Biology subjects

Ghosh, S. K.

Publications and source records attributed to Ghosh, S. K..

At least 19 recordsLinked to original sources

bioRxiv2026

Screening of Potential Vaccine Candidate and Sera Based Diagnostic Markers from N-linked Surface Glycoproteins of Entamoeba histolytica

Pathogenesis inflicted by Entamoeba histolytica causes amoebic diarrhea and liver abscesses and is one of the leading causes of mortality from parasitic disease worldwide. Preventive therapeutics in the form of a vaccine could be highly effective at providing umbrella protection for a vulnerable community. In this study, a group of ten putative hypothetical surface N-linked glycoproteins was examined to assess their potential as vaccine candidates and/or diagnostic markers against amoebic intestinal colitis and liver abscesses. To evaluate the immunogenicity of putative surface glycoproteins, we used Entamoeba histolytica-infected patients sera from Bangladesh and then assessed the titer of antibody these glycoproteins elicited in patients by enzyme-linked immunosorbent assay (ELISA) and immunoblot. Based on this study, eight of ten surface glycoproteins were found immunogenic, as specific antibodies against these glycoproteins were detected in patients sera. Three of the immunogenic glycoproteins showed strong IgG antibody responses in both patients with intestinal amoebiasis and those with liver abscess. On the other hand, the other two glycoproteins showed the presence of specific serum antibodies exclusively in patients with amoebic liver abscesses, not in individuals with amoebic colitis. The remaining two glycoproteins showed more specific sera antibodies against ALA, but the preference was not very distinct. Immunolocalization with a specific antibody against the most immunogenic glycoproteins further confirmed their presence in the cell membrane. This differential immunogenicity of these glycoproteins in the two groups of patients qualifies them to become prospective sera-based diagnostic markers and also has the potential to become vaccine candidates for amoebiasis protection.

immunology↗

bioRxiv2025

Scm3 interacts with the N-terminal tail of Cse4 to regulate kinetochore assembly in budding yeast

The kinetochore is a multiprotein complex formed at the centromeres and is essential for the faithful chromosome segregation. In most of the organisms the kinetochore is assembled on a specialized centromeric nucleosome where histone H3 is replaced by a variant, named CENP-A. In budding yeast, the CENP-A homolog, Cse4 is recruited to the centromeric nucleosome through an interaction between its C-terminal domain and a specific chaperone, named Scm3. Interestingly, following Cse4 recruitment during S phase, Scm3 persists at the centromeres throughout the cell cycle. Recent in vitro studies have reported that Scm3 also interacts with N-terminal of Cse4 (N-C) that in turn facilitates a better interaction of Ame1-Okp1 (AO) of COMA subcomplex with N-C, which promotes kinetochore assembly. In this work, using genetic and biochemical assays we provide in vivo evidence of the interaction between Scm3 and N-C. Additionally, by artificially tethering Scm3, we show that their association has the potential to stabilize a missegregating chromosome with inactive centromere, perhaps by forming an active kinetochore at an ectopic site. We propose that at the centromeres, Scm3 has two jobs in tandem - Cse4 deposition and stabilization of N-C, which together culminates into proper kinetochore assembly. This work has clinical significance as both CENP-A and its chaperone are known to be upregulated in human cells under disease states which can predispose the cells to aneuploidy due to increased chances of forming ectopic kinetochores.

cell biology↗

bioRxiv2025

CTG clade-specific proteins of the RSC chromatin remodeling complex regulate cell cycle progression of a critical priority fungal pathogen, Candida albicans

The RSC and the homologous chromatin remodeling complexes are known to regulate cell cycle progression in various organisms, including Saccharomyces cerevisiae, Drosophila, and Homo sapiens. In this work, we characterized the role of two novel CTG clade-specific proteins (Nri1 and Nri2) of the RSC complex in the regulation of cell cycle progression in a critical priority fungal pathogen, Candida albicans. We observed that Nri1, alone or along with Nri2, regulates cell cycle progression at multiple stages. The nri1{Delta}/{Delta} and nri1{Delta}/{Delta} nri2{Delta}/{Delta} mutants exhibited transient cell cycle arrest, defective spindle morphology, and cytokinesis. Transcriptomic analysis supported these mutant phenotypes and indicated a broad role of Nri proteins in the cell cycle. From our results, we conclude that Nri proteins are crucial for C. albicans proliferation and fitness. ImportanceThe composition of the essential RSC chromatin remodeling complex exhibits species-specific divergence, harboring unique subunits with distinct functions. In this study, we report that two fungal CTG clade-specific proteins of the C. albicans RSC complex, namely Nri1 and Nri2 can promote C. albicans fitness through regulating its cell cycle progression at multiple stages. Fitness defect along with stressor sensitivity and differential expression of the genes regulating pathogenesis in the nri mutants indicate potentiality of the Nri proteins as anti-Candida drug targets.

cell biology↗

bioRxiv2025

Yeast two-hybrid screening identifies Dad1 as a candidate substrate of Hrr25 kinase in Saccharomyces cerevisiae

In Saccharomyces cerevisiae, sister kinetochores are mono-oriented during meiosis I, ensuring that homologous chromosomes segregate to opposite poles, a process critically dependent on the kinase activity of Hrr25, a casein kinase. However, the direct substrates of Hrr25 involved in this mechanism remain poorly defined. In this study, we used a yeast two-hybrid (Y2H) approach to screen for and identify physical interactors of Hrr25. The HRR25 gene was cloned into a Y2H bait vector, and its functional expression was confirmed by complementation of a temperature-sensitive hrr25-ts mutant. Screening independent Y2H libraries in three reading frames, followed by validation via reporter assays, restriction analysis, and sequencing, we identified six putative interactors: HED1, DAD1, YDR015C (from clone C1-5), REP1 (C2-15), and CYR1 and SYS1 (C3-7). Phosphorylation site prediction and AlphaFold 3.0 structural modeling identified high-confidence Hrr25 target residues, including S70/T73 on Hed1 and S63 on Dad1, S323 on Rep1, and S198/S527 on Cyr1, each located in structurally accessible and potentially functional regions. Plasmid-dependent assays confirmed that reporter activation in C1-5 depended on the presence of the prey plasmid, and restriction mapping demonstrated that C1-5 contained a full-length DAD1 ORF. Given Dad1s known role in DASH/Dam1 kinetochore complex and its function in kinetochore-microtubule attachment, along with previous findings that DAD1 mutations cause meiosis I defects, our data suggest that Dad1 may be a substrate of Hrr25. We propose that Hrr25-mediated phosphorylation of Dad1 could facilitate sister kinetochore co-orientation during meiosis I. These results provide new insights into the molecular mechanisms of chromosome segregation and identify Dad1 as a potential candidate substrate for Hrr25 in meiotic regulation.

cell biology↗

bioRxiv2025

Biometric Aging Clock for Practical Utility in Clinical Settings

Aging clocks are an essential tool for biogerontological research that enables a quick assessment of ones pace of aging. In research settings, clocks trained on -omics data types are the most popular since they allow for an inspection of the most basic cellular processes of aging. In clinical settings, however, -omics biomarkers of aging are impractical since they are linked to extra logistical load and costs, and they require personnel training. In this article, we present a cost-efficient aging clock that can be implemented easily in most clinics and hospitals to measure patients aging rates. The clock requires only 22 biomarkers, including 17 blood test parameters, and four biometric measures (blood pressure, body mass index, and waist circumference). The clock predicts ones chronological age with a mean average error of 7.35 years, and it reveals associations with hypertension, cancer, and obesity.

bioinformatics↗

bioRxiv2024

Performance Evaluation of Ciprofloxacin loaded Micellar Bioconjugate Carrier for Eradication of Bacterial Biofilms

This study reports the utilization of a Curcumin-Exopolysaccharide (Cur-EPS) micellar bioconjugate comprising acid-labile ester bond as a carrier for ciprofloxacin (Cip) to improve its ability to eradicate mature bacterial biofilms. The most significant attributes of Cip-loaded Cur-EPS micelles, including hydrodynamic particle size, polydispersity index (PDI), and encapsulation efficiency (EE), were statistically optimized to be 305.43 {+/-} 1.47 nm, 0.38 {+/-} 1.23, and 84.50 {+/-} 2.01 %, respectively. At an acidic pH of 5.6, the pH-responsive release of Cip from Cur-EPS micelles was 82.51 {+/-} 2.95 % over a duration of 100 h, but it was reduced to 39.10 {+/-} 1.55 % at a pH of 7.4 over the same period. This pH-dependent Cip release may be attributed to the destabilisation of Cur-EPS micelles due to hydrolysis of the succinic acid linker in the acidic pH range. Thus, the acidic pH microenvironment of biofilms formed by pathogenic bacterial strains, such as methicillin resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa, enhances the pH-responsive release of Cip from Cur-EPS micelles, thereby leading to the eradication of mature biofilms. This enhanced antibiofilm effect of Cip-Cur-EPS, in contrast to the free form of Cip, was evident from the significant decrease in the biofilm biomass. Furthermore, mature biofilms that formed on the surface of the urinary catheter were almost eradicated when treated in vitro with Cip-Cur-EPS for 72 h in the presence of artificial urine media (AUM). Moreover, Cip-Cur-EPS could also eradicate the biofilms formed by P. aeruginosa and MRSA in a catheterized bladder model employing recirculated artificial urine. Overall, this study demonstrated the antibiofilm potential of Cip-Cur-EPS micelles, rendering them suitable therapeutic agents for addressing biofilm-associated infections in clinical settings. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=125 SRC="FIGDIR/small/630303v1_ufig1.gif" ALT="Figure 1"> View larger version (45K): org.highwire.dtl.DTLVardef@135f68org.highwire.dtl.DTLVardef@1dcd61forg.highwire.dtl.DTLVardef@16d9976org.highwire.dtl.DTLVardef@762425_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

bioRxiv2024

Evidence of centromeric histone 3 chaperone involved in DNA damage repair pathway

The centromeric protein-A (CENP-A) is an evolutionary conserved histone H3 variant that marks the identity of the centromeres. Several mechanisms regulate the centromeric deposition of CENP-A as its mislocalization causes erroneous chromosome segregation, leading to aneuploidy-based diseases, including cancers. The most crucial deposition factor is a CENP-A specific chaperone, HJURP (Scm3 in budding yeast), which specifically binds to CENP-A. However, the discovery of HJURP as a DDR (DNA damage repair) protein and evidence of its binding to Holliday junctions in vitro indicate a CENP-A-deposition-independent role of these chaperones. In this study, using budding yeast, we demonstrate that Scm3 is crucial for the DDR pathway as scm3 cells are sensitive to DNA damage. We further observe that the scm3 mutant interacts with the rad52 DDR mutant and is compromised in activating DDR-mediated arrest. We demonstrate that Scm3 associates with the DNA damage sites and undergoes posttranslational modifications upon DNA damage. Overall, from this report and earlier studies on HJURP, we conclude that DDR functions of CENP-A chaperones are conserved across eukaryotes. Thus, the revelation that these chaperones confer genome stability in more than one pathway has clinical significance.

cell biology↗

bioRxiv2023

Evidence of 14-3-3 proteins contributing to kinetochore integrity and chromosome congression during mitosis

The 14-3-3 family of proteins are conserved across eukaryotes and serve myriad important regulatory functions of the cell. Homo/heterodimers of these protein homologs, majorly recognize their ligands via conserved motifs from a plethora of cellular proteins to modulate the localization and functions of those effector ligands. In most of the genetic backgrounds of Saccharomyces cerevisiae, disruption of both 14-3-3 homologs (Bmh1 and Bmh2) are either lethal or survive with severe growth defects showing gross chromosomal missegregation and prolonged cell cycle arrest. To elucidate their contributions to chromosome segregation, in this work we investigated their centromere/kinetochore-related functions. Analysis of appropriate deletion mutants shows that Bmh isoforms have cumulative and unshared isoform-specific contributions in maintaining the proper integrity of the kinetochore ensemble. Consequently, bmh mutant cells exhibited perturbations in kinetochore-microtubule (KT-MT) dynamics, characterized by kinetochore declustering, mis-localization of kinetochore proteins, and Mad2-mediated transient G2/M arrest. These defects also caused an asynchronous chromosome congression in bmh mutants during metaphase. In summary, this report advances the knowledge on contributions of budding yeast 14-3-3 proteins in chromosome segregation by demonstrating their roles in kinetochore integrity and chromosome congression.

cell biology↗

bioRxiv2023

Higher meiotic chromosome condensation: a potential function of kinetochore through polo-like kinase

Chromosome condensation plays a pivotal role during faithful chromosome segregation, hence understanding the factors that drive condensation is crucial to get mechanistic insight into chromosome segregation. Previously we showed that in budding yeast, the absence of the non-essential kinetochore proteins affects chromatin-condensin association in meiosis but not in mitosis. A differential organization of the kinetochores, that we and others observed earlier during mitosis and meiosis may contribute to the meiotic-specific role. Here, with our in-depth investigation using in vivo chromosome condensation assays in cells sans a non-essential kinetochore protein, Ctf19, we establish that these proteins have roles in achieving a higher meiotic condensation without influencing much of the mitotic condensation. We further observed an accumulation of the polo-like kinase Cdc5 owing to its higher protein stability in ctf19{Delta} meiotic cells. High Cdc5 activity causes hyper-phosphorylation of the condensin resulting in its reduced stability and concomitant decreased association with the chromatin. Overall, our findings highlight the role of Ctf19 in promoting meiotic chromosome condensation by influencing the activity of Cdc5 and thereby affecting the stability and association of condensin with the chromatin.

cell biology↗

bioRxiv2023

The selfish yeast plasmid exploits a SWI/SNF-type chromatin remodeling complex for hitchhiking on chromosomes and ensuring high-fidelity propagation

Extra-chromosomal selfish DNA elements can evade the risk of being lost at every generation by behaving as chromosome appendages, thereby ensuring high fidelity segregation and stable persistence in host cell populations. The yeast 2-micron plasmid and episomes of the mammalian gammaherpes and papilloma viruses that tether to chromosomes and segregate by hitchhiking on them exemplify this strategy. We document for the first time the utilization of a SWI/SNF-type chromatin remodeling complex as a conduit for chromosome association by a selfish element. One principal mechanism for chromosome tethering by the 2-micron plasmid is the bridging interaction of the plasmid partitioning proteins (Rep1 and Rep2) with the yeast RSC2 complex and the plasmid partitioning locus STB. We substantiate this model by multiple lines of evidence derived from genomics, cell biology and interaction analyses. We describe a Rep-STB bypass system in which a plasmid engineered to non-covalently associate with the RSC complex mimics segregation by chromosome hitchhiking. Given the ubiquitous prevalence of SWI/SNF family chromatin remodeling complexes among eukaryotes, it is likely that the 2-micron plasmid paradigm or analogous ones will be encountered among other eukaryotic selfish elements.

molecular biology↗

bioRxiv2022

Evidence of kinesin motors involved in stable kinetochore assembly during early meiosis

The characteristic bi-lobed organization of the kinetochores observed during mitotic metaphase is a result of separation of the sister kinetochores into two clusters upon their stable end-on attachment to the microtubules emanating from opposite spindle poles. In contrast, during metaphase I of meiosis despite bi-orientation of the homologs, we observe that the kinetochores are linearly dispersed between the two spindle poles indicating that pole-distal and pole-proximal kinetochores are attached laterally and end-on, respectively to the microtubules. Colocalization studies of kinetochores and kinesin motors suggest that budding yeast kinesin 5, Cin8 and Kip1 perhaps localize to the end-on attached kinetochores while kinesin 8, Kip3 resides at all the kinetochores. Unlike mitosis in budding yeast, in meiosis, the outer kinetochores assemble much later after prophase I. From the findings including co-appearance of kinesin 5 and the outer kinetochore protein Ndc80 at the centromeres after prophase I and a reduction in Ndc80 level in Cin8 null mutant, we propose that kinesin motors are required for reassembly and stability of the kinetochores during early meiosis. Thus, this work reports yet another meiosis specific function of kinesin motor.

cell biology↗

bioRxiv2022

Template-Directed RIG-I Agonist Assembly for Targeted Cancer Immunotherapy

Recent developments in the use of pattern recognition receptors (PRRs) aim to harness the innate power of the immune system for cancer therapy. Understanding how to recruit PRRs, such as RIG-I, in a tumor-selective manner is critical for its adoption in the clinic. We describe the use of a tumor-selective template-based agonist of RIG-I to induce type-I IFN signaling and tumor cell apoptosis. The agonist, termed ss-ppp-miRNA-21, comprises a single stranded RNA oligonucleotide modified with a 5-triphosphate and complementary to an endogenous miRNA enriched in tumor cells. We demonstrate the efficacy of the template-directed approach and detail mechanistic studies validating the hypothesis of a template-directed RIG-I agonist assembly using miRNA-21 as a target. The template-directed strategy described here moves us closer to making RIG-I a clinically relevant target in oncology because it achieves targeted activation of innate immunity in the tumor microenvironment in the context of systemic agonist injection.

bioengineering↗

bioRxiv2022

Redefining the structure of tip-links in hair-cells

Tip-link, the gating-spring in hearing, modulates the mechanoelectrotransduction process and protects the hearing sensory machineries from overexcited auditory inputs. Tip-links are formed by two exceptionally long cadherins, cadherin-23 and protocadherin-15. It is proposed that both the cadherins first independently form cis-dimers through lateral interactions and finally, engage in trans-interactions to form the heterotetrameric assembly in tip-links. In fact, the molecular details of the protocadherin cis-dimers are well-understood. However, little structural and molecular details are known for independent cis-dimer of cadherin-23. Using photo-induced cross-linking of unmodified proteins in solution and on lipid-membranes, we observed no trace of cadherin-23 cis-dimers, rather we captured trans-homodimers of cadherin-23. We thus, redefine the tip-link structure where protocadherin-15 remains as cis-dimers, however, two individual cadherin-23 binds to protocadherin-15 cis-dimer for tip linking.

biochemistry↗

bioRxiv2022

Insights into the microevolution of SARS-ACE2 Interactions: In-silico analysis of glycosylation and SNP pattern

The prefatory protein-glycan interaction and stabilizing protein-protein interaction of severe acute respiratory syndrome viruses with angiotensin-converting enzyme 2 play a significant role in complex formation thereby promoting endocytosis. The microevolution of SARS-CoV-2 over a period of time has a significant role in increasing the affinity of receptor-binding domain against angiotensin converting-enzyme 2. In the current study, we have corroborated the vitality of acquired SNPs over a period of time with increased affinity by using docking studies. The results indicate that the virus modulates the undesirable glycosylation sites by a series of substitution and deletion mutations. It uses bulky residues such as Tyr/Phe for dynamic arrest for quick stabilization of the complex, and Lys residues for stabilizing via hydrogen bond formation besides increasing the binding affinity to ease the cell entry.

evolutionary biology↗

bioRxiv2021

The genome trilogy of Anopheles stephensi, an urban malaria vector, reveals structure of a locus associated with adaptation to environmental heterogeneity

BackgroundAnopheles stephensi is the most menacing malaria vector to watch for in newly urbanizing parts of the world. The fitness is reported to be a direct consequence of the vector adapting to laying eggs in over-head water tanks with street-side water puddles polluted by oil and sewage. Large frequent inversions of malaria vectors are implicated in adaptation. ResultsWe report the assembly of a strain of An. stephensi of the type-form, collected from a construction site from Chennai (IndCh) in 2016. The genome completes the trilogy with respect to a 16 Mbp inversion (2Rb) in An. stephensi associated with adaptation to environmental heterogeneity. Comparative genome analysis revealed breakpoint structure and allowed extraction of 22,650 segregating SNPs for typing this inversion. Using whole genome sequencing of 82 individual mosquitoes, we conclude that one third of both wild and laboratory populations maintain heterozygous genotype of 2Rb. The large number of SNPs are tailored to assign inversion genotype directly from 1740 exonic SNPs 80% of which are expressed in various developmental stages. ConclusionsThe genome trilogy approach accelerates study of fine structure and typing of important inversions in malaria vectors putting the genome resources for the much understudied An. stephensi, on par with the extensively studied malaria vector, Anopheles gambiae. We argue that the IndCh genome is relevant for field translation work compared to those reported earlier by showing that individuals from diverse populations cluster with IndCh pointing to significant commerce between cities, perhaps, allowing for survival of the fittest strain.

genomics↗

bioRxiv2021

Molecular dynamics simulations and functional studies reveal that hBD-2 binds SARS-CoV-2 spike RBD and blocks viral entry into ACE2 expressing cells

New approaches to complement vaccination are needed to combat the spread of SARS-CoV-2 and stop COVID-19 related deaths and long-term medical complications. Human beta defensin 2 (hBD-2) is a naturally occurring epithelial cell derived host defense peptide that has antiviral properties. Our comprehensive in-silico studies demonstrate that hBD-2 binds the site on the CoV-2-RBD that docks with the ACE2 receptor. Biophysical and biochemical assays confirm that hBD-2 indeed binds to the CoV-2-receptor binding domain (RBD) (KD [~] 300 nM), preventing it from binding to ACE2 expressing cells. Importantly, hBD-2 shows specificity by blocking CoV-2/spike pseudoviral infection, but not VSV-G mediated infection, of ACE2 expressing human cells with an IC50 of 2.4{+/-} 0.1 M. These promising findings offer opportunities to develop hBD-2 and/or its derivatives and mimetics to safely and effectively use as novel agents to prevent SARS-CoV-2 infection.

biochemistry↗

bioRxiv2020

Shugoshin promotes efficient activation of spindle assembly checkpoint and timely spindle disassembly

Shugoshin proteins are evolutionary conserved across eukaryotes with some species-specific cellular functions ensuring the fidelity of chromosome segregation. Shugoshin being present at various subcellular locales, acts as an adaptor to mediate various protein-protein interactions in a spatio-temporal manner. Here, we characterize shugoshin (Sgo1) in the human fungal pathogen, Candida albicans. Interestingly, we discover a novel in vivo localization of Sgo1 along the length of the mitotic spindle. Further, Sgo1 performs a hitherto unknown function of facilitating timely disassembly of spindle in this organism. We observe that Sgo1 retains its centromeric localization and performs its conserved functions that include regulating the centromeric condensin localization, chromosome passenger complex (CPC) maintenance and sister chromatid biorientation. We identify novel roles of Sgo1 as a spindle assembly checkpoint (SAC) component with functions in maintaining the SAC proteins, Mad2 and Bub1, at the kinetochores, in response to faulty kinetochore-microtubule attachments. These findings provide an excellent evidence of the functional rewiring of shugoshin in maintaining genomic stability.

cell biology↗

bioRxiv2020

Hitchhiking on condensed chromatin promotes plasmid persistence in yeast without perturbing chromosome function

Equipartitioning by chromosome hitchhiking and copy number correction by DNA amplification are at the heart of the evolutionary success of the selfish yeast 2-micron plasmid. The present analysis reveals plasmid presence near centromeres and telomeres in mitotic cells, with a preference towards the latter. The observed correlation of plasmid missegregation with non-disjunction of rDNA and telomeres under Cdc14 inactivation, higher plasmid missegregation upon induced missegregation of chromosome XII but not chromosome III, requirement of condensin for plasmid stability and the interaction of the condensin subunit Brn1 with the plasmid partitioning system lend functional credence to condensed chromatin being favored for plasmid tethering. By homing to condensed/quiescent chromosome locales, and not over-perturbing genome homeostasis, the plasmid may minimize fitness conflicts with its host. Analogous persistence strategies may be utilized by other extrachromosomal selfish genomes, for example, episomes of mammalian viruses that also hitchhike on host chromosomes for their stable maintenance.

cell biology↗