Biology subjects

Sinha, A.

Publications and source records attributed to Sinha, A..

At least 73 records · Page 4Linked to original sources

bioRxiv2023

The NeuroML ecosystem for standardized multi-scale modeling in neuroscience

Data-driven models of neurons and circuits are important for understanding how the properties of membrane conductances, synapses, dendrites and the anatomical connectivity between neurons generate the complex dynamical behaviors of brain circuits in health and disease. However, the inherent complexity of these biological processes make the construction and reuse of biologically detailed models challenging. A wide range of tools have been developed to aid their construction and simulation, but differences in design and internal representation act as technical barriers to those who wish to use data-driven models in their research workflows. NeuroML, a model description language for computational neuroscience, was developed to address this fragmentation in modeling tools. Since its inception, NeuroML has evolved into a mature community standard that encompasses a wide range of model types and approaches in computational neuroscience. It has enabled the development of a large ecosystem of interoperable open source software tools for the creation, visualization, validation and simulation of data-driven models. Here, we describe how the NeuroML ecosystem can be incorporated into research workflows to simplify the construction, testing and analysis of standardized models of neural systems, and supports the FAIR (Findability, Accessibility, Interoperability, and Reusability) principles, thus promoting open, transparent and reproducible science.

neuroscience↗

bioRxiv2023

Evolution of tandem repeats in putative CSP to enhance its function: A recent and exclusive event in Plasmodium vivax in India

The molecular hitchhiking model proposes that linked non-coding regions also undergo fixation, while fixing a beneficial allele in a population. This concept can be applied to identify loci with functional and evolutionary significance. Putative circumsporozoite protein (CSP) in Plasmodium vivax (PvpuCSP) identified following the molecular hitchhiking model, holds evolutionary significance. We investigated the extent of genetic polymorphism in PvpuCSP and the role of natural selection which shapes the genetic composition and maintains the diversity in P. vivax isolates from India. Sequencing the putative CSP of P. vivax (PvpuCSP) in 71 isolates revealed a well-conserved N- and C-terminal, constituting around 80% of the gene. PCR amplification and sequencing validated extensive diversity in the repeat region, ranging from 1.8 to 2.2 kb towards the C-terminal, identifying 37 different alleles from 71 samples. The recent and exclusive accumulation of repeats in puCSP within P. vivax highlights its highly variable length polymorphism, making it a potential marker for estimating diversity and infection complexity. Episodic diversifying selection in the PvpuCSP repeat region, evidenced by statistically significant p-values and likelihood ratios, enhances amino acid diversity at various phylogenetic levels, facilitating adaptation for accommodating different substrates for degradation.

evolutionary biology↗

bioRxiv2023

ACSL4 activity drives TNBC metastasis by positively regulating Histone H3 Acetylation mediated SNAIL expression

Triple-Negative Breast Cancer (TNBC) has profound unmet medical need globally for its devastating clinical outcome associated with rapid metastasis and lack of targeted therapies. Recently, lipid metabolic reprogramming has emerged as a major driver of breast cancer metastasis. Here, we unveil a strong association between the heightened expression of fatty acid metabolic enzyme, acyl-CoA synthetase 4 (ACSL4) and TNBC, which is primarily attributed by the selective absence of progesterone receptor (PR). Loss of ACSL4 function, either through genetic ablation or pharmacological inhibition significantly reduces metastatic potential of TNBC. Global transcriptome analysis reveals that ACSL4 activity markedly influences the gene expression pattern associated with TNBC migration. Mechanistically, ACSL4 alters fatty acid oxidation (FAO) and cellular acetyl-CoA levels, leading to the hyper-acetylation of particularly H3K27Ac and H3K9Ac marks resulting in overexpression of SNAIL during the course of TNBC metastatic spread to lymph node and lungs. Further, human TNBC metastasis exhibits positive correlation between ACSL4 and SNAIL expression. Altogether, our findings provide new molecular insights regarding the intricate interplay between metabolic alterations and epigenetic modifications, intertwined to orchestrate TNBC metastasis and posit a rational understanding for the development of ACSL4 inhibitors as a targeted therapy against TNBC.

cancer biology↗

bioRxiv2023

Rational development of a small-molecule activator of CK1γ2 that decreases C99 and beta-amyloid levels

Alzheimers disease (AD) is a debilitating neurodegenerative disorder characterized by the accumulation of beta-amyloid (A{beta}), C99, and Tau in vulnerable areas of the brain. Despite extensive research, current strategies to lower A{beta} levels have shown limited efficacy in slowing the cognitive decline associated with AD. Recent findings suggest that C99 may also play a crucial role in the pathogenesis of AD. Our laboratory has discovered that CK1{gamma}2 phosphorylates Presenilin 1 at the {gamma}-secretase complex, leading to decreased C99 and A{beta} levels. Thus, CK1{gamma}2 activation appears as a promising therapeutic target to lower both C99 and A{beta} levels. In this study, we demonstrate that CK1{gamma}2 is inhibited by intramolecular autophosphorylation and describe a high-throughput screen designed to identify inhibitors of CK1{gamma}2 autophosphorylation. We hypothesize that these inhibitors could lead to CK1{gamma}2 activation and increased PS1-Ser367 phosphorylation, ultimately reducing C99 and A{beta} levels. Using cultured cells, we investigated the impact of these compounds on C99 and A{beta} concentrations and confirmed that CK1{gamma}2 activation effectively reduces their levels. Our results provide proof of concept that CK1{gamma}2 is an attractive therapeutic target for AD. Future studies should focus on the identification of specific compounds that can inhibit CK1{gamma}2 autophosphorylation and evaluate their efficacy in preclinical models of AD. These studies will pave the way for the development of novel therapeutics for the treatment of AD.

pharmacology and toxicology↗

bioRxiv2023

PERK arm of UPR selectively regulates ferroptosis in colon cancer cells by modulating the expression of SLC7A11 (System Xc-)

Ferroptosis, a genetically and biochemically distinct form of programmed cell death, is characterised by an iron-dependent accumulation of lipid peroxides. Therapy-resistant tumor cells display vulnerability toward ferroptosis. Endoplasmic Reticulum (ER) stress and Unfolded Protein Response (UPR) play a critical role in cancer cells to become therapy resistant. Tweaking the balance of UPR to make cancer cells susceptible to ferroptotic cell death could be an attractive therapeutic strategy. To decipher the emerging contribution of ER-stress in the ferroptotic process, we observe that ferroptosis inducer RSL3 promotes UPR (PERK, ATF6, and IRE1), along with overexpression of cystine-glutamate transporter SLC7A11 (System Xc-). Exploring the role of a particular UPR arm in modulating SLC7A11 expression and subsequent ferroptosis, we notice that PERK is selectively critical in inducing ferroptosis in colorectal carcinoma. PERK inhibition reduces ATF4 expression and recruitment to the promoter of SLC7A11 and results in its downregulation. Loss of PERK function not only primes cancer cells for increased lipid peroxidation but also limits in vivo colorectal tumor growth, demonstrating active signs of ferroptotic cell death in situ. Further, by performing TCGA data mining and using colorectal cancer patient samples, we demonstrate that the expression of PERK and SLC7A11 is positively correlated. Overall, our experimental data indicate that PERK is a negative regulator of ferroptosis and loss of PERK function sensitizes colorectal cancer cells to ferroptosis. Therefore, small molecule PERK inhibitors hold huge promise as novel therapeutics and their potential can be harnessed against the apoptosis-resistant condition.

cancer biology↗

bioRxiv2023

CD38 promotes hematopoietic stem cell dormancy via c-Fos

A subpopulation of deeply quiescent, so-called dormant hematopoietic stem cells (dHSCs) resides at the top of the hematopoietic hierarchy and serves as a reserve pool for HSCs possessing the greatest long-term blood repopulation capacity. The state of dormancy protects the HSC pool from exhaustion throughout life, however excessive dormancy may block an efficient response to hematological stresses. The mechanisms of HSC dormancy remain elusive, mainly due to the absence of surface markers that allow dHSC prompt isolation. Here, we identify CD38 as a novel surface marker for murine dHSCs that is broadly applicable. Moreover, we demonstrate that cyclic adenosine diphosphate ribose (cADPR), the product of CD38 cyclase activity, regulates the expression of the transcription factor c-Fos by increasing cytoplasmic Ca2+ concentration. Strikingly, we uncover that c-Fos drives HSCs dormancy through the induction of the cell cycle inhibitor p57Kip2. Moreover, we found that CD38 ecto-enzymatic activity at the neighboring CD38-positive cells can promote human HSC quiescence. Together, CD38/cADPR/Ca2+/cFos/p57Kip2 axis maintains HSC dormancy. Pharmacological manipulations of this pathway can provide new strategies to expand dHSCs for transplantation or to activate them during hematological stresses.

cell biology↗

bioRxiv2023

Metagenomics of culture isolates and insect tissue illuminate the evolution of Wolbachia, Rickettsia and Bartonella symbionts in Ctenocephalides spp. fleas

While fleas are often perceived simply as a biting nuisance and cause of allergic dermatitis, they represent important disease vectors worldwide, especially for bacterial zoonoses such as plague (transmitted by rodent fleas) and some of the rickettsioses and bartonelloses. The cosmopolitan cat (Ctenocephalides felis) and dog (C. canis) fleas, as well as C. orientis (restricted to tropical and subtropical Asia), breed in human dwellings and are vectors of cat-scratch fever (caused by Bartonella spp.) and Rickettsia spp. of the so-called "transitional group". The latter includes R. felis (agent of flea-borne spotted fever) and R. asembonensis, an emerging pathogen. The relatively depauperate flea microbiome can also contain arthropod-specific endosymbionts, including a diverse range of Wolbachia strains. Here, we present circularized genome assemblies for two C. orientis-associated pathogens (Bartonella clarridgeiae and R. asembonensis) from Malaysia, a novel Wolbachia strain (wCori), and the C. orientis mitochondrion; all obtained by direct metagenomic sequencing of flea tissues. Moreover, we isolated two Wolbachia strains from Malaysian C. felis into tick cell culture and recovered circularized genome assemblies for both, one of which (wCfeF) is newly sequenced. We demonstrate that the three Wolbachia strains are representatives of different major clades ("supergroups"), two of which appear to be flea-specific. These Wolbachia genomes exhibit unique combinations of features associated with reproductive parasitism or mutualism, including prophage WO, cytoplasmic incompatibility factors, and the biotin operon of obligate intracellular microbes. The first circularized assembly for R. asembonensis includes a plasmid with a markedly different structure and gene content compared to the published plasmid; moreover, this novel plasmid was also detected in cat flea metagenomes from the US. Analysis of loci under positive selection in the transitional group revealed genes involved in host-pathogen interactions that may facilitate host switching. Finally, the first B. clarridgeiae genome from Asia exhibited largescale genome stability compared to isolates from other continents, except for SNPs in regions predicted to mediate interactions with the vertebrate host. These findings highlight the paucity of data on the genomic diversity of Ctenocephalides-associated bacteria and raise questions regarding how interactions between members of the flea microbiome might influence vector competence. Impact statementWhile fleas are vectors for several globally-distributed bacterial zoonoses, including plague, murine typhus, cat-scratch fever, and flea-borne spotted fever, data on the genomic diversity of the etiological agents of these diseases and other members of the flea microbiome are scant. Here, we focused on Ctenocephalides spp. fleas associated with dogs and cats in Malaysia, a region where flea microbiome data are particularly sparse. Using a combination of direct metagenomic sequencing of flea tissues coupled with culture isolation and sequencing of infected cell lines, we obtained the first genomes for a cat-scratch agent (Bartonella clarridgeiae) and an emerging human pathogen (Rickettsia asembonensis) from Asia, as well as genomes for three different Wolbachia endosymbiont strains and the first mitochondrial genome for Ctenocephalides orientis. The pathogen genomes reveal evidence of selection in genes involved in host-pathogen interactions and in the case of R. asembonensis, a novel plasmid genome was obtained. Two of the Wolbachia genomes are entirely novel and exhibit highly unusual combinations of features associated with potential reproductive manipulations and mutualistic roles. Our findings highlight major gaps in our understanding of flea microbiome diversity worldwide, with implications for the emergence of bacterial pathogens transmitted by these ubiquitous ectoparasites associated with human dwellings. Data summarySequencing reads and genome assemblies produced in study have been deposited in NCBI Genbank under BioProject accession PRJNA909216. Accession numbers for symbiont genomes: wCfeF (CP116767), wCfeJ (CP116768), wCori (CP116769), Rickettsia asembonensis str. Perak (CP116496), pRAS02 (CP116495) and Bartonella clarridgeiae str. Perak (CP116497). Ctenocephalides orientis mitochondrial assembly was deposited with the accession number OQ383237. The authors confirm all supporting data, code and protocols have been provided within 86 the article or through supplementary data files.

genomics↗

bioRxiv2023

Genomes of the human filarial parasites Mansonella perstans and Mansonella ozzardi

The filarial parasites Mansonella ozzardi and Mansonella perstans, causative agents of mansonellosis, infect hundreds of millions of people worldwide, yet remain among the most understudied of the human filarial pathogens. M. ozzardi is highly prevalent in Latin American countries and Caribbean Islands, while M. perstans is predominantly found in sub-Saharan Africa as well as in a few areas in South America. In addition to the differences in their geographical distribution, the two parasites are transmitted by different insect vectors, as well as exhibit differences in their responses to commonly used anthelminthic drugs. The lack of genome information has hindered investigations into the biology and evolution of Mansonella parasites and understanding the molecular basis of the clinical differences between species. In the current study, high quality genomes of two independent clinical isolates of M. perstans from Cameroon and two M. ozzardi isolates one from Brazil and one from Venezuela are reported. The genomes are approximately 76 Mb in size, encode about 10,000 genes each, and are largely complete based on BUSCO scores of about 90%, similar to other completed filarial genomes. These sequences represent the first genomes from Mansonella parasites and enabled a comparative genomic analysis of the similarities and differences between Mansonella and other filarial parasites. Horizontal DNA transfers (HDT) from mitochondria (nuMTs) as well as transfers from genomes of endosymbiotic Wolbachia bacteria (nuWTs) to the host nuclear genome were identified and analyzed. Sequence comparisons and phylogenetic analysis of known targets of anti-filarial drugs diethylcarbamazine (DEC), ivermectin and mebendazole revealed that all known target genes were present in both species, except for the DEC target encoded by gon-2 gene, which is fragmented in genome assemblies from both M. ozzardi isolates. These new reference genome sequences will provide a valuable resource for further studies on biology, symbiosis, evolution and drug discovery.

genomics↗

bioRxiv2022

Niche partitioning and coexistence of sympatric macaques in a fragmented habitat of the Upper Brahmaputra Valley, northeastern India

How closely related species co-exist, especially under conditions of resource limitation remains an intriguing problem in ecology. Having to share space and resources, such species are expected to have evolved a variety of mechanisms to reduce competition. In this study, we examined the niche partitioning of three congeneric species, northern pig-tailed macaque Macaca leonina, the rhesus macaque Macaca mulatta, and stump-tailed macaque Macaca arctoides, in a tropical lowland rainforest fragment of less than 21 square kilometer, the Hollongapar Gibbon Sanctuary, northeastern India. We conducted behavioural observations on two troops each of the three species over 23 months from March 2008 to January 2010 resulting in 6,471, 11,062 and 16,723 individual scans of pig-tailed, rhesus and stump-tailed macaques respectively. We compared activity budgets and examined niche partitioning among the macaques along space and food axes. The activity budgets of three macaques shows differences in their feeding and foraging behaviours: the stump-tailed were active throughout the day, pig-tailed macaques were most active during mid-day while the rhesus macaques activity peaked after late afternoon. The rhesus macaque utilised the forest edge and surrounding areas whereas the stump-tailed and pig-tailed macaques use forest interiors. Pig-tailed macaques used significantly higher canopy strata, while stump-tailed macaques were predominantly terrestrial while rhesus macaques used the mid-lower canopy. Dietary overlap was highest between rhesus and pig-tailed macaques and lowest between pig-tailed and stump-tailed macaques. Rhesus macaques showed the broadest dietary niche breadth, whereas pig-tailed and stump-tailed macaques exhibited narrower and more specialised diets. Our results demonstrate that fine-scale partitioning of space and food resources facilitates the long-term coexistence of three congeneric macaques within a small and isolated rainforest fragment. These findings provide important insights into primate persistence and conservation in fragmented tropical landscapes.

ecology↗

bioRxiv2022

Possible Self-awareness in Wild Adelie Penguins Pygoscelis adeliae

This preliminary study, conducted in January-February 2020, investigates the potential presence of self-awareness in a population of wild Adelie penguins on the Dogs Neck Ice Shelf and on Svenner Island in East Antarctica. It is based on the responses and reactions of individual penguins to images, generated in mirrors during three experimental paradigms: a group-behaviour test; a modified mirror test and a hidden-head test. We believe that this set of experiments constitutes possibly the first investigations into the potential presence of self-awareness in any penguin species and is pioneering in conducting a set of cognitive experiments on free-ranging individuals of a nonhuman species in its natural environment, without any prior familiarisation, conditioning or acclimatisation to the experimental paradigms employed. Future studies, integrating the socioecology and cognitive ethology of penguins, may provide insights into whether our experimental paradigms could provide evidence to confirm the presence of self-awareness and even of self-recognition in this species and examine whether the observed social awareness may have evolved due to the social needs of individual penguins to engage in cooperative behaviour with conspecific individuals, while maintaining their independent decision-making capacities, throughout their communal lives.

animal behavior and cognition↗

bioRxiv2022

An RNA modification enzyme directly senses reactive oxygen species for translational regulation in Enterococcus faecalis

Bacteria possess elaborate systems to manage reactive oxygen and nitrogen species (ROS) arising from exposure to the mammalian immune system and environmental stresses. Here we report the discovery of an ROS-sensing RNA-modifying enzyme that regulates translation of stress-response proteins in the gut commensal and opportunistic pathogen Enterococcus faecalis. We analyzed the tRNA epitranscriptome of E. faecalis in response to reactive oxygen species (ROS) or sublethal doses of ROS-inducing antibiotics and identified large decreases in N2-methyladenosine (m2A) in both 23S ribosomal RNA and transfer RNA. This we determined to be due to ROS-mediated inactivation of the Fe-S cluster-containing methyltransferase, RlmN. Genetic knockout of RlmN gave rise to a proteome that mimicked the oxidative stress response, with increased levels of superoxide dismutase and decreased virulence proteins. While tRNA modifications are established to be dynamic for fine-tuning translation, here we report the first instance of a dynamically regulated, environmentally responsive rRNA modification. These studies lead to model in which RlmN serves as a redox-sensitive molecular switch, directly relaying oxidative stress to modulating translation through the rRNA and the tRNA epitranscriptome, revealing a new paradigm for understanding direct regulation of the proteome by RNA modifications.

microbiology↗

bioRxiv2022

Female Friendships: Social Network Analysis as a tool to understand intra-group affiliation in semi-commensal lion-tailed macaques (Macaca silenus)

The lion-tailed macaque is a gregarious, rainforest-adapted species, that has, in certain locations across its natural distribution, recently begun to explore and utilise surrounding human-dominated habitats. In many primate species, exposure to novel human-use food resources and potential provisioning has previously been associated with changes in intra-group social structure, often categorised by increased aggression and, more importantly, increased affiliation, possibly as a means of reconciliation. Here we quantify the changing affiliative relationships among the female members of two groups with varying degrees of habituation, using social network analysis. We examine frequencies of pair-wise affiliation between the female members of each group, ranked within the prevailing linear hierarchy, to measure individual attributes or the local importance of individuals in a network, and group attributes or the global role of all individuals in the whole network. We found that the subordinate individuals in the less habituated group maintained a higher number of connections with group members, an expected outcome, as key affiliative behaviours such as grooming are known to be directed upwards in the hierarchy. This pattern was observed to be inverse in the highly habituated group, with dominant individuals maintaining more connections, suggesting that under the conditions of increased competition for the novel food resource, dominance rank was highly contested. In support of this theory, we also found multiple fluctuations in dominance rank over time for this highly habituated group, with nearly no fluctuations in the less habituated group. This study demonstrates that varied intensities of human presence and dependence on human-use foods have differential effects on the intra-group sociality of lion-tailed macaques.

animal behavior and cognition↗

bioRxiv2022

Gemini calling! First occurrence of successful twinning in wild, endangered lion-tailed macaques Macaca silenus in the Anamalai hills of the Western Ghats, India

We recorded the first occurrence of surviving twins in lion-tailed macaques Macaca silenus from the Anamalai hills of the Western Ghats, India. The Puthuthottam population of liontailed macaques has historically been restricted to a rainforest fragment measuring 92ha, situated adjacent to human settlements, however, direct interactions between macaques and humans have been observed only in the last ten years. The population now visit settlements at a rate of 0.52/ day, and exploits anthropogenic foods. We recorded two sets of twins from the Puthuthottam population within a six-month period. Two previous cases of twins have been observed in this population since 2000, however, in both cases the twins did not survive beyond a few weeks. We followed and opportunistically collected ad libitum behavioural data on both sets of twins for a year between March 2019 to March 2020. Both mothers carrying the twins used the ground substrate extensively, however the mother with the younger set of twins also used the canopy and other precarious substrates such as cable wires. Our report shows that twinning occurs in lion-tailed macaques and twins are able to survive successfully. This report also supports previous evidence that twinning may occur in higher frequencies as a direct response to provisioning, with the mothers behaviour contributing to the successful survival of both twins.

animal behavior and cognition↗

bioRxiv2022

Ranging patterns of the rainforest-adapted lion-tailed macaque Macaca silenus in a human-dominated landscape in the Anamalai hills of the Western Ghats, India

The ranging patterns of five lion-tailed macaque Macaca silenus troops, forming the Puthuthottam sub-population, were studied over a three year period to determine road/habitation visitation rate, home ranges and habitat preference. Each troop visited the road or human habitation at varying rates, with the largest troop visiting most frequently. Home ranges sizes were observed to be highly reduced when compared to wild populations, and also greatly varied across troops, with relatively low overlap given the macaque density in the available area. All five macaque troops showed a preference for human-modified habitats such as roads and human settlements where anthropogenic food was easily available. Our study shows an increasing dependence amongst members of the Puthuthottam troops on anthropogenic foods, which has led to many threats faced by individuals including fatal collisions with vehicular traffic and electrocutions.

ecology↗

bioRxiv2022

Expansion Sequencing of RNA Barcoded Neurons in the Mammalian Brain: Progress and Implications for Molecularly Annotated Connectomics

Mapping and molecularly annotating mammalian neural circuits is challenging due to the inability to uniquely label cells while also resolving subcellular features such as synaptic proteins or fine cellular processes. We argue that an ideal technology for connectomics would have the following characteristics: the capacity for robust distance-independent labeling, synaptic resolution, molecular interrogation, and scalable computational methods. The recent development of high-diversity cellular barcoding with RNA has provided a way to overcome the labeling limitations associated with spectral dyes, however performing all-optical circuit mapping has not been demonstrated because no method exists to image barcodes throughout cells at synaptic-resolution. Here we show ExBarSeq, an integrated method combining in situ sequencing of RNA barcodes, immunostaining, and Expansion Microscopy coupled with an end-to-end software pipeline that automatically extracts barcode identities from large imaging datasets without data processing bottlenecks. As a proof of concept, we applied ExBarSeq to thick tissue sections from mice virally infected with MAPseq viral vectors and demonstrated the extraction of 50 barcoded cells in the visual cortex as well as cell morphologies uncovered via immunostaining. The current work demonstrates high resolution multiplexing of exogenous barcodes and endogenous synaptic proteins and outlines a roadmap for molecularly annotated connectomics at a brain-wide scale.

neuroscience↗

bioRxiv2022

Cohesin is involved in transcriptional repression of stage-specific genes in the human malaria parasite

The most virulent human malaria parasite, Plasmodium falciparum, has a complex life cycle between its human host and mosquito vector. Each stage is driven by a specific transcriptional program, but with a relatively high ratio of genes to specific transcription factors, it is unclear how genes are activated or silenced at specific times. The P. falciparum genome is relatively euchromatic compared to the mammalian genome, except for specific genes that are uniquely heterochromatinized via HP1. There seems to be an association between gene activity and spatial organization; however, the molecular mechanisms behind genome organization are unclear. While P. falciparum lacks key genome-organizing proteins found in metazoans, it does have all core components of the cohesin complex. In other eukaryotes, cohesin is involved in sister chromatid cohesion, transcription, and genome organization. To investigate the role of cohesin in P. falciparum, we combined genome editing, mass spectrometry, chromatin immunoprecipitation and sequencing (ChIP-seq), and RNA sequencing to functionally characterize the cohesin subunit Structural Maintenance of Chromosomes protein 3 (SMC3). SMC3 knockdown in early stages of the intraerythrocytic developmental cycle (IDC) resulted in significant upregulation of a subset of genes involved in erythrocyte egress and invasion, which are normally expressed at later stages. ChIP-seq of SMC3 revealed that over the IDC, enrichment at the promoter regions of these genes inversely correlates with their expression and chromatin accessibility levels. These data suggest that SMC3 binding helps to repress specific genes until their appropriate time of expression, revealing a new mode of stage-specific, HP1-independent gene repression in P. falciparum.

molecular biology↗

bioRxiv2022

A Multifunctional Anchor for Multimodal Expansion Microscopy

In situ imaging of biomolecular location with nanoscale resolution enables mapping of the building blocks of life throughout biological systems in normal and disease states. Expansion microscopy (ExM), by physically enlarging specimens in an isotropic fashion, enables nanoimaging on standard light microscopes. Key to ExM is the equipping of different kinds of molecule, with different kinds of anchoring moiety, so they can all be pulled apart by polymer swelling. Here we present a multifunctional anchor, an acrylate epoxide, that enables multiple kinds of molecules (e.g., proteins and RNAs) to be equipped with anchors in a single experimental step. This reagent simplifies ExM protocols and greatly reduces cost (by 2-10 fold for a typical multiplexed ExM experiment) compared to previous strategies for equipping RNAs with anchors. We show that this unified ExM (uniExM) protocol can be used to preserve and visualize RNA transcripts, proteins in biologically relevant ultrastructure, and sets of RNA transcripts in patient-derived xenograft (PDX) cancer tissues, and can support the visualization of other kinds of biomolecular species as well. Thus, uniExM may find many uses in the simple, multimodal nanoscale analysis of cells and tissues.

bioengineering↗

bioRxiv2022

INFLAMMATION INDUCED DOWNREGULATION OF SUCCINATE DEHYDROGENASE REGULATES ADRENOCORTICAL FUNCTION

The hypothalamus-pituitary-adrenal (HPA) axis is activated in response to inflammation leading to increased production of anti-inflammatory glucocorticoids by the adrenal cortex, thereby representing an endogenous feedback loop. However, severe inflammation reduces the responsiveness of the adrenal gland to adrenocorticotropic hormone (ACTH) although the underlying mechanisms are poorly understood. Here, we show by transcriptomic, proteomic and metabolomic analyses that LPS-induced systemic inflammation triggers profound metabolic changes in steroidogenic adrenocortical cells, including downregulation of the TCA cycle and oxidative phosphorylation. Inflammation disrupts the TCA cycle at the level of succinate dehydrogenase (SDH) leading to succinate accumulation and disturbed steroidogenesis. Mechanistically, IL-1{beta} reduces SDHB expression through upregulation of DNA methyltransferase 1 (DNMT1) and methylation of the SDHB promoter. Consequently, increased succinate levels impair oxidative phosphorylation and ATP synthesis, leading to reduced steroidogenesis. Together, we demonstrate that the IL-1{beta}-DNMT1-SDHB-succinate axis disrupts steroidogenesis. Our findings not only provide a mechanistic explanation for the adrenal dysfunction in severe inflammation but also a potential target for therapeutic intervention.

immunology↗