Biology subjects

Das, S.

Publications and source records attributed to Das, S..

At least 217 records · Page 12Linked to original sources

bioRxiv2023

Spatial and single-nucleus transcriptomic analysis of genetic and sporadic forms of Alzheimer's Disease

The pathogenesis of Alzheimers disease (AD) depends on environmental and heritable factors, with remarkable differences evident between individuals at the molecular level. Here we present a transcriptomic survey of AD using spatial transcriptomics (ST) and single-nucleus RNA-seq in cortical samples from early-stage AD, late-stage AD, and AD in Down Syndrome (AD in DS) donors. Studying AD in DS provides an opportunity to enhance our understanding of the AD transcriptome, potentially bridging the gap between genetic mouse models and sporadic AD. Our analysis revealed spatial and cell-type specific changes in disease, with broad similarities in these changes between sAD and AD in DS. We performed additional ST experiments in a disease timecourse of 5xFAD and wildtype mice to facilitate cross-species comparisons. Finally, amyloid plaque and fibril imaging in the same tissue samples used for ST enabled us to directly link changes in gene expression with accumulation and spread of pathology.

neuroscience↗

bioRxiv2023

The role of greater competitive ability in countering age disadvantages in larval competition in Drosophila melanogaster

BackgroundPopulations of Drosophila melanogaster adapted to high larval densities evolve increased larval competitive ability compared to low density controls. However, traits contributing to greater competitive ability can differ across such populations, depending on the exact details of high-density conditions imposed. In the current study, we consider three sets of D. melanogaster populations adapted to three different kinds of high-density scenarios. These population sets have evolved different degrees of increases in egg size and decreases in egg hatching time as correlates of increased larval competitive ability. QuestionWe asked two related questions: O_LIDo populations adapted to larval crowding counter an imposed initial age disadvantage in larval competition, better than their controls? C_LIO_LIDo differences in egg size and egg hatching time among crowding-adapted populations matter in competitive performance when suffering age disadvantage? C_LI Study systemWe used three sets of outbred laboratory Drosophila melanogaster populations selected for larval crowding with different egg number, food volume and vial type combinations (named MCU, CCU and LCU). We also used one set of low-density ancestral control populations (named MB). MethodsWe implemented high density cultures with half the eggs from one of the study (focal) populations, and the other half from a common marked competitor population (Orange Eye: OE). We provided head start durations of 0, 3, 5, or 7 hours to the eggs of the common competitor. This imposed the respective age disadvantage to the larvae of the focal population. Pre-adult development time of survivors was the indicator of competitive performance used. ResultsAll crowding-adapted populations countered an initial age disadvantage better than the control populations. We did not see any differences among crowding adapted populations regarding their performance in countering the age disadvantage. The common competitors showed the best competitive performance against the populations with the greatest egg size and shortest hatching time. ConclusionsAdaptation to crowding leads to significantly better chances against age disadvantages in larval competition. Temporal head starts need not be of overwhelming advantage in larval competition against superior competitors in Drosophila. Among crowding adapted populations, evolving greater egg size and shorter hatching time does not seem to better offset the effects of age disadvantage to larvae. Differences in larval effectiveness and tolerance of the populations are likely to explain these results.

evolutionary biology↗

bioRxiv2023

Differential expression of miRNAs between Young-Onset and Late-Onset Indian colorectal carcinoma patients

Early-Onset Colorectal Carcinoma (EOCRC) is a growing concern as reports indicate a worldwide increase in the incidence of CRC among young adults (<50 years old). In an effort to understand the different mode of pathogenesis in young-onset CRC, we performed a pilot study wherein we looked at colorectal tumors from both young (< 50 years old) and old patients (>55 years old) and screened them to eliminate tumors positive for Microsatellite Instability (MSI) and showing activation of the Wnt pathway, known canonical factors in CRC pathogenesis. RNA isolated from EOCRC and Late-Onset (LOCRC) tumors and paired normal tissues without MSI, nuclear {beta}-catenin and APC mutations were sent for small RNA seq to identify miRNA alterations between the two subsets. Comparative analysis revealed differential expression of 23 miRNAs specific to EOCRC and 11 miRNAs specific to LOCRC. We validated the top 10 EOCRC DEMs in TCGA-COAD cohorts followed by validation in additional EOCRC and LOCRC cohorts. Our integrated analysis revealed upregulation of hsa-miR-1247-3p, hsa-miR-148a-3p and hsa-miR-27a-5p and downregulation of hsa-miR-326 between the two subsets. Experimentally validated targets of the above miRNAs were compared with differentially expressed genes in the TCGA dataset to identify targets with physiological significance in EOCRC development. Our analysis revealed downregulation of epithelial gene expression and intercellular junction proteins potentially leading to dissolution of epithelial intercellular junctions in EOCRC development and EMT progression. Upregulated targets included genes whose expression have been reported to correlate with CRC tumor invasion, liver metastasis, disease recurrence and poor prognosis.

cancer biology↗

bioRxiv2023

Intercalation of the disulfide bond between the A2 and A4 loop of Cellobiohydrolase (Cel7A) of Aspergillus fumigatus enhances catalytic activity and thermostability

Disulfide bond is important for maintaining the structural conformation and stability of the protein. Introducing new disulfide bond is a promising strategy for rational protein design. In this report, disulfide bond engineering has been applied to improve the stability of an industrially important enzyme, Glycoside Hydrolase family GH 7 cellobiohydrolase (GH7 CBHs) or Cel7A of A fumigatus origin. Disulfide by Design 2.0 (DbD2), an online tool, was used for the detection of the mutation sites and created four mutations (D276C-G279C; DSB1, D322C-G327C; DSB2, T416C-I432C; DSB3, G460C-S465C; DSB4) both inside and outside of the peripheral loops but, not in the catalytic region. The disulfide bond formed between the A2 and A4 loop of DSB3 showed higher thermostability (70% activity at 700C), higher substrate affinity (Km= 0.081mM) and higher catalytic activity (Kcat =9.75 min-1; Kcat/Km = 120.37 mM min-1) compared to wild type AfCel7A (50% activity at 700C; Km= 0.128mM; Kcat = 4.833 min-1; Kcat/Km = 37.75 mM min -1). The other three mutants with high B factor showed loss of thermostability and catalytic activity. Molecular dynamic simulations revealed that the mutation T416C-I432C makes the tunnel wider (DSB3:13.6 [A]; Wt: 5.3 [A]) at the product exit site; giving flexibility in the entrance region and mobility of the substrate. It may facilitate substrate entry into the catalytic tunnel and releases the product faster than the wild type. Whereas in other mutants, the tunnel is not prominent (DSB4), the exit is lost (DSB1), and the ligand binding site is absent (DSB2). This is the first report of the gain of function of both thermostability and enzyme activity of cellobiohydrolase Cel7A by disulfide bond engineering in the loop. ImportanceCellulases are important for their role in the production of bioethanol, the cleanest renewable replacement of fossil fuels. Engineering of the cellulases is a chalange to increase their catalytic activity and thermostability for production of cheap ethanol. In this report we have introduced disulfide bond and successfully increased the both thermostabilty and catalytic activity of AfCel7A.

biochemistry↗

bioRxiv2023

Single Chain Fragment Variable (scFv) corresponding to a novel epitope within HCV envelope protein restricts virus entry into hepatocytes

Hepatitis C virus (HCV) is a leading cause of chronic viral hepatitis. The use of neutralizing antibodies could be a more effective therapeutic option. Previously we reported the discovery of a novel epitope at the C terminus of HCV-E2 protein, that induced potent neutralizing antibodies in the infected patients. Furthermore, monoclonal antibodies generated against this epitope could also significantly reduce virus replication in a cell culture system. In this study, we have focused on the generation of single chain variable fragments of this unique neutralizing monoclonal antibody A8A11 raised against the conserved epitope. The nucleotide sequence of the neutralizing monoclonal antibody A8A11 was determined and the scFv gene was constructed followed by cloning into the expression plasmid for recombinant protein expression. The scFv mimicked the antibody in binding to the hepatitis C virus like particles (HCV-LP). As expected, the scFv inhibited HCV-LP binding to hepatocytes and could effectively reduce viral replication in the cell culture system. More importantly, scFv A8A11 could restrict serum HCV RNA levels in HCV-infected chimeric mice harboring human hepatocytes. Results provide a basis for developing a promising scFv-based entry inhibitor, which could be more effective against viruses refractory to drugs targeting viral enzymes.

microbiology↗

bioRxiv2023

Interaction of genital microbiota in infertile couples

Bacteria colonise most of the human body and the genital tract is not an exception. While it has been known for decades that a vaginal microbiota exists, other genital sites have traditionally been viewed as sterile environments, with bacterial presence associated only with pathological conditions. However, recent studies identified specific patterns of bacterial colonisation in most genital sites. Shifts in the bacterial colonisation of the female genital tract have been linked to impairment of reproduction and adverse pregnancy outcomes, such as preterm birth. The goal of this project is to understand the association between the genital microbiota of couples seeking assisted procreation aid and the outcome of this treatment. Male and female partners will be studied as a unit ("couple microbiota") and the interaction between their microbiota will be evaluated. We have characterised microbial samples coming from vaginal and penile swabs, as well as follicular fluid and semen, using next generation sequencing (16S rRNA profiling). The results were linked to clinical data of the patients included in the study and particularly to the results of the fertility treatment process. With this project, we aim to gain a better understanding of how the male genital microbiota could influence the lower (vagina) and upper (follicular fluid) female genital tracts.

microbiology↗

bioRxiv2023

Combined effects of P25 TiO2 nanoparticles and disposable face mask leachate on microalgae Scenedesmus obliquus: Analysing the effects of heavy metals

Disposable surgical masks have been extensively employed as protective medical equipment due to the widespread breakout and transmission of the COVID-19 virus across the globe. These masks were made up of plastic polymer materials that would emit microplastics after entering the environment. Therefore, their careless disposal might lead to new and bigger microplastic contamination. The impacts of plastics that seep into waterways and their subsequent interactions with aquatic life are yet largely unexplored. In this study, we determined the quantity and kind of microplastics that were discharged from disposable surgical face masks. Furthermore, we also quantified heavy metals leached from the face masks (HML). In contrast, the increasing usage of nTiO2 in consumer items has led to its ubiquitous presence in freshwater systems. Four different concentrations of nTiO2, 0.5, 1, 2, and 4 mg L-1 were mixed with face mask leachates (FML) to perform the mixture toxicity test on freshwater algae, Scenedesmus obliquus. Reduced cell viability and photosynthetic activity were noticed in the treatment groups containing nTiO2 and FML. This was accompanied by increased oxidative stress and antioxidant activities. Furthermore, the heavy metals leached from the face masks were also tested for toxicity. In addition to that, changes in the cellular morphology were also studied with the help of FE-SEM and FTIR analysis. Our study reveals that leachates from disposable surgical face masks along with nTiO2 possess a serious threat to the environment. Environmental significanceDuring COVID-19, surgical face masks were widely used and discarded. These discarded face masks end up in lakes, rivers, and ponds. The facemasks were composed of polypropylene and other polymers. These masks release microplastics and heavy metals when discarded into water bodies. The current research focuses on assessing the environmental toxicity of the microplastics and heavy metals leached from the masks using algae as a model system. Our work further demonstrates the combined toxic effects of nTiO2 in the presence of the face mask leachate. Algae plays a crucial role as the primary producer in the freshwater ecosystem. These emerging contaminants may act as environmental stressors to the microalgae, and this may impair the ecosystems structure and function.

pharmacology and toxicology↗

bioRxiv2023

A new pipeline SPICE identifies novel JUN-IKZF1 composite elements

Transcription factor partners can cooperatively bind to DNA composite elements to augment gene transcription. Here, we report a novel protein-DNA binding screening pipeline, termed Spacing Preference Identification of Composite Elements (SPICE), that can systematically predict protein binding partners and DNA motif spacing preferences. Using SPICE, we successfully identified known composite elements, such as AP1-IRF composite elements (AICEs) and STAT5 tetramers, and also uncovered several novel binding partners, including JUN-IKZF1 composite elements. One such novel interaction was identified at CNS9, an upstream conserved noncoding region in the human IL10 gene, which harbors a non-canonical IKZF1 binding site. We confirmed cooperative binding of JUN and IKZF1 and showed that the activity of an IL10-luciferase reporter construct in primary B and T cells depended on both this site and the AP1 binding site within this composite element. Overall, our findings reveal an unappreciated global association of IKZF1 and AP1 and establish SPICE as a valuable new pipeline for predicting novel transcription binding complexes.

systems biology↗

bioRxiv2023

Separate Gut Plasma Cell Populations Produce Autoantibodies Against Transglutaminase 2 and Transglutaminase 3 in Dermatitis Herpetiformis

Dermatitis herpetiformis (DH) is an inflammatory skin disorder often considered as an extraintestinal manifestation of celiac disease (CeD). Hallmarks of CeD and DH are autoantibodies to transglutaminase 2 (TG2) and transglutaminase 3 (TG3), respectively. DH patients have autoantibodies reactive with both transglutaminase enzymes. We here report that in DH both gut plasma cells and serum autoantibodies are specific for either TG2 or TG3 with no TG2-TG3 cross-reactivity. By generating monoclonal antibodies from TG3-specific duodenal plasma cells of DH patients, we define three conformational epitope groups. Both TG2-specific and TG3-specific gut plasma cells have few immunoglobulin mutations, and the two transglutaminase-reactive populations show distinct selection of certain heavy and light chain V-genes. Mass spectrometry analysis of TG3-specific serum IgA corroborates preferential usage of IGHV2-5 in combination with IGKV4-1. Collectively, our results demonstrate parallel induction of anti-TG2 and anti-TG3 autoantibody responses involving separate B-cell populations in DH patients.

immunology↗

bioRxiv2023

Exomap1 mouse: a transgenic model for in vivo studies of exosome biology

Exosomes are small extracellular vesicles (sEVs) of [~]30-150 nm in diameter that have the same topology as the cell, are enriched in selected exosome cargo proteins, and play important roles in health and disease. To address large unanswered questions regarding exosome biology in vivo, we created the exomap1 transgenic mouse model. In response to Cre recombinase, exomap1 mice express HsCD81mNG, a fusion protein between human CD81, the most highly enriched exosome protein yet described, and the bright green fluorescent protein mNeonGreen. As expected, cell type-specific expression of Cre induced the cell type-specific expression of HsCD81mNG in diverse cell types, correctly localized HsCD81mNG to the plasma membrane, and selectively loaded HsCD81mNG into secreted vesicles that have the size ([~]80 nm), topology (outside out), and content (presence of mouse exosome markers) of exosomes. Furthermore, mouse cells expressing HsCD81mNG released HsCD81mNG-marked exosomes into blood and other biofluids. Using high-resolution, single-exosome analysis by quantitative single molecule localization microscopy, we show here that that hepatocytes contribute [~]15% of the blood exosome population whereas neurons contribute <1% of blood exosomes. These estimates of cell type-specific contributions to blood EV population are consistent with the porosity of liver sinusoidal endothelial cells to particles of [~]50-300 nm in diameter, as well as with the impermeability of blood-brain and blood-neuron barriers to particles >5 nm in size. Taken together, these results establish the exomap1 mouse as a useful tool for in vivo studies of exosome biology, and for mapping cell type-specific contributions to biofluid exosome populations. In addition, our data confirm that CD81 is a highly-specific marker for exosomes and is not enriched in the larger microvesicle class of EVs.

cell biology↗

bioRxiv2023

Honeycomb-inspired SERS nano-bowls for rapid capture and analysis of extracellular vesicles and liposomes in suspension

Nanoscale carriers such as liposomes and extracellular vesicles (EVs) are readily being explored for personalized medicine or disease prediction and diagnostics, respectively. Owing to their small size, such nanocarriers can undergo endocytosis or exocytosis, providing means to either transport cargo to the cells (liposomes) or to serve as a biomarker (EVs). When looking at current analysis methods, there is a growing need for detailed characterization of the content and composition of such nanocarriers in their natural state in aqueous media. This can be achieved through surface-enhanced Raman spectroscopy (SERS), which provides a molecular fingerprint of the analytes while reducing the detection limit. In this paper, we utilize a nano-structured SERS substrate to study different bio-nanoparticles such as liposomes, EVs and DNA nanogel in suspension. A silver-coated polydimethylsiloxane (PDMS) film-based honeycomb shaped nano-bowl surface passively traps and reduces the mobility of the nanosized bio-particles, improving the intensity and the reproducibility of the SERS signal. FDTD simulations are used for substrate geometry optimization, and a detection limit of 10-15 M is demonstrated for Rhodamine 6G (R6G). The potential of the proposed SERS nano-bowl is shown through distinct spectral features following surface-(polyethylene glycol) and bilayer-(cholesterol) modification of empty liposomes. For DNA nanogels, the characterization of highly crosslinked DNA specimens exhibits enhanced peaks for nitrogenous bases, sugar, and phosphate groups. EVs isolated from various cells provided spectral signatures of specific protein content, lipid components, and nucleic acids. Concluding, the findings of the spectral signatures of a wide range of molecular complexes and chemical morphology of bio-membranes in their natural state highlight the possibilities of using SERS as a sensitive and instantaneous characterization alternative.

biophysics↗

bioRxiv2023

SRF-deficient astrocytes provide neuroprotection in mouse models of excitotoxicity and neurodegeneration

Reactive astrogliosis is a common pathological hallmark of central nervous system (CNS) injury, infection, and neurodegeneration, where reactive astrocytes can be protective or detrimental to normal brain functions. Currently, the mechanisms regulating neuroprotective astrocytes and the extent of neuroprotection are poorly understood. Here, we report that conditional deletion of serum response factor (SRF) in adult astrocytes causes reactive-like hypertrophic astrocytes throughout the mouse brain. These SrfGFAP-ERCKO astrocytes do not affect neuron survival, synapse numbers, synaptic plasticity or learning and memory. However, the brains of Srf knockout mice exhibited neuroprotection against kainic-acid induced excitotoxic cell death. Relevant to human neurodegenerative diseases, SrfGFAP-ERCKO astrocytes abrogate nigral dopaminergic neuron death and reduce {beta}-amyloid plaques in mouse models of Parkinsons and Alzheimers disease, respectively. Taken together, these findings establish SRF as a key molecular switch for the generation of reactive astrocytes with neuroprotective functions that attenuate neuronal injury in the setting of neurodegenerative diseases.

neuroscience↗

bioRxiv2023

An expanded palette of ATP sensors for subcellular and multiplexed imaging

Genetically encoded fluorescent biosensors that detect changes in ATP levels in live cells have enabled the discovery of novel roles for ATP in cellular processes and signaling. Many of these available ATP biosensors have a limited dynamic range, or have ATP affinities that are not suitable for sensing the physiological concentrations of ATP in mammalian cells. To address these limitations, we developed a FRET-based ATP biosensor with enhanced dynamic range and signal to noise ratio, eATeam. Using eATeam, we uncovered distinct spatiotemporal dynamics of ATP changes upon inhibition of cellular energy production. We also developed dimerization-dependent GFP and RFP-based ATP biosensors with enhanced dynamic ranges compared to the current standard in the field. Using the single-color ATP biosensors, we visualized the complex interplay between AMPK activity, ATP, lactate, and calcium by multiplexed imaging in single cells. This palette of ATP sensors expands the toolbox for interrogating subcellular ATP regulation and metabolic signaling in living cells.

cell biology↗

bioRxiv2023

Pharmacological Induction of mesenchymal-epithelial transition chemosensitizes breast cancer cells and prevents metastatic progression

The epithelial-mesenchymal transition (EMT) is a developmental program co-opted by tumor cells that aids the initiation of the metastatic cascade. Tumor cells that undergo EMT are relatively chemoresistant, and there are currently no therapeutic avenues specifically targeting cells that have acquired mesenchymal traits. We show that treatment of mesenchymal-like triple-negative breast cancer (TNBC) cells with the microtubule-destabilizing chemotherapeutic eribulin, which is FDA-approved for the treatment of advanced breast cancer, leads to a mesenchymal-epithelial transition (MET). This MET is accompanied by loss of metastatic propensity and sensitization to subsequent treatment with other FDA-approved chemotherapeutics. We uncover a novel epigenetic mechanism of action that supports eribulin pretreatment as a path to MET induction that curtails metastatic progression and the evolution of therapy resistance. SummaryWhile the advent of targeted therapy has led to vast improvements in outcomes for certain types of breast cancer, a mainstay for triple-negative breast cancer (TNBC) remains cytotoxic chemotherapy. A major clinical hurdle in successfully managing this disease is the eventual development of therapeutic resistance and disease relapse in more aggressive forms. Our data reveal that epigenetic modulation of EMT state using the FDA-approved therapeutic eribulin curtails metastatic propensity of breast tumors and, when administered in the treatment-naive setting, sensitizes to subsequent treatment with other chemotherapeutics.

cancer biology↗

bioRxiv2023

Native and engineered human megakaryocytic extracellular vesicles for targeted non-viral cargo delivery to blood stem cells

Native and engineered extracellular vesicles (EVs) generated from human megakaryocytes (huMkEVs) or from the human megakaryocytic cell line CHRF (CHEVs) interact with tropism delivering their cargo to both human and murine hematopoietic stem and progenitor cells (HSPCs). 24 hours after intravenous infusion of huMkMPs into NOD-scid IL2R{gamma}null (NSG) mice, they induced a nearly 50% increase in murine platelet counts relative to saline control, thus demonstrating the potential of these EVs, which can be stored frozen, for treating thrombocytopenias. PKH26-labeled huMkMPs or CHEVs localized to the HSPC-rich bone marrow preferentially interacting with murine HSPCs. Using engineered huMkEVs or CHEVs, their receptor-mediated tropism for HSPCs was explored to functionally deliver synthetic cargo, notably plasmid DNA coding for a fluorescent reporter, to murine HSPCs both in vitro and in vivo. These data demonstrate the potential of these EVs as a non-viral, HSPC-specific cargo vehicle for gene therapy applications to treat hematological diseases. Native and engineered human megakaryocytic extracellular vesicles for targeted non-viral cargo delivery to blood stem cells (Table of Contents): O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=111 SRC="FIGDIR/small/536479v1_ufig1.gif" ALT="Figure 1"> View larger version (49K): org.highwire.dtl.DTLVardef@664768org.highwire.dtl.DTLVardef@d9fe13org.highwire.dtl.DTLVardef@1b795eforg.highwire.dtl.DTLVardef@1d48e2d_HPS_FORMAT_FIGEXP M_FIG C_FIG Graphical Overview: Native and engineered human megakaryocytic extracellular vesicles (huMkEVs) for provide targeted non-viral cargo delivery to blood stem cells. We demonstrate that huMkEVs as a transformational cargo-delivery system to blood stem cells (hematopoietic stem and progenitor cells, HSPCs) in NOD-scid IL2R{gamma}null (NSG) mice. Intravenous delivery of native huMkEVs enhances de novo platelet biogenesis by inducing megakaryocytic differentiation of murine HSPCs, thus demonstrating the desirable strong tropism of huMkEVs for murine HSPCs. Based on this tropism, we demonstrate that engineered huMkEVs can deliver functional plasmid-DNA cargo specifically to HSPCs.

bioengineering↗

bioRxiv2023

Hagfish genome illuminates vertebrate whole genome duplications and their evolutionary consequences

Whole genome duplications (WGDs) are major events that drastically reshape genome architecture and are causally associated with organismal innovations and radiations1. The 2R Hypothesis suggests that two WGD events (1R and 2R) occurred during early vertebrate evolution2, 3. However, the veracity and timing of the 2R event relative to the divergence of gnathostomes (jawed vertebrates) and cyclostomes (jawless hagfishes and lampreys) is unresolved4-6 and whether these WGD events underlie vertebrate phenotypic diversification remains elusive7. Here we present the genome of the inshore hagfish, Eptatretus burgeri. Through comparative analysis with lamprey and gnathostome genomes, we reconstruct the early events in cyclostome genome evolution, leveraging insights into the ancestral vertebrate genome. Genome-wide synteny and phylogenetic analyses support a scenario in which 1R occurred in the vertebrate stem-lineage during the early Cambrian, and the 2R event occurred in the gnathostome stem-lineage in the late Cambrian after its divergence from cyclostomes. We find that the genome of stem-cyclostomes experienced two additional, independent genome duplications (herein CR1 and CR2). Functional genomic and morphospace analyses demonstrate that WGD events generally contribute to developmental evolution with similar changes in the regulatory genome of both vertebrate groups. However, appreciable morphological diversification occurred only after the 2R event, questioning the general expectation that WGDs lead to leaps of morphological complexity7.

genomics↗

bioRxiv2023

p53 translational isoform {triangleup}40p53 orchestrates cellular SGSH levels via microRNA-4671-5p to modulate cell cycle

{Delta}40p53, the only translational isoform of p53, modulates the full-length p53 (FLp53) activity and independently regulates targets such as the miR-186-5p-YY1 axis. To identify additional miRNAs regulated by {Delta}40p53, we performed small RNA sequencing. We found that overexpression of {Delta}40p53, but not FLp53, significantly downregulated miR-4671-5p. Expression of both isoforms at varying ratios revealed that miR-4671-5p may be modulated by FLp53 in a {Delta}40p53-dependent manner. In silico analysis identified SGSH (N-sulfoglucosamine sulfohydrolase) as a potential miR-4671-5p target. SGSH expression showed inverse correlation with miR-4671-5p in cancer datasets and prognostic significance. SGSH mRNA and protein levels were reduced upon miR-4671-5p overexpression or si{Delta}40p53 treatment, confirming regulatory linkage. Functionally, miR-4671-5p overexpression induced intra-S-phase cell cycle arrest, implicating SGSH in cell cycle regulation. These results reveal a novel {Delta}40p53-miR-4671-5p-SGSH axis that impacts cell cycle progression and may contribute to cancer outcomes. Our findings highlight the distinct regulatory role of {Delta}40p53, independent of FLp53, in maintaining cellular and metabolic homeostasis via miRNA-mediated mechanisms.

molecular biology↗

bioRxiv2023

Incidences of Helicobacter infection in pigs and tracing occupational hazard in pig farmers

Helicobacter species (H. sp.) is a gram-negative spiral-shaped motile bacteria that causes gastritis in pigs and also colonizes the human stomach. The current study seeks to assess the prevalence of various H. sp. in the gastric mucosa of slaughtered and dead pigs, as well as the prevalence of Helicobacter infection among pig farmers. A total of 403 stomach samples from various pig slaughter points, 74 necropsy samples from various pig farms and 97 stool samples from pig farmers were collected from Assam, India. Among 477 pig stomach samples tested, 214 samples with gastritis (20.09%) showed Gram negative, spiral-shaped organisms in brush cytology from the mucosal surface, and the rest of the 263 stomach samples without any gastric lesion showed only 3.04% Gram negative, spiral-shaped organisms. In ultrastructure investigation, Scanning Electron Microscopy (SEM) of the four urease positive stomach samples revealed a tightly coiled Helicobacter bacterium (spiral-shaped) found in the mucous lining of the stomach. In histopathological examination of pars esophagia, cardiac and fundic mucosa showed chronic gastritis associated with hemorrhagic necrosis, leucocytic infiltration with neutrophils and macrophages, and lymphoid aggregates (lymphoid follicles) etc. PCR confirmed 16S rRNA genes of Helicobacter suis (H. suis) where a total of 42 (19.63%) out of 214 pig stomach samples and 2 (2.08%) out of 96 stool samples of pig farmers were found positive for H. suis. of these 96 stool samples of pig farmers 3 (3.12%) were confirmed positive for Helicobacter pylori (H. pylori) Phosphoglucosamine mutase gene in PCR. Phylogenic analysis of the 16S rRNA gene of H. suis showed distinct clusters with other H. sp. In conclusion, this study provides evidence for the prevalence of Helicobacter both in pig gastric mucosa and human stool. The findings highlight the need for improved sanitation and hygiene practices among pig farmers to minimize the risk of Helicobacter infection in humans.

microbiology↗