Biology subjects

Dutta, S.

Publications and source records attributed to Dutta, S..

At least 127 records · Page 7Linked to original sources

bioRxiv2022

Glycyrrhizin, an inhibitor of HMGB1 induces autolysosomal degradation function and inhibits H. pylori infection

Helicobacter pylori a key agent for causing gastric complications is linked with peptic ulcer, gastritis, and in severe cases gastric cancer. In response to infection, host cells stimulate autophagy to maintain cellular homeostasis. However, H. pylori have evolved the ability to usurp the hosts autophagic machinery. High mobility group box1 (HMGB1), an alarmin molecule is a regulator of autophagy and its expression is augmented in gastric cancer and many other cancers. Therefore, this study aims to explore the role of glycyrrhizin (a known inhibitor of HMGB1) in autophagy during H. pylori infection. Human gastric cancer (AGS) cells were infected with H. pylori SS1 strain and further treatment was done with glycyrrhizin. Western blot was used to examine the expression levels of autophagy proteins. Autophagy and lysosomal activity were monitored by immunofluorescence. We have performed knockdown of HMGB1 to verify the effect of glycyrrhizin by siRNA transfection method. H. pylori-infection in vivo C56BL/6 mice model was established and the effect of glycyrrhizin treatment was studied. We found that the autophagy-lysosomal pathway was impaired due to a significant increase in lysosomal membrane permeabilization during H. pylori infection in AGS cells. Subsequently, glycyrrhizin treatment restored the lysosomal membrane integrity, accompanied by an increase in cathepsin B activity and reduction of ROS and inflammatory cytokine IL-8. The recovered lysosomal function enhanced autolysosome formation and concomitantly attenuated the intracellular H. pylori growth by eliminating the pathogenic niche from gastric cells. Additionally, glycyrrhizin treatment inhibited inflammation and improved gastric tissue damages in mice.

microbiology↗

bioRxiv2022

Recombinant Human ACE2-Fc : A promising therapy for SARS-CoV2 infection

SARS-CoV2 entry is mediated by binding of viral spike-protein(S) to the transmembrane Angiotensin-Converting Enzyme-2 (ACE2) of the host cell. Thus, to prevent transmission of disease, strategies to abrogate the interaction are important. However, ACE2 cannot be blocked since its normal function is to convert the Angiotensin II peptide to Angiotensin(1-7) to reduce hypertension. This work reports a recombinant cell line secreting soluble ACE2-ectopic domain (MFcS2), modified to increase binding and production efficacy and fused to human immunoglobulin-Fc. While maintaining its enzymatic activity, the molecule trapped and neutralized SARS CoV2 virus in vitro with an IC50 of 64 nM. In vivo, with no pathology in the vital organs, it inhibited the viral load in lungs in SARS-CoV2 infected Golden-Syrian-hamster. The Intravenous pharmacokinetic profiling of MFcS2 in hamster at a dose of 5 mg/Kg presented a maximum serum concentration of 23.45 {micro}g/mL with a half-life of 29.5 hrs. These results suggest that MFcS2 could be used as an effective decoy based therapeutic strategy to treat COVID19. This work also reports usage of a novel oral-cancer cell line as in vitro model of SARS-Cov2 infection, validated by over expressing viral-defence pathways upon RNA-seq analysis and over-expression of ACE2 and TMPRSS upon growth in hyperglycaemic condition.

molecular biology↗

bioRxiv2022

Genomic analysis unveils the role of genome degradation events and gene flux in the emergence and persistence of S. Paratyphi A lineages

Paratyphoid fever caused by S. Paratyphi A is endemic in parts of Asia and Sub-Saharan Africa. The proportion of enteric fever cases caused by S. Paratyphi A has substantially increased, yet only limited data is available on the population structure and genetic diversity of this serovar. We examined the phylogenetic distribution and evolutionary trajectory of S. Paratyphi A isolates collected as part of the Indian enteric fever surveillance study "Surveillance of Enteric Fever in India (SEFI)." In the study period (2017-2020), S. Paratyphi A comprised 17.6% (441/2503) of total enteric fever cases in India, with the isolates highly susceptible to all the major antibiotics used for treatment except fluoroquinolones. Phylogenetic analysis clustered the global S. Paratyphi A collection into seven lineages (A-G), and the present study isolates were distributed in lineages A, C and F. Our analysis documented that the genome degradation events and gene acquisitions or losses play a major role in the evolution of new S. Paratyphi A lineages/sub-lineages. A total of 10 pseudogene-forming mutations possibly associated with the emergence of lineages were identified. Pan-genome analysis identified the insertion of P2/PSP3 phage and acquisition of IncX1 plasmid during the selection in 2.3.2/2.3.3 and 1.2.2 genotypes, respectively. We also identified that the six characteristic missense mutations associated with the lipopolysaccharide (LPS) biosynthesis genes of S. Paratyphi A confer only a low structural impact and would therefore have minimal impact on vaccine effectiveness. Since S. Paratyphi A is human restricted, high levels of genetic drift are not expected unless these bacteria transmit to naive hosts. However, public-health investigation and intervention by means of genomic surveillance would be continually needed to avoid S. Paratyphi A serovar becoming a public health threat similar to the S. Typhi of today.

genomics↗

bioRxiv2022

Activation Mechanism of the Human Smoothened Receptor

Smoothened (SMO) is a membrane protein of the Class F subfamily of G-Protein Coupled Receptors (GPCRs) and maintains homeostasis of cellular differentiation. SMO undergoes conformational change during activation, transmitting the signal across the membrane, making it amenable to bind to its intracellular signaling partner. Receptor activation has been studied at length for Class A receptors, but the mechanism of Class F receptor activation remain unknown. Agonists and antagonists bound to SMO at sites in the Transmembrane Domain (TMD) and the Cysteine Rich Domain have been characterized, giving a static view of the various conformations SMO adopts. While the structures of the inactive and active SMO outline the residue-level transitions, a kinetic view of the overall activation process remains unexplored for Class F receptors. We describe SMOs activation process in atomistic detail by performing 300 s of molecular dynamics simulations and combining it with Markov state model theory. A molecular switch, conserved across Class F and analogous to the activation-mediating D-R-Y motif in Class A receptors, is observed to break during activation. We also show that this transition occurs in a stage-wise movement of the transmembrane helices - TM6 first, followed by TM5. To see how modulators affect SMO activity, we simulated agonist and antagonist-bound SMO. We observed that agonist-bound SMO has an expanded hydrophobic tunnel in SMOs core TMD, while antagonist-bound SMO shrinks this tunnel, further supporting the hypothesis that cholesterol travels through a tunnel inside Smoothened to activate it. In summary, this study elucidates the distinct activation mechanism of Class F GPCRs and shows that SMOs activation process rearranges the core transmembrane domain to open a hydrophobic conduit for cholesterol transport.

biophysics↗

bioRxiv2022

Differential structural organization and gene expression regulatory networks of lamin A Ig fold domain mutants of muscular dystrophy.

In metazoan cells, lamins form a proteinaceous meshwork as a major structural component of the nucleus and also play a role in regulating essential cellular processes. Lamins, along with their interactors, act as determinants for chromatin organization throughout the nucleus. The major dominant missense mutations responsible for autosomal dominant forms of muscular dystrophies reside in the Ig fold domain of lamin A. However, how lamin A contributes to the distribution of heterochromatin and balances euchromatin, and how it relocates epigenetic marks to shape chromatin states, remains poorly defined, making it difficult to draw conclusions about the prognosis of lamin A-mediated muscular dystrophies.In the first part of this report, we identified the in-vitro organization of full-length lamin A proteins due to two well-documented Ig LMNA mutations, R453W and W514R, through biophysical and electron microscopy observations. We further demonstrated that both lamin A/C mutant cells predominantly expressed nucleoplasmic aggregates with reduced amounts at the nuclear envelope. Labeling specific markers of epigenetics, such as H3K9me3, H3K27me3, H3K36me3, and HP1, allowed correlation of lamin A mutations with epigenetic mechanisms. Immunofluorescence and biochemical analyses indicated transcriptional upregulation. In addition to manipulating epigenetic mechanisms, our proteomic studies traced diverse expressions of transcription regulators, RNA synthesis and processing proteins, protein translation components, and posttranslational modifications. These data suggest severe perturbations in targeting other proteins to the nucleus.Our study also predicts specific structural configurations of lamin A determined by its interacting partners patterns. SignificanceO_LIThis study investigates the effects of two severe mutations in the LMNA gene associated with muscular dystrophies: R453W and W514R. C_LIO_LIThis research aims to provide new insights into the structural environment of full-length lamin A protein due to single-point mutations. C_LIO_LIOur study employs proteomics to map expression levels of regulators involved in epigenetic states, RNA synthesis and processing, and protein synthesis and processing. C_LIO_LIIt assesses potential effects on binding affinity and transcription, as well as alterations in heterochromatin protein distribution and RNA polymerase II functionality due to mutations affecting lamin A. C_LIO_LIA systematic investigation of lamin A interactors is conducted to explore their associations with other proteins. C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=67 SRC="FIGDIR/small/493956v2_ufig1.gif" ALT="Figure 1"> View larger version (18K): org.highwire.dtl.DTLVardef@10ac33dorg.highwire.dtl.DTLVardef@197fa20org.highwire.dtl.DTLVardef@a679daorg.highwire.dtl.DTLVardef@18a03a7_HPS_FORMAT_FIGEXP M_FIG Figure: Schematic diagram representing the results. Mutations in lamin A protein evicts the nucleosome from nuclear periphery and consequently promotes aberrant gene expression. C_FIG

biochemistry↗

bioRxiv2022

Machine learning for lumbar and pelvis kinematics clustering

Clustering algorithms such as k-means and agglomerative hierarchical clustering (HCA) may provide a unique opportunity to analyze time-series kinematic data. Here we present an approach for determining number of clusters and which clustering algorithm to use on time-series lumbar and pelvis kinematic data. Cluster evaluation measures such as silhouette coefficient, elbow method, Dunn Index, and gap statistic were used to evaluate the quality of decision making. The result show that multiple clustering evaluation methods should be used to determine the ideal number of clusters and algorithm suitable for clustering time-series data for each dataset being analyzed.

physiology↗

bioRxiv2022

ERK pathway activation inhibits ciliogenesis and causes defects in motor behavior, ciliary gating, and cytoskeletal rearrangement

Mitogen-activated protein kinase (MAPK) pathways are well known regulators of the cell cycle but they have also been found to control ciliary length in a wide variety of organisms and cell types from Caenorhabditis elegans neurons to mammalian photoreceptors through unknown mechanisms. ERK1/2 is a MAP kinase in human cells that is predominantly phosphorylated by MEK1/2 and dephosphorylated by the phosphatase DUSP6. We have found that the ERK1/2 activator/DUSP6 inhibitor, (E)-2-benzylidine-3-(cyclohexylamino)-2,3-dihydro-1H-inden-1-one (BCI), inhibits ciliary maintenance in Chlamydomonas and hTERT-RPE1 cells and assembly in Chlamydomonas. These effects involve inhibition of total protein synthesis, microtubule organization, membrane trafficking, and partial kinesin-2 motor dynamics. Our data provide evidence for various avenues for BCI-induced ciliary shortening and impaired ciliogenesis that gives mechanistic insight into how MAP kinases can regulate ciliary length.

cell biology↗

bioRxiv2022

Thermal endurance by a hot-spring-dwelling phylogenetic relative of the mesophilic Paracoccus

High temperature growth/survival was revealed in a phylogenetic relative (strain SMMA_5) of the mesophilic Paracoccus isolated from the 78-85{degrees}C water of a Trans- Himalayan sulfur-borax spring. After 12 h at 50{degrees}C, or 45 minutes at 70{degrees}C, in mineral salts thiosulfate (MST) medium, SMMA_5 retained [~]2% colony-forming units (CFUs), whereas comparator Paracoccus had 1.5% and 0% CFU left at 50{degrees}C and 70{degrees}C respectively. After 12 h at 50{degrees}C, the thermally-conditioned sibling SMMA_5_TC exhibited [~]1.5 time increase in CFU-count; after 45 minutes at 70{degrees}C, SMMA_5_TC had 7% of the initial CFU-count intact. 1000-times diluted Reasoners 2A medium, and MST supplemented with lithium, boron or glycine-betaine (solutes typical of the SMMA_5 habitat), supported higher CFU-retention/CFU-growth than MST. With or without lithium/boron/glycine-betaine in MST, a higher percentage of cells always remained viable (cytometry data), compared with what percentage remained capable of forming single colonies (CFU data). SMMA_5, compared with other Paracoccus, contained 335 unique genes, mostly for DNA replication/recombination/repair, transcription, secondary metabolites biosynthesis/transport/catabolism, and inorganic ion transport/metabolism. Its also exclusively enriched in cell wall/membrane/envelope biogenesis, and amino acid metabolism, genes. SMMA_5 and SMMA_5_TC mutually possessed 43 nucleotide polymorphisms, of which 18 were in protein-coding genes with 13 nonsynonymous and seven radical amino acid replacements. Such biochemical and biophysical mechanisms could be involved in thermal stress mitigation which streamline the cells energy and resources towards system-maintenance and macromolecule-stabilization, thereby relinquishing cell-division for cell-viability. Thermal conditioning apparently helped memorize those potential metabolic states which are crucial for cell-system maintenance, while environmental solutes augmented the indigenous stability-conferring mechanisms. IMPORTANCEFor a holistic understanding of microbial lifes high-temperature adaptation it is imperative to explore the biology of the phylogenetic relatives of mesophilic bacteria which get stochastically introduced to geographically and geologically diverse hot spring systems by local geodynamic forces. Here, in vitro endurance of high heat up to the extent of growth under special (habitat-inspired) conditions was discovered in a hot- spring-dwelling phylogenetic relative of the mesophilic Paracoccus species. Thermal conditioning, extreme oligotrophy, metabolic deceleration, presence of certain habitat- specific inorganic/organic solutes, and typical genomic specializations were found to be the major enablers of this conditional (acquired) thermophilicity. Feasibility of such phenomena across the taxonomic spectrum can well be paradigm-changing for the established scopes of microbial adaptation to the physicochemical extremes. Applications of conditional thermophilicity in microbial process biotechnology may be far reaching and multi-faceted.

microbiology↗

bioRxiv2022

Assessment of local potato cultivars found in Cis-Himalayan region of West Bengal through morphology and biochemical profiling

Potato is a major global food crop grown for tubers (underground storage stems) that are high in carbohydrates, minerals, and vitamins. The presence of anthocyanins in tuber skin and flesh can have a significant impact on customer preferences. Potatoes are also high in resistant starches, which have a variety of health benefits, including enhanced fullness, cholesterol reduction, and a reduced risk of colon cancer etc. In West Bengal, diabetes is prevalent. Similarly, according to ICMR studies, colon cancer affects 8.9% of all cancer patients in West Bengal cancer which is caused by ill eating habits or the consumption of foods that are low in antioxidants. 5-10% of colon cancers are genetically caused, while the remainder are caused by poor eating habits or the consumption of foods that are low in antioxidants. To address these issues, one strategy is to eat foods with a low glycemic index and high antioxidant content as a staple food. The tuber tissues of the potato (S. tuberosum L.) accumulate various quantities of anthocyanins, which are commonly consumed around the world. Anthocyanins are pigments that range in colour from red to purple and are found throughout the plant kingdom. Anthocyanins are powerful antioxidants that are water soluble. The red skin potato is well-known among the general public. The epidermal layer contains a high quantity of red anthocyanins, which causes the skin to be red. With this background the present study has been undertaken to address the following objectives morphological (quantitative and qualitative traits) and biochemical characterization of local potato cultivars and identification of most stable genotypes based on the anthocyanin content and tuber yield of local potato cultivars. For our recent research 14 local potato cultivar from North Bengal were collected and evaluated for DUS characterization showed wide range of variability with respect to different phenotypic variants of ten characters. Overall predominant DUS characters of local potato cultivars found in North Bengal are medium sized apical length of sprout, short stem height, compact foliage structure, open leaf structure, ovate lanceolate type leaflet shape, purple leaf sprout predominant colour, spherical leaf sprout shape, medium intensity of anthicyanin coloraton at the base, light intensity of anthicyanin coloraton at the tip and weak nature of light sprout pubescence base. The results of the present investigation suggest that local potato cultivars collected from the northern part of West Bengal showed a high level of genetic variation. The differences between genotypes were highly significant at the 1% and 5% alpha level for all characters like tuber yield, length and weight of tuber, length of sprout, chlorophyll content, canopy temperature, and number of tuber per plant, according to the analysis of variance. Three quantitative traits namely tuber yield, tuber breadth, tuber length played major role in the genetic variance. Anthocyanin content had more contribution to diversify the local potato cultures according to biochemical characterization AMMI analysis suggested that Jalpai from CoochBehar is the most stable potato cultivars in respect to anthocyanin content and tuber yield per plant.

plant biology↗

bioRxiv2022

A critical developmental interval of coupling axon branching to synaptic degradation during neural circuit formation

The emergence of neuronal wiring specificity requires stabilization of dynamic axonal branches at sites of selective synapse formation. Models that explain how axonal branching is coupled to synaptogenesis postulate molecular regulators acting in a spatiotemporally restricted fashion. We report that Epidermal Growth Factor Receptor (EGFR) activity is required in presynaptic axonal branches during two distinct temporal intervals to regulate circuit wiring in the developing Drosophila visual system. EGFR is required early to regulate primary axonal branching and independently again later to prevent autophagic degradation of the synaptic active zone protein Bruchpilot (Brp). The protection of synaptic material during this later interval of wiring ensures the stabilization of terminal branches, circuit connectivity and appropriate visual behavior. Phenotypes of EGFR inactivation were rescued by increasing Brp levels or downregulating autophagic genes. We identify a temporally restricted molecular mechanism required for coupling axonal branching and synaptic stabilization that contributes to the emergence of neuronal wiring specificity.

neuroscience↗

bioRxiv2022

Evolution of Delta variant by non-Spike signature co-appearing mutations: trailblazer of COVID-19 disease outcome

The high transmissibility and infectivity of a SARS-CoV-2 variant is usually ascribed to the Spike mutations, while emerging non-spike mutations might be a serious threat to the current Spike-recombinant vaccines. In addition to mutations in structural Spike glycoprotein, rapid accumulation of mutations across non-structural genes is leading to continuous virus evolution, altering its pathogenicity. We performed whole genome sequencing of SARS-CoV-2 positive samples collected from different clinical groups from eastern India, during the second pandemic wave (April-May, 2021). In addition to the several common spike mutations in Delta variant, two mutually explicit signature constellations of non-spike co-appearing mutations were identified, driving symptomatic and asymptomatic infections. We attempted to correlate these unique signatures of non-Spike co-appearing mutations to COVID-19 disease outcome. Results revealed that the Delta strains harboring a unique constellation of 9 non-spike co-appearing mutations could be the wheeler and dealer of symptomatic infection, even post vaccination. The strains predominantly driving asymptomatic infection possessed 7 non-spike co-appearing mutations, which were mutually exclusive in contrast to the set of mutations causing symptomatic disease. Phylodynamic analysis depicted high probability of emergence of these unique sub-clusters within India, with subsequent spread worldwide. Interestingly, some mutations of this signature were selected in Omicron and IHU variants, which suggest that gradual accumulation of such co-existing mutations may lead to emergence of more "vaccine-evading variants" in future. Hence, unfaltering genome sequencing and tracking of non-Spike mutations might be significant in formulation of any future vaccines against emerging SARS-CoV-2 variants that might evade the current vaccine-induced immunity.

genomics↗

bioRxiv2022

Interconnection between molecular regulators in LMNA related muscular dystrophy

Nuclear lamina is composed of different A-type and B-type lamin proteins and acts as major regulator of DNA replication, transcription, heterochromatin-euchromatin machinery. The majority of mutations in A type lamin are associated with some forms of muscular dystrophies,that majorly distinguishable by a common clinical feature : progressive skeletal muscle wasting. This speculates impaired skeletal muscle differentiation during development and after injury with the influence of myriad of signalling pathways. The molecular mechanism behind phenotypes of lamin A associated muscular dystrophies are still elusive . Here, we used genome wide expression analysis platform during the differentiation of murine C2C12 skeletal muscle cells to investigate the factors have been attributed to the disease. We compared the expression levels of the components of pathways that indicates important role of skeletal muscle as well as identified gene regulatory networks at two different time points of muscle differentiation in wild type and mutant cells. We also report significant perturbations in the expression and activation of Wnt signalling pathway in mutant cells among 40 dysregulated signalling pathways , presenting pronounced regulation of normal downstream myogenic signalling . Finally, with this largest data sets we evaluate in depth characterization of molecular effectors for myogenic differentiation, which could allow greater insight into development of therapeutic strategies for the remission of patients with LMNA linked muscle - related pathologies.

cell biology↗

bioRxiv2022

CD36 homologs determine microbial resistance to the Lyme disease spirochete

Pattern recognition receptors sense pathogens in arthropods and mammals through distinct immune processes. Whether these molecules share a similar function and recognize the same microbe in evolutionarily distant species remain ill-defined. Here, we establish that the CD36 superfamily is required for Borrelia burgdorferi resistance in both the arthropod vector and humans. Using the blacklegged tick Ixodes scapularis and an electronic health record-linked biobank, we demonstrate that CD36 members elicit immunity to the Lyme disease spirochete. In ticks, the CD36-like protein Croquemort recognizes lipids and initiates the immune deficiency and jun N-terminal kinase pathways against B. burgdorferi. In humans, exome sequencing and clinical information reveal that individuals with CD36 loss-of-function variants have increased prevalence of Lyme disease. Altogether, we discovered a conserved mechanism of anti-bacterial immunity. One Sentence SummaryLipid receptors belonging to the CD36 superfamily exhibit a shared immune function in both ticks and humans.

immunology↗

bioRxiv2022

Evaluation of lamin A/C mechanotransduction under different surface topography in LMNA related muscular dystrophy.

Most of the single point mutations of the LMNA gene are associated with distinct muscular dystrophies, marked by heterogenous phenotypes but primarily the loss and symmetric weakness of skeletal muscle tissue. The molecular mechanism and phenotype-genotype relationships in these muscular dystrophies are poorly understood. An effort has been here to delineating the adaptation of mechanical inputs into biological response by mutant cells of lamin A associated muscular dystrophy. In this study we implement engineered smooth and pattern surfaces of particular young modulus to mimic muscle physiological range. Using fluorescence and atomic force microscopy we present distinct architecture of the actin filament along with abnormally distorted cell and nuclear shape in mutants, which showed a tendency to deviate from wild type cells. Topographic features of pattern surface antagonizes the binding of the cell with it. Correspondingly, from the analysis of genome wide expression data in wild type and mutant cells, we report differential expression of the gene products of the structural components of cell adhesion as well as LINC (linkers of nucleoskeleton and cytoskeleton) protein complexes. This study also reveals mis expressed downstream signaling processes in mutant cells, which could potentially lead to onset of the disease upon the application of engineered materials to substitute the role of conventional cues in instilling cellular behaviors in muscular dystrophies. Collectively, these data support the notion that lamin A is essential for proper cellular mechanotransduction from extracellular environment to the genome and impairment of the muscle cell differentiation in the pathogenic mechanism for lamin A associated muscular dystrophy.

cell biology↗

bioRxiv2021

Engineered High-Affinity ACE2 Peptide Mitigates ARDS and Death Induced by Multiple SARS-CoV-2 Variants

Vaccine hesitancy and continuing emergence of SARS-CoV-2 variants of concern that may escape vaccine-induced immune responses highlight the urgent need for effective COVID-19 therapeutics. Monoclonal antibodies used in the clinic have varying efficacies against distinct SARS-CoV-2 variants; thus, there is considerable interest in engineered ACE2 peptides with augmented binding affinities for SARS-CoV-2 Spike protein. These could have therapeutic benefit against multiple viral variants. Using molecular dynamics simulations, we show how three amino acid substitutions in an engineered soluble ACE2 peptide (sACE22.v2.4-IgG1) markedly increase affinity for the SARS-CoV-2 Spike (S) protein. We demonstrate high binding affinity to S protein of the early SARS-CoV-2 WA-1/2020 isolate and also to multiple variants of concern: B.1.1.7 (Alpha), B.1.351 (Beta), P.1 (Gamma), and B.1.617.2 (Delta) SARS-CoV-2 variants. In humanized K18-hACE2 mice, prophylactic and therapeutic administration of sACE22.v2.4-IgG1 peptide prevented acute lung vascular endothelial injury and lung edema (essential features of ARDS) and significantly improved survival after infection by SARS-CoV-2 WA-1/2020 as well as P.1 variant of concern. These studies demonstrate for the first time broad efficacy in vivo of an ACE2 decoy peptide against multiple SARS-CoV-2 variants and point to its therapeutic potential.

pharmacology and toxicology↗

bioRxiv2021

BrnQ-type Branched-chain Amino Acid Transporters Influence Bacillus anthracis Growth and Virulence

Bacillus anthracis, the anthrax agent, exhibits robust proliferation in diverse niches of mammalian hosts. Metabolic attributes of B. anthracis that permit rapid growth in multiple mammalian tissues have not been established. We posit that branched-chain amino acid (BCAA: Isoleucine, leucine and valine) metabolism is key to B. anthracis pathogenesis. Increasing evidence indicates relationships between B. anthracis virulence and expression of BCAA-related genes. Expression of some BCAA-related genes is altered during culture in bovine blood in vitro and the bacterium exhibits valine auxotrophy in a blood serum mimic medium. Transcriptome analyses have revealed that the virulence regulator AtxA, that positively affects expression of the anthrax toxin and capsule genes, negatively regulates genes predicted to be associated with BCAA biosynthesis and transport. Here, we show that B. anthracis growth in defined media is severely restricted in the absence of exogenous BCAAs, indicating that BCAA transport is required for optimal growth in vitro. We demonstrate functional redundancy among multiple BrnQ-type BCAA transporters. Three transporters are associated with isoleucine and valine transport, and deletion of one, BrnQ3, attenuates virulence in a murine model for anthrax. Interestingly, an ilvD-null mutant lacking dihydroxy-acid dehydratase, an enzyme essential for BCAAs synthesis, exhibits unperturbed growth when cultured in media containing BCAAs, but is highly attenuated in the murine model. Finally, our data show that BCAAs enhance AtxA activity in a dose-dependent manner, suggesting a model in which BCAAs serve as a signal for virulence gene expression. IMPORTANCEInfection with B. anthracis can result in systemic disease with large numbers of the bacterium in multiple tissues. We found that BCAA synthesis is insufficient for robust growth of B. anthracis; access to branched-chain amino acids (BCAAs) is necessary for proliferation of the pathogen during culture and during infection in a murine model for anthrax. B. anthracis produces an unusually large repertoire of BCAA-related transporters. We identified three isoleucine/valine transporters with partial functional redundancy during culture. Deletion of one of these transporters, BrnQ3, resulted in attenuated virulence. Interestingly, a BCAA biosynthesis mutant grew well in medium containing BCAAs, but like BrnQ3, was attenuated for virulence. These results suggest that BCAAs are limiting in multiple niches during infection and furthers understanding of nutritional requirements of this important pathogen.

microbiology↗

bioRxiv2021

Telomere-Dependent Interleukin-1 Receptor Activation Promotes Immune Suppression in Triple-Negative-Breast Cancer

Telomeres are crucial for cancer progression. Immune signalling in the tumour microenvironment has been shown to be very important in cancer prognosis. However, the mechanisms by which telomeres might affect tumour immune response remain poorly understood. Here, we observed that interleukin-1 signalling is telomere-length dependent in cancer cells. Mechanistically, non-telomeric TRF2 (Telomeric Repeat binding Factor 2) binding at the IL-1-receptor type-1 (IL1R1) promoter was found to be affected by telomere length. Enhanced TRF2 binding at the IL1R1 promoter in cells with short telomeres directly recruited the histone-acetyl-transferase (HAT) p300, and consequent H3K27 acetylation activated IL1R1. This altered NF-kappa B signalling and affected downstream cytokines like IL6, IL8 and TNF. Further, IL1R1 expression was telomere-sensitive in triple-negative breast cancer (TNBC) clinical samples. Infiltration of tumour-associated macrophages (TAM) was also sensitive to the length of tumour cell telomeres and highly correlated with IL1R1 expression. The use of both IL1 Receptor antagonist (IL1RA) and IL1R1 targeting ligands could abrogate M2 macrophage infiltration in TNBC tumour organoids. In summary, using TNBC cancer tissue (>90 patients), tumour-derived organoids, cancer cells and xenograft tumours with either long or short telomeres, we uncovered a heretofore undeciphered function of telomeres in modulating IL1 signalling and tumour immunity.

cancer biology↗

bioRxiv2021

SARS-CoV-2 Delta derivatives impact on neutralization of Covishield recipient sera

The emergence of SARS-CoV-2 Delta variant and its derivatives has created grave public health problem worldwide. The high transmissibility associated with this variant has led to daily increase in the number of SARS-CoV-2 infections. Delta variant has slowly dominated the other variants of concern. Subsequently, Delta has further mutated to Delta AY.1 to Delta AY.126. Of these, Delta AY.1 has been reported from several countries including India and considered to be highly infectious and probable escape mutant. Considering the possible immune escape, we had already evaluated the efficacy of the BBV152 against Delta and Delta AY.1 variants. Here, we have evaluated the neutralizing potential of sera of COVID-19 naive vaccinees (CNV) immunized with two doses of vaccine, COVID-19 recovered cases immunized with two doses of vaccine (CRV) and breakthrough infections (BTI) post immunization with two doses of vaccine against Delta, Delta AY.1 and B.1.617.3 using 50% plaque reduction neutralization test (PRNT50). Our study observed low NAb titer in CNV group against all the variants compared to CRV and BTI groups. Delta variant has shown highest reduction of 27.3-fold in NAb titer among CNV group compared to other groups and variants. Anti-S1-RBD IgG immune response among all the groups was also substantiated with NAb response. Compromised neutralization was observed against Delta and Delta AY.1 compared B.1 in all three groups. However, it provided protection against severity of the disease and fatality.

molecular biology↗