyrus
C_LIO_LIBDNF is released from MC axons and GC proximal dendrites following repetitive activation of MC axons
C_LIO_LIPresynaptic BDNF release is inhibited by activation of CB1R expressed on MC axons
C_LIO_LIBDNF is sufficient to trigger postsynaptic BDNF release from GCs in a TrkB- and calcium-dependent manner
C_LIO_LIEnriched environment induces BDNF-dependent LTP of MC-GC synapses in vivo
C_LI

In BriefBerthoux et al. describe a novel mechanism of presynaptic LTP that requires BDNF-induced BDNF release. This BDNF-mediated plasticity can occur in vivo and is tightly controlled by cannabinoid activity.


title: Optogenetic inhibition of the dorsal hippocampus CA3 region during early-stage cocaine-memory reconsolidation disrupts subsequent context-induced cocaine seeking in rats

abstract: The dorsal hippocampus (DH) is key to the long-term maintenance of cocaine memories following retrieval-induced memory destabilization; even though, it is not the site of protein synthesis-dependent memory reconsolidation. Here, we took advantage of the temporal and spatial specificity of an optogenetic manipulation to examine the role of the cornu ammonis 3 subregion of the DH (dCA3) in early-stage cocaine-memory reconsolidation. Male Sprague-Dawley rats expressing eNpHR3.0 in the DH were trained to self-administer cocaine in a distinct context and underwent extinction training in a different context. Rats then received a 15-min memory-reactivation session, to destabilize cocaine memories and trigger reconsolidation, or remained in their home cages (no-reactivation controls). Optogenetic inhibition of the dCA3 for 1 h immediately, but not 1 h, after memory reactivation resulted in cocaine-memory impairment as indicated by reduction in drug-seeking behavior selectively in the cocaine-paired context 3 d later, at test, relative to responding in no-inhibition, no-reactivation, and no-eNpHR3.0 controls. Cocaine-memory impairment was associated with reduced c-Fos expression, an index of neuronal activation, in the dCA3 stratum lucidum (SL) and stratum pyramidale (SP) at test. Based on these observations and extant literature, we postulate that recurrent circuits in the SP are activated during early-stage memory reconsolidation to maintain labile cocaine memories prior to protein synthesis-dependent restabilization in another brain region, such as the basolateral amygdala. Furthermore, SL and SP interneurons may enhance memory reconsolidation by limiting synaptic noise in the SP and also contribute to recall as elements of the updated cocaine engram or retrieval links.


title: Kinetics Of Interferon-{lambda} And Receptor Expression In Response To In Vitro Respiratory Viral Infection

abstract: The major protective immune response against viruses is production of type I and III interferons (IFNs). IFNs induce the expression of hundreds of IFN-stimulated genes (ISGs) that block viral replication and further viral spread. The ability of respiratory viruses to suppress induction of IFN-mediated antiviral defenses in infected epithelial cells may be a factor contributing to the particular pathogenicity of several strains. In this report, we analyzed expression of IFNs and some ISGs in an alveolar epithelial cell subtype (A549) in response to infection with: influenza A viruses (A/California/07/09pdm (H1N1), A/Texas/50/12 (H3N2)); influenza B virus (B/Phuket/3073/13); adenovirus type 5 and 6; or respiratory syncytial virus (strain A2). IFNL and ISGs expression significantly increased in response to infection with all RNA viruses 24 hpi. Nevertheless, only IBV led to early increase in IFNL and ISGs mRNA level. IBV and H1N1 infection led to elevated proinflammatory cytokine production. We speculate that augmented IFN-, IFN-{beta}, IL-6 levels negatively correlate to SOCS1 expression. Importantly, we showed a decrease in IFNLR1 mRNA in case of IBV infection that implies the existence of negative ISGs expression regulation at IFN{lambda}R level. It could be either a specific feature of IBV or a consequence of early IFNL expression.


title: Reconstitution of the S. aureus agr quorum sensing pathway reveals a direct role for the integral membrane protease MroQ in pheromone biosynthesis

abstract: In Staphylococcus aureus, virulence is under the control of a quorum sensing (QS) circuit encoded in the accessory gene regulator (agr) genomic locus. Key to this pathogenic behavior is the production and signaling activity of a secreted pheromone, the autoinducing peptide (AIP), generated following the ribosomal synthesis and post-translational modification of a precursor polypeptide, AgrD, through two discrete cleavage steps. The integral membrane protease AgrB is known to catalyze the first processing event, generating the AIP biosynthetic intermediate, AgrD (1-32) thiolactone. However, the identity of the second protease in this biosynthetic pathway, which removes an N-terminal leader sequence, has remained ambiguous. Here, we show that MroQ, an integral membrane protease recently implicated in the agr response, is directly involved in AIP production. Genetic complementation and biochemical experiments reveal that MroQ proteolytic activity is required for AIP biosynthesis in agr specifiy groups -I and -II, but not group-III. Notably, as part of this effort, the biosynthesis and AIP-sensing arms of the QS circuit were reconstituted together in vitro. Our experiments also reveal the molecular features guiding MroQ cleavage activity, a critical factor in defining agr specificity group identity. Collectively, our study adds to the molecular understanding of the agr response and Staphylococcus aureus virulence.


title: Pronounced sequence specificity of the TET enzyme catalytic domain guides its cellular function

abstract:
TET (ten-eleven translocation) enzymes catalyze the oxidation of 5-methylcytosine bases in DNA, thus driving active and passive DNA demethylation. Here, we report that the catalytic cores of mammalian TET enzymes favor CpGs embedded within bHLH and bZIP transcription factor binding sites, with 250-fold preference in vitro. Crystal structures and molecular dynamics calculations show that sequence preference is caused by intra-substrate interactions and CpG flanking sequence indirectly affecting enzyme conformation. TET sequence preferences are physiologically relevant as they explain the rates of DNA demethylation in TET-rescue experiments in culture and in vivo within the zygote and germline. Most and least favorable TET motifs represent DNA sites that are bound by methylation-sensitive immediate-early transcription factors and OCT4, respectively, illuminating TET function in transcriptional responses and pluripotency support.

One-Sentence SummaryThe catalytic domains of the enzymes that facilitate passive and drive active DNA demethylation have intrinsic sequence preferences that target DNA demethylation to bHLH and bZIP transcription factor binding sites.

category: molecular biology


title: iTRVZ: liquid nano-platform for signal integration on the plasma membrane

abstract:
Crosstalk of cellular signaling pathways is essential for coordinated cell responses. Therefore, we suspect the existence of scaffolding platforms that specifically undertake signal crosstalk and integration. Using advanced single-molecule imaging, we found a nanometer-scale liquid-like platform for integrating the signals downstream from GPI-anchored receptors and receptor-type tyrosine kinases. The platform employs some of the focal adhesion proteins, including integrin, talin, RIAM, VASP, and zyxin, but is distinct from focal adhesions, and is thus termed iTRVZ. The iTRVZ formation is driven by the protein liquid-liquid phase separation and the interactions with the raft domains in the plasma membrane and cortical actin. iTRVZ non-linearly integrates the two distinctly different receptor signals, and thus works as an AND gate and noise filter.

One-Sentence SummaryNanometer-scale liquid-like protein platform for signal integration working as an AND gate was found on the plasma membrane.


title: Learning the histone codes of gene regulation with large genomic windows and three-dimensional chromatin interactions using transformer

abstract: The quantitative characterization of the transcriptional control by histone modifications (HMs) has been challenged by many computational studies, but still most of them exploit only partial aspects of intricate mechanisms involved in gene regulation, leaving a room for improvement. We present Chromoformer, a new transformer-based deep learning architecture that achieves the state-of-the-art performance in the quantitative deciphering of the histone codes of gene regulation. The core essence of Chromoformer architecture lies in the three variants of attention operation, each specialized to model individual hierarchy of three-dimensional (3D) transcriptional regulation including (1) histone codes at core promoters, (2) pairwise interaction between a core promoter and a distal cis-regulatory element mediated by 3D chromatin interactions, and (3) the collective effect of the pairwise cis-regulations. In-depth interpretation of the trained model behavior based on attention scores suggests that Chromoformer adaptively exploits the distant dependencies between HMs associated with transcription initiation and elongation. We also demonstrate that the quantitative kinetics of transcription factories and polycomb group bodies, in which the coordinated gene regulation occurs through spatial sequestration of genes with regulatory elements, can be captured by Chromoformer. Together, our study shows the great power of attention-based deep learning as a versatile modeling approach for the complex epigenetic landscape of gene regulation and highlights its potential as an effective toolkit that facilitates scientific discoveries in computational epigenetics.