Biology subjects

Arora, N.

Publications and source records attributed to Arora, N..

At least 19 recordsLinked to original sources

bioRxiv2026

Microbial community profiles of genital and non-genital skin lesions from Ghana

Skin ulcers, such as those observed in cases of the non-venereal disease yaws and the venereal disease syphilis, are traditionally diagnosed based on their anatomical location and visible appearance. Yet various evidence suggests that the pathogens that cause these ulcers may not be restricted to genital or non-genital body sites. We explored this by profiling the bacterial communities of genital and non-genital skin ulcers collected in Ghana using 16S rRNA gene amplicon sequencing. Swab samples were collected from 156 clinically diagnosed skin lesions (110 non-genital and 46 genital). The V1-V2 region of the 16S rRNA gene was sequenced to characterize bacterial communities. Microbial diversity, community composition, differential abundance, and associations between lesion diversity and common skin bacteria were analysed using bioinformatics and statistical methods. Following quality filtering, 135 samples were retained, revealing 23 bacterial phyla, 143 families, and 406 genera. Alpha diversity indicated similar bacterial richness and diversity in genital and non-genital skin lesions. Beta diversity analyses indicated differences between genital and non-genital lesions, accompanied by significant differences in within group dispersion. A log2 fold change analysis evidenced genital lesions were enriched in Gardnerella, whereas non-genital lesions showed more abundance of Staphylococcus, Streptococcus, Corynebacterium, Bradyrhizobium, and Pelomonas. Clinically important genera, including Treponema, Haemophilus, and Mycobacterium, were also detected, even in samples with low sequencing depths. Statistical modelling indicated that greater microbial diversity was associated with intermediate abundances of common skin taxa, particularly Corynebacteriaceae, Micrococcaceae, Cutibacterium, and Staphylococcaceae, thereby attesting the presence of polymicrobial communities in skin ulcers. Overall, genital and non-genital skin lesions showed similar microbial diversity but differed in composition. Our findings support the view of skin lesions as ecological niches containing diverse bacterial communities rather than being consistently dominated by a single bacterial taxon. They provide a basis for investigating possible polymicrobial interactions in skin ulcers.

microbiology↗

bioRxiv2026

Lipid acyl chain remodeling governs KRAS nanoclustering and oncogenic functions through membrane composition control

KRAS signaling is spatially organized within the dynamic proteolipid nanoclusters on the plasma membrane (PM) that selectively concentrate anionic phospholipids, yet the molecular determinants governing this lipid sorting remain incompletely defined. Here we identify phospholipid acyl chain composition as a critical regulator of KRAS spatial organization and signal output. Using genetic and pharmacological perturbation of lysophosphatidylcholine acyltransferase 3 (LPCAT3), a key enzyme that incorporates polyunsaturated fatty acids into phospholipids, we demonstrate that remodeling of membrane acyl chain unsaturation alters the phosphatidylserine (PS) lipidome without changing total PS abundance. Loss of LPCAT3 depletes polyunsaturated PS species, disrupts KRAS PM association and nanoclustering, and attenuates downstream MAPK signaling and oncogenic activity across multiple KRAS mutant contexts. Acute restoration of polyunsaturated, but not saturated, lipids rescue KRAS nanoclustering, establishing a direct requirement for lipid unsaturation in sustaining signaling-competent assemblies. Mechanistically, these findings define membrane acyl chain composition as a determinant of proteolipid organization that encodes KRAS signaling capacity. Consistent with this model, LPCAT3 inhibition selectively impairs KRAS-dependent proliferation and sensitizes tumor cells to direct KRAS inhibition. Together, this work uncovers lipid acyl chain remodeling as a fundamental regulator of membrane-associated signaling and identifies membrane composition as an exploitable vulnerability in oncogenic KRAS systems.

cancer biology↗

bioRxiv2026

HuMMANet: A Harmonized Cross-Study Resource for Integrative Analysis of Human Gut Microbiome Metabolome Associations

Deciphering gut microbiome to host metabolome interaction is critical for understanding how microbial communities generate bioactive signals that shape host physiology and disease. Progress, however, has been hindered by inconsistent metabolite annotations, poor interoperability across studies, and the absence of integrated resources placing microbiome-derived metabolites within their functional, microbial, physiological, and clinical context. Here we present HuMMANet (Human Microbiome Metabolome Annotation Network), a harmonized resource integrating 46 paired gut microbiome metabolome studies (59 study-units; 14,405 samples; 13 disease categories plus a healthy/control reference category) with a scalable metabolite-harmonization framework. HuMMANet resolves heterogeneous annotations through a multi-stage workflow spanning RefMet, HMDB, PubChem, Metabolomics Workbench, SMPDB, MiMeDB 2.0, GNPS/microbeMASST, DrugBank, and DrugCentral, yielding a reference atlas of 54,914 unique metabolites, annotated with standardized chemical identifiers, biochemical pathways, microbial producer associations, physiological distributions, disease links, and structural relationships to approved therapeutics, a unified reference framework for microbiome metabolome research. Applying HuMMANet to a multi-cohort integration of adult serum and fecal metabolomes, we identified 519 serum and 322 fecal metabolites reproducibly associated with gut microbial community composition (PERMANOVA, P < 0.05 in at least 50% of studies in which detected), enriched for specific biomolecular classes and pathways. Cross-referencing these against Health Associated Core Keystone (HACK) taxa revealed 58 serum and 25 fecal metabolites (HACK positive) whose taxon-level associations tracked positively with the taxon specific HACK indices. These reproducible metabolomic signatures of microbiome health included indole3propionic acid, a gut barrier-protective microbial tryptophan metabolite, and 3phenylpropionate. Drug similarity annotation within HuMMANet linked 16 of this serum and 13 fecal HACK positive metabolites to therapeutics used in neurological, inflammatory, and vascular disease. Conversely, 38 serum and 65 fecal metabolites, including imidazole propionate and long-chain acylcarnitines such as ACar 18:0, showed HACK negative signatures previously associated with dysbiosis-linked disease. GNPS/microbeMASST and MiMeDB 2.0 annotations further traced subsets of these metabolites to putative bacterial producers. HuMMANet thus provides a standardized framework for reproducible microbiome metabolome integration, enabling cross study discovery and translational prioritization of conserved microbiome derived metabolic signatures across human populations and disease states.

bioinformatics↗

bioRxiv2026

MifRix: An Integrated microbiome framework for predicting generic and disease-specific risks investigating inter-disease diagnostic cross-talks and intra-disease signature variability

The human gut microbiome is increasingly recognized as a diagnostic indicator across diverse diseases, yet unified frameworks integrating taxonomic and functional features for multi-disease risk assessment, while capturing disease-specific variability in microbiome-alteration signatures, remain limited. Here we present MifRix (Microbiome-inferred Risk-scores with explainability), a two-step ensemble machine-learning framework integrating microbial composition with composition-derived functional signatures to predict generic and disease-specific risk scores across 10 major diseases, coupled with profiling of risk-explainable microbiome features. MifRix was trained using 38,054 gut microbiomes spanning 150 cohorts and 48 nationalities, leveraging taxa abundance and taxa-inferred functional profiles derived from 57,743 functional features mapped across 4,814 species-level taxa. On unseen validation datasets (4,649 microbiomes, 32 cohorts), MifRix outperformed established microbiome health metrics, with disease-specific risk scores achieving strong discrimination (AUC: 0.92-0.99). The explainability module revealed shared microbial signatures across disease pairs, organizing all ten diseases along a continuous gastrointestinal-to-neurological risk gradient, driven not by whole-disease microbiome alteration signatures but by specific, reproducible sub-signatures within each disease. Applied across independent cohorts, MifRix scores further identified population subgroups and individuals at elevated risk for related diseases, flagged precursor/pre-disease states, and tracked therapy-associated response, establishing an interpretable framework for microbiome-based precision diagnostics and disease-risk stratification.

microbiology↗

bioRxiv2026

Community needs for FAIR pathogen data

BackgroundDatasets related to infectious diseases are essential for public health decision-making, yet their reuse remains limited by persistent barriers to data sharing and integration. Achieving data that are Findable, Accessible, Interoperable, and Reusable (FAIR) is widely recognized as essential for accelerating scientific discovery and enabling coordinated responses to emerging threats, but the needs of the global pathogen data community have not been systematically characterized. AimThis study, conducted by the Pathogen Data Network (PDN), aims to identify infrastructural and educational priorities among stakeholders working with infectious disease-related data in order to guide community-responsive support for data sharing and interoperability. MethodsA cross-sectional stakeholder survey was disseminated to a well-defined expert population within PDN networks and via open professional channels. A total of 136 responses from researchers, healthcare professionals, bioinformaticians, and educators were analyzed descriptively to identify prioritized barriers, training needs, and preferred support mechanisms. ResultsRespondents consistently identified structural constraints as the primary impediments to effective data use, including limited funding (74%), data-aggregation challenges (68%), and a shortage of skilled personnel (52%). Respondents identified bioinformatics for infectious disease research (68%) as the highest priority for training, followed by guidance on using the integrated pathogen data and tools portal provided by the PDN, the Pathogens Portal (51%). The Pathogens Portal was also ranked as the most essential PDN resource (72%). Preferred training formats included virtual short courses (68%) and webinars (66%). Notably, while researchers emphasized technical subjects like machine learning, educators prioritized foundational case studies. ConclusionThese findings provide an evidence-based diagnostic of community needs and suggest that barriers to FAIR pathogen data are predominantly systemic rather than purely technological. The survey framework and openly available dataset offer a reusable template for assessing needs in other communities and regions. By aligning training, infrastructure development, and outreach with empirically identified priorities, organizations supporting infectious disease research can strengthen the interoperability and reuse of data and establish a benchmark for future community-driven improvements.

scientific communication and education↗

bioRxiv2026

DHHC7 palmitoylates KRAS4A and promotes mutant KRAS-driven pancreatic cancers

KRAS mutations underlie many human cancers. While inhibitors such as Sotorasib and Adagrasib targeting KRAS mutants have shown promise, additional strategies are required to address the broader spectrum of KRAS-driven cancers, particularly those displaying drug resistance. Thus, there is a need to better understand KRAS signaling and develop new therapeutic strategies. Here we show that KRAS4A is palmitoylated on Cys180 by a palmitoyl transferase, DHHC7 (gene name ZDHHC7). Palmitoylation promotes KRAS4A plasma membrane localization, and more importantly, nanoclustering. This in turn promotes the activation of ARAF and RAF1, but not BRAF. DHHC7 and KRAS4A Cys180 palmitoylation are important for the normal and anchorage independent growth of pancreatic cancer cell lines. Depletion of ZDHHC7 dramatically inhibits pancreatic tumor growth in mouse xenograft models. These studies provide new understandings about how palmitoylation regulates KRAS4A activity and suggest DHHC7 as a promising new target for KRAS mutant cancers.

cancer biology↗

bioRxiv2025

Cerebral blood flow in elastin haploinsufficient and 3xTg-AD mice

Artery structural properties and Alzheimers disease (AD) pathology are individually associated with impaired cerebrovascular function; however, the interaction of these factors is unclear. Furthermore, while elastin haploinsufficient (Eln+/-) mice are known to have impaired cerebrovascular function, sex differences for this effect have not been previously studied. To answer these questions, we crossed middle-aged and old Eln+/-mice with 3xTg-AD mice. We measured cerebral blood flow (CBF) using arterial spin labeling MRI at rest and during hypercapnia to calculate cerebrovascular reactivity (CVR). We also assessed neuroinflammation by microglia and astrocyte cell counts. We found that Eln+/- mice had lower resting blood flow rate in the cerebral cortex compared with Eln+/+ mice, but Eln+/- mice had an intact hypercapnic response, resulting in better CVR compared with Eln+/+ in hippocampus. Sex did not impact resting blood flow or CVR. 3xTg-AD mice had a lower resting CBF than non-AD mice, and there was an interaction between Eln genotype and AD mutations on CVR, such that Eln+/-x 3xTg-AD mice had the poorest hippocampal CVR of all groups. Glia cell counts were highly dependent on brain region, with Eln+/- having more microglia but fewer astrocytes, while 3xTg-AD having higher both microglia and astrocytes. While sex also impacted glial cell counts, we found no interactions between sex and Eln genotype. Our results demonstrate that elastin haploinsufficiency and AD mutations individually result in lower resting CBF, and the combination of these leads to impaired CVR. NEW & NOTEWORTHYThe findings of this study demonstrate that elastin haploinsufficiency leads to lower resting cerebral blood flow, but also greater cerebrovascular reactivity. However, elastin haploinsufficiency interacts with Alzheimers disease mutations to impair cerebrovascular reactivity. These results suggest that multiple insults, such as changes to the extracellular matrix combined with genetic risk factors, are needed to impact cerebrovascular reactivity.

physiology↗

bioRxiv2025

A new typing scheme demonstrates high discriminatory power for Treponema pallidum subspecies

The global resurgence of treponematoses, particularly syphilis, poses a growing public health challenge. Despite recent advances in sequencing technologies, obtaining complete Treponema pallidum genome sequences for epidemiological studies remains time-consuming and challenging due to the difficulty related to procuring clinical samples with sufficient treponemal burden to fulfil the sequencing requirements. There is an urgent need for rapid, cost-effective and accessible typing methods suitable for laboratories with Sanger sequencing resources. Based on the analysis of 121 T. pallidum genomes from geographically diverse regions, we selected seven highly variable genes to form the basis of this new typing system. These seven genes show high discrimination capacity, identifying many allelic profiles among T. pallidum isolates. Importantly, the scheme employs a single-step PCR protocol for the amplification and sequencing of all seven targets enabling straightforward implementation in standard laboratory settings. The MLST was validated using a diverse set of T. pallidum clinical samples from across the globe. A significant proportion of the tested samples showed macrolide resistance, emphasizing the need for epidemiological surveillance. Utilizing this new tool, we have analyzed the genetic variation within and between populations of T. pallidum, considering the geographical origin of the samples. Population structure analysis revealed distinct genetic clusters, underlining complex transmission dynamics of T. pallidum, shaped by local epidemiological factors. The MLST scheme is publicly accessible through the PubMLST database, encouraging widespread adoption in standard laboratories due to this database being user-friendly, intuitive, and fast to implement. The novel MLST scheme offers a promising tool to advance the study of the molecular epidemiology of T. pallidum, facilitate tracking transmission, and establish a global surveillance network with the overall goal of strengthening public health interventions for syphilis control.

microbiology↗

bioRxiv2024

Standardising a microbiome pipeline for body fluid identification from complex crime scene stains

BackgroundRecent advances in next-generation sequencing have opened up new possibilities for utilizing the human microbiome in various fields, including forensics. Researchers have capitalized on the site-specific microbial communities found in different parts of the body to identify body fluids from biological evidence. Despite promising results, microbiome-based methods have not yet been fully integrated into forensic practice due to the lack of standardized protocols and systematic testing of methods on forensically relevant samples. Our study addresses critical decisions in establishing these protocols, focusing on bioinformatics choices and the use of machine learning to present microbiome results in court for forensically relevant and challenging samples. ResultsWe propose using Operational Taxonomic Units (OTUs) for read data processing and creating heterogeneous training datasets for training a random forest classifier. Our classifier incorporates six forensically relevant classes: saliva, semen, hand skin, penile skin, urine, and vaginal/menstrual fluid. Across these classes, our classifier achieved a high weighted average F1 score of 0.89. Systematic testing on mixed-source samples and underwear revealed reliable detection of at least one component of the mixture and the identification of vaginal fluid from underwear substrates. Additionally, when investigating the sexually shared microbiome (sexome) of heterosexual couples, our classifier shows promising results for the inference of sexual activity. ConclusionIn our study, we recommend the use of a novel random forest classifier trained on a heterogenous dataset for obtaining predictions from samples mimicking forensic evidence. We also highlight the potential of the sexome for assessing the nature of sexual activities in forensic investigations, while delineating areas that warrant further research. Furthermore, we underscore key considerations when presenting machine learning results for classifying mixed-source samples.

bioinformatics↗

bioRxiv2024

Structural Basis for the Activation of Proteinase-Activated Receptors (PARs) by Endogenous Ligands

The proteinase-activated receptor (PAR) subfamily of G protein-coupled receptors (GPCRs) include four members, PAR1-PAR4, that play critical roles in hemostasis, thrombosis, embryonic development, wound healing, inflammation, and cancer progression. The PARs share a unique activation mechanism driven by proteinase cleavage at a specific site within the extracellular amino-terminus, exposing a tethered ligand that self-activates the receptor. Subsequent activation allows PAR family members to initiate complex intracellular signaling networks via traditional G protein-mediated pathways and beta-arrestin signaling and, in this way, the PARs link extracellular protease signaling molecules to cellular functions. Despite a primary reliance on biochemical studies for understanding tethered ligand recognition, direct structural visualization of these ligand-receptor complexes has been elusive. Here, we present structural snapshots of activated PAR1 and PAR2 bound to their endogenous tethered ligands, revealing, for the first time, shallow and constricted orthosteric binding pockets and highlighting critical residues involved in ligand recognition and receptor activation. Surprisingly, comparisons with antagonist-bound structures show minimal conformational changes in the TM6 helix, a typical signature of GPCR activation, with large movements of TM7 observed upon activation. These insights lead to the identification of a common mechanism for PAR1 and PAR2 activation and provide a structural template for designing novel antagonists targeting the orthosteric binding site, potentially opening new avenues for therapeutic interventions.

biochemistry↗

bioRxiv2024

Lysophosphatidylcholine acyltransferase 1 suppresses nanoclustering and function of KRAS

Small GTPase KRAS mutated at hotspots, such as G12, G13 and Q61, are major drivers of cancer and display allele-specific oncogenic properties, which are not well understood. KRAS mutants require precise spatiotemporal distribution to the proteolipid nanoclusters on the plasma membrane (PM) for efficient signaling. We recently reported allele-specific lipid sensing of KRAS mutants. KRASG12D, KRASG12V and KRASQ61H favor the unsaturated phosphatidylserine (PS), while KRASG12C and KRASG13D gain enrichment of the saturated PS, cholesterol and/or phosphoinositol 4,5-bisphosphate (PIP2). We, here, examined how the allele-specific lipid sensing of KRAS mutants contributes to their allele-specific signaling and activities. We now show that the stable expression of lysophosphatidylcholine acyltransferase 1 (LPCAT1) elevates the saturated phospholipids and reduces the mixed-chain lipids, especially PS species. Our super-resolution electron microscopy (EM)-spatial analysis revealed that the LPCAT1 expression perturbs the PM nanoclustering of KRASG12D, without affecting that of KRASG12C. LPCAT1 suppresses the KRAS-dependent mitogen-activated protein kinases (MAPKs) signaling and the MAPK-regulated proliferation and colony formation of the KRASG12D-expressing human pancreatic PANC1 cells, while promoting those of the KRASG12C-expressing MiaPaCa-2 cells. Mouse embryonic fibroblasts (MEF) transformed with KRASG12C contain more saturated lipids than those expressing KRASG12D. Concordantly, patient tumor genomics analysis illustrated that expression of LPCAT1 and KRAS mutants negatively correlate in pancreatic adenocarcinoma with KRASG12D as a dominant driver, but loses correlation in lung adenocarcinoma with KRASG12C as a major driver. Thus, the allele-specific lipid sensing of KRAS mutants contributes to their pathological activities.

cancer biology↗

bioRxiv2024

Deletion of Robo4 worsens neuroinflammation and motor coordination in a mouse model of Alzheimer's disease

Declines in vascular integrity are potential contributors to Alzheimers disease (AD) as these result in increased blood-brain barrier permeability and, as a consequence, accelerate neuroinflammation and cognitive impairment. Roundabout guidance receptor 4 (Robo4) is primarily expressed in endothelial cells and stabilizes the vasculature, and thus, has the potential to protect the brain in AD. To study the effect of Robo4 on neuroinflammation and cognitive function in the context of AD, we compared Robo4 knockout and wildtype mice crossed with mice with and without AD mutations (APP/tau). We found that the knockout of Robo4 led to greater astrocyte activation, as demonstrated by GFAP content, but this was dependent on the brain region studied. The knockout of Robo4 also led to greater activated microglia, as assessed by Iba1 content, but only in the presence of AD-related mutations. We found that AD mutations, but not Robo4, were associated with cognitive dysfunction measured by a nest-building test. In contrast, Robo4 deletion, but not AD mutations, was associated with impaired motor coordination. Lastly, Robo4 deletion was associated with greater arterial stiffness, but this trend did not reach statistical significance. In summary, these results demonstrate that Robo4 impacts neuroinflammation, motor coordination, and arterial stiffness, however, the impact on neuroinflammation is dependent on the presence/absence of AD-related mutations and the brain region examined.

physiology↗

bioRxiv2024

Inhibition of NFAT promotes loss of tissue resident uterine natural killer cells and attendant pregnancy complications in humans

Uterine natural killer cells (uNKs) are a tissue resident lymphocyte population that are critical for pregnancy success. Although mouse models have demonstrated that NK deficiency results in abnormal placentation and poor pregnancy outcomes, the generalizability of this knowledge to humans remains unclear. Here we identify uterus transplant (UTx) recipients as a human population with reduced uNK cells and altered pregnancy phenotypes. We show that the NK reduction in UTx correlates with impaired transcriptional programming of NK tissue residency arising from the inhibition of NFAT-mediated signaling. Our observations suggest that NFAT-dependent genes modulate multiple molecular tissue residency programs in uNKs. These include early residency programs involving AP-1-family transcription factors and TGF-{beta}-mediated upregulation of surface integrins. Collectively, our data identify a previously undescribed role for NFAT in uterine NK tissue residency and provide novel mechanistic insights into the biologic basis of pregnancy complications due to alteration of tissue resident NK subsets in humans. One Sentence SummaryRole of NFAT in uterine NK cell tissue residency

immunology↗

bioRxiv2024

A Mycobacterium tuberculosis secreted virulence factor disrupts host snRNP biogenesis

We earlier reported that Mycobacterium tuberculosis (Mtb) alters host RNA splicing to help its intracellular growth. Here, we report an unprecedented mechanism whereby a secreted virulence factor from Mtb interferes with the biogenesis of key spliceosomal components, causing an altered RNA splicing pattern. A high-throughput yeast-2-hybrid screen identified several Mtb-secreted proteins that can interact with the host RNA splicing factors (SFs). Through custom-designed in-cell assays, we show that one of those proteins, Rv1435c, targets specific exon-skipping events to alter RNA splicing. We show that Rv14345c or host splicing regulator 1 (hsr1) facilitates direct interaction between Mtb phagosomes and U5 snRNA and SNRPF, key components of the snRNPs. Genetic deletion of hsr1 reverses the specific exon-skipping events caused by WT Mtb infection. The{Delta} hsr1 strain shows compromised growth during ex vivo infection in the macrophages and in vivo infection in the mice. Tissue sections from the WT Mtb or{Delta} hsr1-infected mice showed significant hsr1-dependent SNRPF staining, a phenomenon also noted in the human intestinal tuberculosis (ITB) biopsies. We infer that hsr1 is a virulence factor which alters RNA splicing by interacting with U5snRNA and SNRPF. The splicing regulators from the host and pathogen are novel targets for anti-tuberculosis therapy.

molecular biology↗

bioRxiv2023

Identification of an H-Ras nanocluster disrupting peptide

The Ras-MAPK pathway is critical to regulate cell proliferation and differentiation. Its dysregulation is implicated in the onset and progression of numerous types of cancers. To be active, Ras proteins are membrane anchored and organized into nanoclusters, which realize high-fidelity signal transmission across the plasma membrane. Nanoclusters therefore represent potential drug targets. However, targetable protein components of signalling nanoclusters are poorly established. We previously proposed that the nanocluster scaffold galectin-1 (Gal1) enhances H-Ras nanoclustering by stabilizing stacked dimers of H-Ras and Raf via a direct interaction of dimeric Gal1 with the Ras binding domain (RBD) in particular of B-Raf. Here, we provide further supportive evidence for this model. We establish that the B-Raf preference emerges from divergent regions of the Raf RBDs that were proposed to interact with Gal1. We then identify the L5UR peptide, which disrupts this interaction by binding with low micromolar affinity to the B-Raf-RBD. Its 23-mer core fragment is thus sufficient to interfere with Gal1-enhanced H-Ras nanocluster, reduce MAPK-output and cell viability in HRAS-mutant cancer cell lines. Our data therefore suggest that the interface between Gal1 and the RBD of B-Raf can be targeted to disrupt Gal1-enhanced H-Ras nanoclustering. Collectively, our results support that Raf-proteins are integral components of active Ras nanoclusters.

biochemistry↗

bioRxiv2023

RAS G-domains fine-tune the sorting of phosphatidylserine acyl chains in the plasma membrane

Mutant RAS are major contributors to cancer and signal from nanoclusters on the plasma membrane (PM), via isoform-specific membrane anchors. However, the same RAS isoform bound to different guanine nucleotides are segregated on the PM. Paradoxically, various segregated RAS nanoclusters all enrich a type of anionic phospholipid, phosphatidylserine (PS). These findings suggest intricate participation of RAS G-domains in their PM distribution, which have not been explored. We now show that wild-types, oncogenic G12V mutants and membrane anchors of isoforms HRAS, KRAS4A and KRAS4B sort distinct PS species. Mechanistically, shifting orientation states of KRAS4B G-domain exposes residues, such as Arg 73, Arg 102 and Arg 135, to the PM, and contributes to PS acyl chain sorting. Oncogenic mutations may shift orientation states of G-domains. We show that G12V, G12D, G12C, G13D and Q61H mutants of KRAS4B sort distinct PS species. Thus, RAS G-domains fine-tune their lateral distribution on the PM.

molecular biology↗

bioRxiv2023

CaV3.1 Channels Facilitate Calcium Wave Generation and Myogenic Tone Development in Mouse Mesenteric Arteries.

BackgroundThe myogenic response is the mechanism whereby intraluminal pressure elicits arterial constriction pursuant to the maintenance of tissue perfusion. Smooth muscle [Ca2+] is a key determinant of constriction, a process intimately tied to L-type (CaV1.2) Ca2+ channels. While important, other Ca2+ channels, in particular T-type, are expressed and could contribute to pressure regulation within defined voltage ranges. This study examined the role of one T-type Ca2+ channel using mesenteric arteries from C57BL/6 wild type and CaV3.1-/- mice. MethodsPatch-clamp electrophysiology, pressure myography, non-invasive blood pressure measurements and rapid Ca2+ imaging were employed to define the CaV3.1-/- phenotype relative to C57BL/6. Proximity ligation assay tested the closeness of CaV3.1 channels to inositol triphosphate receptors (IP3R). Nifedipine (0.3 M) and 2-APB (50 M) were used to block L-type Ca2+ channels and IP3Rs, respectively. ResultsInitial experiments confirmed the absence of CaV3.1 expression and whole-cell current in global deletion mice, a change that coincided with a reduction in systemic blood pressure. Mesenteric arteries from CaV3.1-/- mice produced less myogenic tone than C57BL/6, particularly at lower pressures (20-60 mmHg) where membrane potential is more hyperpolarized. This reduction in myogenic tone correlated with diminished Ca2+ wave generation in the CaV3.1-/- mice. These asynchronous events are dependent upon Ca2+ release from the sarcoplasmic reticulum which is insensitive to L-type Ca2+ channel blockade. A close physical association (<40 nm) between IP3R1 and CaV3.1 was confirmed by proximity ligation assay; blockade of IP3R in nifedipine-treated C57BL/6 arteries rendered a CaV3.1-/- contractile phenotype. ConclusionFindings indicate that Ca2+ influx through CaV3.1 channels contributes to myogenic tone development at hyperpolarized voltages by triggering a Ca2+-induced Ca2+ release mechanism tied to the sarcoplasmic reticulum. This study helps establish CaV3.1 as a potential therapeutic target in the control of blood pressure. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=160 SRC="FIGDIR/small/528095v1_ufig1.gif" ALT="Figure 1"> View larger version (44K): org.highwire.dtl.DTLVardef@1b3fe9forg.highwire.dtl.DTLVardef@11954borg.highwire.dtl.DTLVardef@607417org.highwire.dtl.DTLVardef@65964c_HPS_FORMAT_FIGEXP M_FIG C_FIG

systems biology↗

bioRxiv2023

Inferring patterns of recombination and divergence with ancient and modern treponemal genomes

BackgroundTreponemal diseases pose significant global health risks, presenting severe challenges to public health due to their serious health impacts if left untreated. Despite numerous genomic studies on Treponema pallidum and the known possible biases introduced by the choice of the reference genome used for mapping, few investigations have addressed how these biases affect phylogenetic and evolutionary analysis of these bacteria. In this study, we assessed the impact of selecting an appropriate genomic reference on phylogenetic and evolutionary analyses of T. pallidum. ResultsWe designed a multiple-reference-based (MRB) mapping strategy using four different reference genomes and compared it to traditional single-reference mapping. To conduct this comparison, we created a genomic dataset comprising 77 modern and ancient genomes from the three subspecies of T. pallidum, including a newly sequenced 17th-century genome (35X coverage) of a syphilis-causing strain (designated as W86). Our findings show that recombination detection was consistent across different references, but the choice of reference significantly affected ancient genome reconstruction and phylogenetic inferences. The high-coverage W86 genome obtained here also provided a new calibration point for Bayesian molecular clock dating, improving the reconstruction of the evolutionary history of treponemal diseases. Additionally, we identified novel recombination events, positive selection targets, and refined dating estimates for key events in the species history. ConclusionsThis study highlights the importance of considering methodological implications and reference genome bias in High-Throughput Sequencing-based whole-genome analysis of T. pallidum, especially of ancient or low-coverage samples, contributing to a deeper understanding of this pathogen and its subspecies.

microbiology↗