Biology subjects

Patra, D.

Publications and source records attributed to Patra, D..

12 recordsLinked to original sources

bioRxiv2026

Cryo-EM structures of the YsaN ATPase reveal dodecameric and tetradecameric assemblies in the Yersinia enterocolitica T3SS

Bacterial pathogens deploy sophisticated nanomachines such as the Type III Secretion System (T3SS) to inject effector proteins directly into host cells, thereby subverting cellular defenses and promoting infection. Central to this machinery is a conserved homo-oligomeric ATPase that fuels substrate unfolding and translocation. Despite extensive structural and biochemical studies, the precise mechanism of T3SS ATPase oligomer assembly and regulation remains incompletely understood, events crucial for the overall functionality of the T3SS. Here, we report high-resolution cryo-electron microscopy (cryo-EM) structures of the Yersinia enterocolitica T3SS ATPase YsaN, captured in two previously uncharacterized oligomeric states, a dodecamer (12-mer) at 3.51 [A] and a tetradecamer (14-mer) at 3.80 [A] resolution. Both assemblies exhibit helical-like symmetry and conformational heterogeneity. A conserved longitudinal fissure interrupts the helical lattice and appears to mediate interconversion between the two states, while an N-terminal interface stabilized by inter-tier disulfide bonds and an extensive non-covalent network locks the assembly together. Molecular dynamics simulations reveal that ATP hydrolysis drives a global, correlated compaction upon transition to the ADP-bound state. Together, these structural, biochemical, and MD simulation data establish T3SS ATPases as dynamic, oligomerically plastic molecular motors and provide a mechanistic framework for chaperone-effector dissociation and substrate translocation.

biophysics↗

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↗

bioRxiv2026

Fibroblast Growth Factor Receptor Signaling in Maturing Osteoblasts Controls Cell-Matrix Interactions Critical for Osteocyte Survival

The terminal differentiation of osteoblasts into osteocytes, the most abundant cell type in cortical bone, is critical for skeletal homeostasis. Osteocyte loss is a hallmark of bone aging and fragility, yet the mechanisms regulating osteocyte formation and survival are poorly understood. We show that inactivation of fibroblast growth factor receptor 1 (Fgfr1) in the mature osteoblast lineage results in extensive osteocyte death, identifying FGFR1 signaling as essential for osteocyte viability and bone integrity. Lineage tracing and analysis of endogenous and induced appositional bone formation revealed that newly embedded osteocytes fail to survive without FGFR1. These osteocytes exhibited ectopic expression of osteocalcin and podoplanin within sclerostin-positive, TUNEL-reactive lacunae, along with defective dendrite formation and disruption of the local lacunocanalicular network. RNA sequencing of cortical bone demonstrated reduced expression of extracellular matrix (ECM) genes and neuronal regulatory genes, while histological and ultrastructural analyses showed disorganized collagen fibrils, diminished osteoid, and abnormal mineralization. In vitro, FGF signaling in Ocy454 cells regulated gene programs involved in development, axon guidance, and bone ECM organization, highlighting a dual function for FGF signaling in which it controls both matrix-dependent and intrinsic cell differentiation mechanisms during the osteoblast-to-osteocyte transition. We propose that FGFR1 deficiency causes ECM disorganization and impaired dendrite formation, disrupting osteocyte communication with neighboring bone and vascular cells, ultimately leading to cell death. These findings establish FGFR signaling as a critical regulator of osteocyte differentiation, viability of bone-embedded osteocytes, and bone homeostasis. Summary StatementFGFR signaling has a profound effect on adult bone extracellular matrix that is vital to maintaining the viability and morphology of newly formed osteocytes, their lacunocanalicular network and the maintenance of bone homeostasis.

developmental biology↗

bioRxiv2025

Crowding drives terminal investment in a generalist pest

Apart from genotype and environment, several fitness related traits of individuals are also influenced by the conditions experienced by their parents, i.e., by parental effects. To uncover the underlying physiological or molecular mechanisms, most studies analyze specific drivers in isolation, such as parental age or the degree of crowding experienced by the parents. However, potential interaction effects between different aspects of parental context are poorly understood. We analyzed the combined effect of parental age and population density on the number, fitness and physiology of offspring, in the red flour beetle Tribolium castaneum. Both factors independently reduce fitness, leading us to predict that old parents from high density should produce fewer offspring of the lowest quality. Surprisingly, older parents exposed to crowding produced more eggs than expected. These eggs were larger, had higher survival in an optimal as well as a suboptimal resource, and were provisioned with more lipids. This result is consistent with terminal reproductive investment in response to high density. The interaction between two ecologically relevant contexts -- ageing and crowding -- thus drives terminal investment via nutrient provisioning, perhaps contributing to the success of this generalist pest.

evolutionary biology↗

bioRxiv2025

The mechanism of DRB7.2:DRB4 mediated sequestering of endogenous inverted-repeat dsRNA precursors in plants

Noncoding transcribable inverted repeat sequences are vital in genome stability, regulation of transposable elements, mutations, and diseases in eukaryotes. In vascular plants, dsRNA Binding Proteins (dsRBPs), DRB7.2 and DRB4, inhibit Dicer-like-protein 3 (DCL3) to stall endogenous inverted-repeat dsRNA (endo-IR dsRNA) mediated gene silencing. As dsRBPs generally assist Dicers, the inhibition of DCL3 by a dsRBP complex is quite enigmatic. Here, we explore how the DRB7.2:DRB4 complex sequesters substrate dsRNA of DCL3 using a structure-based mechanistic approach. Intriguingly, the crucial step of endo-IR dsRNA precursor sequestration is the high affinity complex formation of interacting domains of DRB7.2 (DRB7.2M) and DRB4 (DRB4D3). Next, we establish that DRB7.2 simultaneously interacts with DRB4 and endo-IR dsRNA precursors, where DRB4 contributes towards enhancement in the affinity of the complex with dsRNA, thereby impairing DCL3 mediated cleavage of endo-IR dsRNA precursors. The uniqueness of the DRB4D3 structure implies that the trans-acting (tasi)/siRNA initiation complex formed by DCL4:DRB4 in plants is diverse from its non-plant higher eukaryotes. Overall, we present considerable insights into endo-IR dsRNA precursors regulation in plants and indicate a differential evolution of RNAi initiation complexes between plants and other higher eukaryotes.

biophysics↗

bioRxiv2024

Resolving the design principles that control postnatal vascular growth and scaling

After birth, tissues grow continuously until reaching adult size, with each organ exhibiting unique cellular dynamics, growth patterns, and (stem or non-stem) cell sources. Using a suite of experimental and computational multiscale approaches, we found that aortic expansion is guided by specific biological principles and scales with the vertebral column rather than animal body weight. Expansion proceeds via two distinct waves of arterial cell proliferation along blood flow that are spatially stochastic, yet temporally coordinated. Each wave exhibits unique cell cycle kinetics and properties, with the first wave exhibiting cell cycle durations as fast as 6 hours. Single-cell RNA sequencing showed changes in fatty acid metabolism concomitant with an increase in cell size. Mathematical modeling and experiments indicated endothelial cell extrusion is essential for homeostatic aortic growth and balancing excess proliferation. In a genetic model of achondroplasia, the aorta achieves proper scaling through enhanced cell extrusion while maintaining normal proliferation dynamics. Collectively, these results provide a blueprint of the principles that orchestrate aortic growth which depends entirely on differentiated cell proliferation rather than resident stem cells.

developmental biology↗

bioRxiv2023

Identification of a myofibroblast differentiation program during neonatal lung development

Alveologenesis is the final stage of lung development in which the internal surface area of the lung is increased to facilitate efficient gas exchange in the mature organism. The first phase of alveologenesis involves the formation of septal ridges (secondary septae) and the second phase involves thinning of the alveolar septa. Within secondary septa, mesenchymal cells include a transient population of alveolar myofibroblasts (MyoFB) and a stable but poorly described population of lipid rich cells that have been referred to as lipofibroblasts or matrix fibroblasts (MatFB). Using a unique Fgf18CreER lineage trace mouse line, cell sorting, single cell RNA sequencing, and primary cell culture, we have identified multiple subtypes of mesenchymal cells in the neonatal lung, including an immature progenitor cell that gives rise to mature MyoFB. We also show that the endogenous and targeted ROSA26 locus serves as a sensitive reporter for MyoFB maturation. These studies identify a myofibroblast differentiation program that is distinct form other mesenchymal cells types and increases the known repertoire of mesenchymal cell types in the neonatal lung. Summary StatementDuring primary alveologenesis, alveolar myofibroblasts comprise a distinct proliferative mesenchymal lineage that matures and populates emerging secondary septa.

developmental biology↗

bioRxiv2022

Targeted Deletion of Fgf9 in Tendon Disrupts Mineralization of the Developing Enthesis

The enthesis is a transitional tissue between tendon and bone that matures postnatally. The development and maturation of the enthesis involve cellular processes likened to an arrested growth plate. In this study, we explored the role of fibroblast growth factor 9 (Fgf9), a known regulator of chondrogenesis and vascularization during bone development, on the structure and function of the postnatal enthesis. First, we confirmed spatial expression of Fgf9 in wildtype tendon and enthesis using in situ hybridization. We then used Cre recombinase driven by the scleraxis promoter (ScxCre) to conditionally inactivate Fgf9 in mouse tendon and enthesis. Characterization of enthesis morphology and mechanical properties in Fgf9ScxCre and wildtype (WT) entheses showed a smaller calcaneal and humeral apophyses, thinner cortical bone at the attachment, increased cellularity, and reduced failure load in mature entheses in Fgf9ScxCre compared to WT littermates. During postnatal development, we found reduced chondrocyte hypertrophy and disrupted type X collagen (Col X) in Fgf9ScxCre entheses. These findings support a model in which tendon-derived Fgf9 regulates the functional development of the enthesis, including its postnatal mineralization.

developmental biology↗

bioRxiv2022

BRCA1 Hypermethylation In Sporadic Breast Cancers: Discovering A Novel Pathway To Tumorigenesis Via Coordinate NBR2 Deregulation And TNBC Transformation

Women with a family history of mutations in the Breast cancer susceptibility gene, BRCA1 will have an increased risk of developing breast neoplasms. However, majority of the breast cancers are sporadic where BRCA1 mutations are very rare. Instead, 5-65% of sporadic cases manifest BRCA1 promoter hypermethylation and 30-40% of such cases develop into Triple Negative Breast Cancers. Even then, the molecular mechanism of BRCA1 hypermethylation mediated breast tumorigenesis has remained an enigma till date. Here, we present a novel tumorigenesis pathway for breast cancers that engenders from BRCA1 hypermethylation by generating site-specific methylations in the BRCA1 promoter using a modified version of CRISPR technology. We report that induction of site-specific methylation on BRCA1 promoter effectuates a downregulation in BRCA1 expression via alteration in the balance between its alternate transcripts {beta} and . Induced BRCA1 hypermethylation is also responsible for the attenuation of a long noncoding RNA, NBR2 (Neighbour of BRCA1 gene 2), which is transcribed through the bidirectional BRCA1 promoter in the reverse direction. Downregulation of NBR2 activates a feedback loop by leading to further downregulation of BRCA1 which is more evident under glucose starvation conditions and is associated with impaired DNA damage repair. BRCA1 hypermethylation also results in significant overexpression of {beta}-hCG (human chorionic gonadotrophin), which was found to be associated with highly aggressive and drug-resistant forms of BRCA1 mutated breast cancers invitro & in vivo in our previous study. Further, we report a change in the hormone receptor levels as the tumor progresses which demonstrates how BRCA1 deficient cells modulate their expression of ER- and ER-{beta} to promote their proliferation in early stages of tumor development and at later stages, transform to a basal tumor subtype by shedding down the expression of ER- & PR. Interestingly, we also discovered that modulation of ER- expression upon BRCA1 hypermethylation is responsible for the alteration in BRCA1 transcript ratio. Finally, in in vivo mouse studies, BRCA1 hypermethylated tumors were found to be much larger, aggressive and invasive as compared to wildtype, BRCA1 and NBR2 knockdown tumors with downregulation of ER- and PR; which explains the most probable reason behind high relapse rates in BRCA1 hypermethylated tumors. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=118 SRC="FIGDIR/small/490082v2_ufig1.gif" ALT="Figure 1"> View larger version (19K): org.highwire.dtl.DTLVardef@1c81155org.highwire.dtl.DTLVardef@1ea1fdcorg.highwire.dtl.DTLVardef@1d1b5e9org.highwire.dtl.DTLVardef@fddd22_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2021

Joint degeneration in a mouse model of pseudoachondroplasia: ER stress, inflammation and autophagy blockage

Pseudoachondroplasia (PSACH), a short limb skeletal dysplasia, associated with premature joint degeneration is caused by misfolding mutations in cartilage oligomeric matrix protein (COMP). Here, we define mutant-COMP-induced stress mechanisms that occur in articular chondrocytes of MT-COMP mice, a murine model of PSACH. The accumulation of mutant-COMP in the ER occurred early in MT-COMP articular chondrocytes and stimulated inflammation (TNF) at 4 wks. Articular chondrocyte death increased at 8 wks and ER stress through CHOP was elevated by 12 wks. Importantly, blockage of autophagy (pS6), the major mechanism which clears the ER, sustained cellular stress in MT-COMP articular chondrocytes. Degeneration of MT-COMP articular cartilage was similar to that observed in PSACH and was associated with increased MMPs, degradative enzymes. Moreover, chronic cellular stresses stimulated senescence. Senescence-associated secretory phenotype (SASP) may play a role in generating and propagating a pro-degradative environment in the MT-COMP murine joint. The loss of CHOP or resveratrol treatment from birth preserved joint health in MT-COMP mice. Taken together, these results indicate that ER stress/CHOP signaling and autophagy blockage are central to mutant-COMP joint degeneration and MT-COMP mice joint health can be preserved by decreasing articular chondrocyte stress. Future joint sparing therapeutics for PSACH may include resveratrol.

pathology↗

bioRxiv2020

SARS-CoV-2 viral budding and entry can be modeled using virus-like particles

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was first discovered in December 2019 in Wuhan, China and expeditiously spread across the globe causing a global pandemic. While a select agent designation has not been made for SARS-CoV-2, closely related SARS-CoV-1 and MERS coronaviruses are classified as Risk Group 3 select agents, which restricts use of the live viruses to BSL-3 facilities. Such BSL-3 classification make SARS-CoV-2 research inaccessible to the majority of functioning research laboratories in the US; this becomes problematic when the collective scientific effort needs to be focused on such in the face of a pandemic. In this work, we assessed the four structural proteins from SARS-CoV-2 for their ability to form viruslike particles (VLPs) from human cells to form a competent system for BSL-2 studies of SARS-CoV-2. Herein, we provide methods and resources of producing, purifying, fluorescently and APEX2-labeling of SARS-CoV-2 VLPs for the evaluation of mechanisms of viral budding and entry as well as assessment of drug inhibitors under BSL-2 conditions.

microbiology↗

bioRxiv2019

Targeting the trypanosome kinetochore with CLK1 protein kinase inhibitors

The kinetochore is a macromolecular structure that assembles on the centromeres of chromosomes and provides the major attachment point for spindle microtubules during mitosis. In Trypanosoma brucei the proteins that make up the kinetochore are highly divergent, with the inner kinetochore comprising at least 20 distinct and essential proteins (KKT1-20) that include four protein kinases, CLK1 (KKT10), CLK2 (KKT19), KKT2 and KKT3. We performed a phenotypic screen of T. brucei bloodstream forms with a Novartis kinase-focused inhibitor library, which identified a number of selective inhibitors with potent pan-kinetoplastid activity. Deconvolution of an amidobenzimidazole series using a selection of 37 T. brucei mutants that over-express known essential protein kinases identified CLK1 as the primary target. Biochemical studies show that the irreversible competitive inhibition of CLK1 is dependent on a Michael acceptor forming an irreversible bond with C215 in the ATP binding pocket, a residue that is not present in human CLK1, thereby providing selectivity. Chemical inhibition of CLK1 impairs inner kinetochore recruitment and compromises cell cycle progression, leading to cell death. We show that KKT2 is a substrate for CLK1 and identify phosphorylation of S508 to be essential for KKT2 function and for kinetochore assembly. We propose that CLK1 is part of a novel signalling cascade that controls kinetochore function via phosphorylation of the inner kinetochore protein kinase KKT2. This work highlights a novel drug target for trypanosomatid parasitic protozoa and a new chemical tool for investigating the function of their divergent kinetochores.

microbiology↗