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Sarkar, S.

Publications and source records attributed to Sarkar, S..

At least 145 records · Page 8Linked to original sources

Home-field advantage affects the local adaptive interaction between Andropogon gerardii ecotypes and rhizobiome

Due to climate change, drought frequencies and severities are predicted to increase across the United States. Plant responses and adaptation to stresses depend on plant genetic and environmental factors. Understanding the effect of those factors on plant performance is required to predict the species responses to environmental change. We used reciprocal gardens planted with distinct regional Andropogon gerardii ecotypes adapted to dry, mesic, and wet environments to characterize their rhizosphere communities using 16S rRNA metabarcode sequencing. Even though the local microbial pool was the main driver of these rhizosphere communities, the significant plant ecotype effect highlighted active microbial recruitment in the rhizosphere driven by ecotype or plant genetic background. Our data also suggest that ecotypes were more successful in recruiting rhizosphere community members unique to their local homesites, supporting the "home field advantage" hypothesis. These unique homesite microbes may represent microbial specialists that are linked to plant stress responses. Further, our data support ecotypic variation in the recruitment of congeneric but distinct bacterial variants, highlighting the nuanced effects of plant ecotypes on the rhizosphere microbiome recruitment. Our results should facilitate expanded studies on understanding the complexity of plant host interactions with local soil microbes and identification of functional potential of recruited microbes. Our study has the potential to aid in predicting ecosystem responses to climate change and the impact of management on restoration practices. ImportanceIn this study, we used reciprocal gardens located across a sharp precipitation gradient to characterize rhizosphere communities of distinct dry, mesic, and wet regional Andropogon gerardii ecotypes. We used16S rRNA amplicon sequencing and focused oligotyping analysis and showed that even though the location was the main driver of the microbial communities, ecotypes could potentially recruit distinct bacterial populations. We showed that different A. gerardii ecotypes were more successful in overall community recruitment and recruitment of microbes unique to the "home" environment, when growing at their "home site". We found evidence for "home field advantage" interactions between the host and associated rhizobiomes, and the capability of ecotypes to recruit specialized microbes that were potentially linked to plant stress responses. Our study provides insights into the understanding of factors effecting the plant adaptation, improving management strategies, and predicting of the future landscape under the changing climate.

microbiology↗

Identifying common genes, proteins, and pathways from human miRNA and gene blood profiles in multiple sclerosis patients.

The molecular pathway associated with Multiple sclerosis (MS) is complex and symptomatic treatments are only available right now. Early diagnosis of MS creates a window for healthcare providers to manage the disease more efficiently. Blood-based biomarker study has been done in the past to identify the upregulated and downregulated genes but in this present study, a novel approach has been taken for identifying genes associated with the disease. In this present study, hub genes are identified and the top ten hub genes were used to identify drugs associated with them. Upregulated genes were identified using the dataset GSE21942 (which contains information related to genes identified in the blood of multiple sclerosis patients) and datasets GSE17846 and GSE61741(which contains information related to microRNAs taken from multiple sclerosis patients). Genes associated with microRNAs were identified using miRWalk. Common genes from both miRWalk and the dataset GSE21942 were identified and were subjected to STRINGdb for the creation of a protein-protein interaction network and this network was then imported to Cytoscape for identifying the top ten hub genes. The top ten hub genes were subjected to EnrichR for enrichment analysis of genes. In our study, it was found that CTNNB1 is the gene with the highest degree (116).

neuroscience↗

Fc mediated pan-sarbecovirus protection after alphavirus vector vaccination

Two group 2B {beta}-coronaviruses (sarbecoviruses) have caused regional and global epidemics in modern history. The mechanisms of cross protection driven by the sarbecovirus spike, a dominant immunogen, are less clear yet critically important for pan-sarbecovirus vaccine development. We evaluated the mechanisms of cross-sarbecovirus protective immunity using a panel of alphavirus-vectored vaccines covering bat to human strains. While vaccination did not prevent virus replication, it protected against lethal heterologous disease outcomes in both SARS-CoV-2 and clade 2 bat sarbecovirus HKU3-SRBD challenge models. The spike vaccines tested primarily elicited a highly S1-specific homologous neutralizing antibody response with no detectable cross-virus neutralization. We found non-neutralizing antibody functions that mediated cross protection in wild-type mice were mechanistically linked to FcgR4 and spike S2-binding antibodies. Protection was lost in FcR knockout mice, further supporting a model for non-neutralizing, protective antibodies. These data highlight the importance of FcR-mediated cross-protective immune responses in universal pan-sarbecovirus vaccine designs.

microbiology↗

IPLS-LDA: An Improved Partial Least Square Discriminant Analysis for Heterogeneous Transcriptomics and Metabolomics Data Analysis

Supervised machine learning (SML) is an approach that learns from training data with known category membership to predict the unlabeled test data. There are many SML approaches in the literature and most of them use a linear score to learn its classifier. However, these approaches fail to elucidate biodiversity from heterogeneous biomedical data. Therefore, their prediction accuracies become low. Partial Least Square Linear Discriminant Analysis (PLS-LDA) is widely used in gene expression (GE) and metabolomics datasets for predicting unlabelled test data. Nevertheless, it also does not consider the non-linearity and heterogeneity pattern of the datasets. Hence, in this study, an improved PLS-LDA (IPLS-LDA) was developed by capturing the heterogeneity of datasets through an unsupervised hierarchical clustering approach. In our approach a non-linear score was calculated by combining all the linear scores obtained from the clustering method. The performance of IPLS-LDA was investigated in a comparison with six frequently used SML methods (SVM, LDA, KNN, Naive Bayes, RF, PLS-LDA) using one simulation data, one colon cancer gene expression data (GED) and one lung cancer metabolomics datasets. The resultant IPLS-LDA predictor achieved accuracy 0.841 using 10-fold cross validation in colon cancer data and accuracy 0.727 from two independent metabolomics data analysis. In both the cases IPLS-LDA outperformed other SML predictors. The proposed algorithm has been implemented in an R package, Uplsda was given in the https://github.com/snotjanu/UplsLda.

bioinformatics↗

Limitation of sulfur-containing amino acid availability by specific bacterial populations during enhanced colitis in IBD mouse model

Members of the Enterobacteriaceae family including Escherichia coli are associated with persistent gut inflammation during disorders like inflammatory bowel disease. This is due to rapid microbial colonization during dysbiosis combined with pathogenic tendencies. We characterized the dysbiotic gut community, defined potential functional pathways, and investigated crosstalk between host gene expression and microbial detections in an intestinal inflammation murine model. Members of Enterobacteriaceae family and the Enterococcus genus were highly detected in dysbiotic mice. These metagenome assembled genomes (MAGs) contained several virulence factors and metabolic pathways necessary to drive perpetual inflammation. Two Enterobacteriaceae MAGs with L-cysteine and L-taurine dioxygenases were strongly correlated with upregulation of the host gene CSAD, responsible for cysteine metabolism. Suggesting these bacteria compete with the host to utilize essential amino acids. We observed that bacterial isolates from dysbiotic mice displayed increased growth rates supplemented with L-cysteine, confirming that these microbes can utilize host nutrients to sustain inflammation. In BriefInflammatory bowel disease is associated with an increase in Enterobacteriaceae and Enterococcus species, however the mechanisms are unclear. Richie et al. show that these bacterial populations use sulfur metabolism and tolerate host-derived immune-response, to drive host inflammation and fuel growth in the dysbiotic colon. Cultured isolates from dysbiotic mice indicated faster growth supplemented with L-cysteine, showing these microbes can utilize these essential host nutrients. HighlightsO_LIMice receiving native microbial FMT showed lower colon inflammation scores, higher microbial diversity, detections and gene expression similar to control mice. C_LIO_LIDysbiotic mice displayed increased colon inflammation, higher detection of potential pathogenic MAGs, and upregulation of cysteine dioxygenase and other inflammation response genes C_LIO_LIMAGs assigned to Enterococcus and Enterobacteriaceae species were more frequently detected in dysbiotic mice, while almost absent in mice receiving FMT or control mice, they also contain several virulence factors and antibiotic resistance genes. C_LIO_LIThese MAGs also display potential functions of utilizing host products and nutrients including nitrate, cysteine, and taurine to further fuel their growth and metabolism, which results in persistent host intestinal inflammation. C_LIO_LIIsolates in the Enterobacteriaceae family from dysbiotic mice utilize L-cysteine for growth, whereas isolates from FMT and control mice show no significant difference, indicating these bacteria can utilize the host derived cysteine. C_LI

genomics↗

Meta-analysis refinement of plasma extracellular vesicle composition identifies proplatelet basic protein as a signaling messenger in type 1 diabetes

Extracellular vesicles (EVs) play important roles in cell-to-cell communication and are potential biomarkers as they carry markers of their derived tissues and disease signatures. However, obtaining pure EV preparations from biofluids is challenging due to contaminants with similar physicochemical properties. Here, we performed a meta-analysis of plasma EV proteomics data deposited in public repositories to refine the protein composition of EVs and investigate potential roles in type 1 diabetes development. With the concept that each purification method yields different proportions of distinct contaminants, we grouped proteins into clusters based on their abundance profiles. This allowed us to separate clusters with classical EV markers, such as CD9, CD40, C63 and CD81, from clusters of well-known contaminants, such as serum albumin, apolipoproteins and components of the complement and coagulation pathways. Two clusters containing a total of 1720 proteins combined were enriched with EV markers and depleted in common contaminants; therefore, they were considered to contain bona fide EV components. As possible origins of plasma EVs, these clusters had markers of tissues such as spleen, liver, brain, lungs, pancreas, and blood/immune cells. These clusters were also enriched in cell surface markers CD antigens, and proteins from cell-to-cell communication and signaling pathways, such as chemokine signaling and antigen presentation. We also show that the EV component and type 1 diabetes biomarker, platelet basic protein (PPBP/CXCL7) regulates apoptosis in both beta and macrophage cell lines. Overall, our meta-analysis refined the composition of plasma EVs, reinforcing a primary function as messengers for cell-to-cell communication and signaling. Furthermore, this analysis identifies optimal avenues to target EVs for development of disease biomarkers.

cell biology↗

Enhancer Reprogramming in Melanoma Immune Checkpoint Therapy Resistance

Immune checkpoint blockade (ICB) therapy has improved long-term survival for patients with advanced melanoma. However, there is critical need to identify potential biomarkers of response and actionable strategies to improve response rates. Through generation and analysis of 148 chromatin modification maps for 36 melanoma samples from patients treated with anti-PD- 1, we identified significant enrichment of active enhancer states in non-responders at baseline. Analysis of an independent cohort of 20 samples identified a set of 437 enhancers that predicted response to anti-PD-1 therapy (Area Under the Curve of 0.8417). The activated non-responder enhancers marked a group of key regulators of several pathways in melanoma cells (including c- MET, TGF{beta}, EMT and AKT) that are known to mediate resistance to ICB therapy and several checkpoint receptors in T cells. Epigenetic editing experiments implicated involvement of c-MET enhancers in the modulation of immune response. Finally, inhibition of enhancers and repression of these pathways using bromodomain inhibitors along with anti-PD-1 therapy significantly decreased melanoma tumor burden and increased T-cell infiltration. Together, these findings identify a potential enhancer-based biomarker of resistance to anti-PD-1 and suggest enhancer blockade in combination with ICB as a potential strategy to improve responses.

cancer biology↗

Genetically-Encoded Discovery of Perfluoroaryl-Macrocycles that Bind to Albumin and Exhibit Extended Circulation in-vivo

In this paper, we report selection of albumin-binding macrocyclic peptides from genetically encoded libraries of peptides modified by perfluoroaryl-cysteine SNAr chemistry. Modification of phage-displayed libraries SXCXnC-phage, n=3-5, where X is any amino acid except for cysteine by decafluoro-diphenylsulfone (DFS), yields genetically-encoded library of octafluoro-diphen-ylsulfone-crosslinked macrocycles (OFS-SXCXnC-phage). Selection from these libraries using albumin as a bait identified a family of significantly enriched perfluoroaryl-macrocycles. Synthesis of perfluoroaryl-macrocycles predicted by phage display and testing their binding properties by 19F NMR and fluorescent polarization identified OFS-macrocycle with SICRFFC sequence as the most potent albumin binder. We observed that OFS-macrocycles slowly react with biological nucleophiles such as glutathione. Replacing decafluoro-diphenylsulfone by nearly isosteric pentafluorophenyl sulfide yielded perfluorophenylsulfide (PFS)-crosslinked macrocycles devoid of undesired reactivity. The augmented lead PFS-macrocycle with SICRFFC sequence exhibited KD = 4-6 M towards human serum albumin and similar affinities towards rat and mouse albumins. When injected in mouse, the PFS-SICRFFCGGG compound was significantly retained in circulation in vivo when compared to control PFS-macrocyclic peptide. The perfluoroaryl-macrocycles with SICRFFC motif are the smallest known peptide macrocycle with significant affinity for human albumin and they are a productive starting point for future development of compact macrocycles with predictable circulation half-life in vivo. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=67 SRC="FIGDIR/small/504611v1_ufig1.gif" ALT="Figure 1"> View larger version (17K): org.highwire.dtl.DTLVardef@cd54d2org.highwire.dtl.DTLVardef@7bd9aborg.highwire.dtl.DTLVardef@10f7515org.highwire.dtl.DTLVardef@1cfc49a_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

Chemoenzymatic Synthesis of Genetically-Encoded Multivalent Liquid N-glycan Arrays

A hallmark of cellular glycosylation is its chemical complexity and heterogeneity, which can be challenging to capture synthetically. Using chemoenzymatic synthesis on M13 phage, we produce a genetically-encoded liquid glycan array (LiGA) of biantennary complex type N-glycans. Ligation of azido-functionalized sialylglycosyl-asparagine derived from egg yolk to phage functionalized with 50-1000 copies of dibenzocyclooctyne produced divergent intermediate that can be trimmed by glycosidases and extended by glycosyltransferases to yield a library of phages with different N-glycans. Post-reaction analysis by MALDI-TOF MS provided a rigorous approach to confirm N-glycan structure and density, both of which were encoded in the bacteriophage DNA. The binding of this N-glycan LiGA by ten lectins, including CD22 or DC-SIGN expressed on live cells, uncovered an optimal structure/density combination for recognition. Injection of the LiGA into mice identified glycoconjugates with structures and avidity necessary for enrichment in specific organs. This work provides an unprecedented quantitative evaluation of the interaction of complex N-glycans with GBPs in vitro and in vivo.

biochemistry↗

Culturomics of Andropogon gerardii rhizobiome revealed nitrogen transforming capabilities of stress-tolerant Pseudomonas under drought conditions

BackgroundClimate change will result in more frequent droughts that impact soil-inhabiting microbiomes in the agriculturally vital North American perennial grasslands. In this study, we used the combination of culturomics and high-resolution genomic sequencing of microbial consortia isolated from the rhizosphere of a tallgrass prairie foundation grass, Andropogon gerardii. We cultivated the plant host-associated microbes under artificial drought-induced conditions and identified the microbe(s) that might play a significant role in the rhizobiome of Andropogon gerardii under drought conditions. ResultsPhylogenetic analysis of the non-redundant metagenome-assembled genomes (MAGs) identified the bacterial population of interest - MAG-Pseudomonas. Further metabolic pathway and pangenome analyses detected genes and pathways related to nitrogen transformation and stress responses in MAG-Pseudomonas. ConclusionsOur data indicate that the metagenome-assembled MAG-Pseudomonas has the functional potential to contribute to the plant hosts growth during stressful conditions. This study provided insights into optimizing plant productivity under drought conditions.

microbiology↗

Nonenzymatic template-directed replication using 2'-3' cyclic nucleotides under wet-dry cycles

RNA World Hypothesis is centred around the idea of a period in the early history of lifes origin, wherein nonenzymatic oligomerization and replication of RNA resulted in functional ribozymes. Previous studies in this endeavour have demonstrated template-directed replication using chemically modified nucleotides and primers. Nonetheless, similar studies that used non-activated nucleotides led to the formation of RNA only with abasic sites. In this study, we report template-directed replication with prebiotically relevant cyclic nucleotides, under dehydration-rehydration (DH-RH) cycles occurring at high temperature (90{degrees}C) and alkaline conditions (pH 8). 2'-3' cyclic nucleoside monophosphates (cNMP) resulted in primer extension, while 3'-5' cNMP failed to do so. Intact extension of up to two nucleotide additions was observed with both canonical hydroxy-terminated (OH-primer) and activated amino-terminated (NH2-primer) primers. We demonstrate primer extension reactions using both purine and pyrimidine 2'-3' cNMPs, with higher product yield observed during cAMP additions. Further, the presence of lipid was observed to significantly enhance the extended product in cCMP reactions. In all, our study provides a proof-of-concept for nonenzymatic replication of RNA, using intrinsically activated prebiotically relevant cyclic nucleotides as monomers.

biochemistry↗

Ageing leads to nonspecific antimicrobial peptide responses in Drosophila melanogaster

Evolutionary theory predicts a late-life decline in the force of natural selection, possibly leading to late-life deregulations of the immune system. A potential outcome of such immune-deregulation is the inability to produce specific immunity against target pathogens. We tested this possibility by infecting multiple Drosophila melanogaster lines (with bacterial pathogens) across age-groups, where either individual or different combinations of Imd- and Toll-inducible antimicrobial peptides (AMPs) were deleted using CRISPR gene editing. We show a high degree of non-redundancy and pathogen-specificity of AMPs in young flies: in some cases, even a single AMP could confer complete resistance. In contrast, ageing led to a complete loss of such specificity, warranting the action of multiple AMPs across Imd- and Toll-pathways during infections. Moreover, use of diverse AMPs either had no survival benefits, or even accompanied survival costs post-infection. These features were also sexually dimorphic: females expressed a larger repertoire of AMPs than males, but extracted equivalent survival benefits. Finally, age-specific expansion of the AMP-pool was associated with downregulation of negative-regulators of the Imd-pathway and a potential damage to renal function, as features of poorly-regulated immunity, Overall, we could establish ageing as an important driver of nonspecific AMP responses, across sexes and bacterial infections.

immunology↗

To βor not to β: Lack of correlation between APC mutation and β-catenin nuclear localization in colorectal cancer

Colorectal cancer (CRC) appears to arise from sequential genetic lesions in tumor suppressor genes (APC, SMAD4 and TP53) and oncogenes (KRAS) leading to the classical adenoma to carcinoma progression. Biallelic APC inactivating genetic aberrations are detected in about 70% of early microadenomas implicating APC inactivation as the first genetic hit in CRC. APC is an essential protein of the Wnt destruction complex; APC inactivation is believed to cause disruption of the complex allowing stabilization and nuclear translocation of {beta}-catenin, resulting in transcriptional activation of cancer-promoting genes. Here, we provide evidence for a surprising lack of correlation between APC mutation and {beta}-catenin nuclear localization in early-onset sporadic rectal cancer samples. {beta}-catenin nuclear localization and APC mutation were validated from serial FFPE sections representing same tumor regions. Several factors including status of KRAS mutation could not explain the absence of {beta}-catenin nuclear localization in tumor samples harboring APC mutation. This lack of correlation was validated in CRC cell lines harboring various APC mutations. Our results provide evidence directly from tumor samples for possible non-canonical role(s) for mutant APC.

cancer biology↗

pH-responsive self-assembled compartments as tuneable model protocellular membrane systems

Prebiotically plausible single-chain amphiphiles are enticing as model protocellular compartments to study the emergence of cellular life owing to their self-assembling properties. Here, we investigated the self-assembly behaviour of mono-N-dodecyl phosphate (DDP) and mixed systems of DDP with dodecyl 1-dodecanol (DOH) at varying pH conditions. Membranes composed of DDP showed pH-responsive vesicle formation in a wide range of pH with a low critical bilayer concentration (CBC). Further, the addition of DOH to DDP membrane system enhanced vesicle formation and stability in alkaline pH regimes. We also compared the high-temperature behaviour of DDP and DDP-DOH membranes with conventional fatty acid membranes. Both, DDP and DDP:DOH mixed membranes possess packing that is similar to decanoic acid membrane. However, the micropolarity of these systems are similar to phospholipid membranes. Finally, the pH-dependent modulation of different phospholipid membranes doped with DDP was also demonstrated to engineer tuneable membranes with potential translational implications.

biochemistry↗

Potent universal-coronavirus therapeutic activity mediated by direct respiratory administration of a Spike S2 domain-specific human neutralizing monoclonal antibody

Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) marks the third novel {beta}-coronavirus to cause significant human mortality in the last two decades. Although vaccines are available, too few have been administered worldwide to keep the virus in check and to prevent mutations leading to immune escape. To determine if antibodies could be identified with universal coronavirus activity, plasma from convalescent subjects was screened for IgG against a stabilized pre-fusion SARS-CoV-2 spike S2 domain, which is highly conserved between human {beta}-coronavirus. From these subjects, several S2-specific human monoclonal antibodies (hmAbs) were developed that neutralized SARS-CoV-2 with recognition of all variants of concern (VoC) tested (Beta, Gamma, Delta, Epsilon, and Omicron). The hmAb 1249A8 emerged as the most potent and broad hmAb, able to recognize all human {beta}-coronavirus and neutralize SARS-CoV and MERS-CoV. 1249A8 demonstrated significant prophylactic activity in K18 hACE2 mice infected with SARS-CoV-2 lineage A and lineage B Beta, and Omicron VoC. 1249A8 delivered as a single 4 mg/kg intranasal (i.n.) dose to hamsters 12 hours following infection with SARS-CoV-2 Delta protected them from weight loss, with therapeutic activity further enhanced when combined with 1213H7, an S1-specific neutralizing hmAb. As little as 2 mg/kg of 1249A8 i.n. dose 12 hours following infection with SARS-CoV Urbani strain, protected hamsters from weight loss and significantly reduced upper and lower respiratory viral burden. These results indicate in vivo cooperativity between S1 and S2 specific neutralizing hmAbs and that potent universal coronavirus neutralizing mAbs with therapeutic potential can be induced in humans and can guide universal coronavirus vaccine development.

immunology↗

Structural Basis of HIV-1 Maturation Inhibitor Binding and Activity

HIV-1 maturation inhibitors (MIs) interfere with the final step in the viral lifecycle by disrupting the ordered proteolytic processing of the viral Gag polyprotein into its individual domains. Bevirimat (BVM) and its analogs interfere with the final catalytic cleavage of spacer peptide 1 (SP1) from the capsid protein (CA) C-terminal domain (CACTD), by binding to and stabilizing the CACTD-SP1 region. MIs are under development as alternative drugs to augment current antiretroviral therapies. Although promising, their mechanism of action and associated virus resistance pathways remain poorly understood at the molecular, biochemical, and structural levels. Here, we report atomic-resolution magic angle spinning (MAS) NMR structures of microcrystalline assemblies of CACTD-SP1 complexed with BVM and/or the assembly cofactor inositol hexakisphosphate (IP6). BVM and IP6 can bind simultaneously to SP1, with BVM positioned in the center of its 6-helix bundle in a unique conformation. Importantly, the NMR-observed structural effects of BVM on IP6 binding suggest that the inhibitor stabilizes the 6-helix bundle in multiple ways. In addition, BVM-resistant SP1-A1V and SP1-V7A variants exhibit distinct conformational and binding characteristics. Taken together, our results reveal a novel allosteric mechanism by which BVM disrupts maturation and provide a structural explanation for BVM resistance as well as important guidance for the design of new MIs.

biophysics↗

CriSNPr: a single interface for the curated and de-novo design of gRNAs for CRISPR diagnostics using diverse Cas systems

Nucleic acid detection and variant calling through CRISPR-based diagnostics (CRISPRDx) has facilitated clinical decision-making, particularly during the COVID-19 pandemic. This has been further accelerated through the discovery of newer and engineered CRISPR effectors, expanding the portfolio of such diagnostic applications to a wide variety of pathogenic and non-pathogenic conditions. However, each diagnostic CRISPR pipeline requires customized detection schemes originating from fundamental principles of the Cas protein used, its guide RNA (gRNA) design parameters, and the assay readout. This is particularly relevant for variant detection, an attractive low-cost alternative to sequencing-based approaches for which no in silico pipeline for the ready-to-use design of CRISPR-based diagnostics currently exists. In this manuscript, we fill this lacuna using a unified webserver CriSNPr (CRISPR based SNP recognition), which provides the user the opportunity to de-novo design gRNAs based on six CRISPRDx proteins of choice (Fn/enFnCas9, LwCas13a, LbCas12a, AaCas12b, and Cas14a) and query for ready-to-use oligonucleotide sequences for validation on relevant samples. In addition, we provide a database of curated pre-designed gRNAs and target/off-target for all human and SARS-CoV-2 variants reported so far. CriSNPr has been validated on multiple Cas proteins and highlights its broad and immediate scope of utilization across multiple detection platforms. CriSNPr is available at URL http://crisnpr.igib.res.in/.

bioinformatics↗

M2 Receptors are required for spatiotemporal sequence learning in mouse primary visual cortex

Acetylcholine is a neurotransmitter that plays a variety of roles in the central nervous system. It was previously shown that blocking muscarinic receptors with a non-selective antagonist prevents a form of experience-dependent plasticity termed "spatiotemporal sequence learning" in the mouse primary visual cortex (V1). Muscarinic signaling is a complex process involving the combined activities of five different G-protein coupled receptors, M1-M5, all of which are expressed in the murine brain but differ from each other functionally and in anatomical localization. Here we present electrophysiological evidence that M2, but not M1, receptors are required for spatiotemporal sequence learning in mouse V1. We show in male mice that M2 is highly expressed in the neuropil in V1, especially in thalamorecipient layer 4, and co-localizes with the soma in a subset of somatostatin expressing neurons in deep layers. We also show that expression of M2 receptors is higher in the monocular region of V1 than it is in the binocular region, but that the amount of experience-dependent sequence potentiation is similar in both regions, and that blocking muscarinic signaling after visual stimulation does not prevent plasticity. This work establishes a new functional role for M2-type receptors in processing temporal information and demonstrates that monocular circuits are modified by experience in a manner similar to binocular circuits. Significance StatementMuscarinic acetylcholine receptors are required for multiple forms of plasticity in the brain and support perceptual functions, but the precise role of the five subtypes (M1-M5) are unclear. Here we show that the M2 receptor is specifically required to encode experience-dependent representations of spatiotemporal relationships in both monocular and binocular regions of mouse V1. Muscarinic receptors are metabotropic in nature and have complex downstream signaling effects that can be difficult to isolate experimentally. In addition to identifying a novel functional role for M2 receptors, this work also suggests that V1 circuits can be used as an in-vivo model to understand the relationship between cholinergic signaling and coding of complex temporal relationships in cortical circuits.

neuroscience↗