Biology subjects

Dave, P.

Publications and source records attributed to Dave, P..

5 recordsLinked to original sources

bioRxiv2026

A broad-spectrum phage-encoded mechanism to disarm bacterial type IV filaments

Phages can modify host cell physiology to thwart competitors. The Pseudomonas aeruginosa-specific phage DMS3 encodes Aqs1, a protein inhibitor of type IV pilus (T4P) function to prevent host cell recognition by other phages that leverage these filaments for infection. Aqs1 disrupts T4P by binding to the hexameric ATPase PilB, required to power pilus filament extension, though several mechanistic details remain unclear. We show that Aqs1 has broad-spectrum activity and can disrupt T4P function in a variety of Gram negative bacteria. This protein inhibits PilB by binding to a solvent-exposed hydrophobic patch on the N2-domain, distal to the active site. Binding destabilizes the hexamer, preventing PilB accumulation at T4P machines. Aqs1 likely disrupts PilB oligomerization by displacing a flexible linker segment between the PilB N1- and N2-domains required for inter-subunit contact. Together, the Aqs1 mode of action provides a design template for broad-spectrum inhibitors of diverse bacterial virulence factors. SignificanceThe phage-encoded protein Aqs1 disables type IV pilus (T4P) production in Pseudomonas aeruginosa by targeting the hexameric ATPase responsible for assembling pilus fibers. We show that despite originating from a P. aeruginosa-specific phage, Aqs1 can also disable T4 ATPase-dependent phenotypes across other pathogenic bacteria and homologous systems. Mechanistically, Aqs1 binds to a conserved patch on the PilB N2-domain away from the active site. Binding here breaks apart the PilB oligomer, preventing it from acting on T4P machines. Aqs1 binding at the N2-domain patch likely displaces a flexible PilB linker segment that binds to this site to stabilize the hexamer. Our work highlights a novel and conserved PilB allosteric site which is exploited by the phage-encoded protein Aqs1 to disrupt diverse T4 systems in multiple bacteria.

microbiology↗

bioRxiv2025

Stress granules formed during different RNA virus infections show remarkable plasticity and substantial virus-specific differences in their formation and composition

Eukaryotic cells evolved a cellular stress response to cope with extrinsic and intrinsic stress stimuli including virus infections. The major result of this response is the shutdown of bulk translation to prevent damage and allow the reprogramming of translation towards stress-resolving pathways. The resulting translationally stalled mRNA and associated proteins are accumulated in membrane-less cytosolic condensates called stress granules (SG). While the inhibitory effect of translation arrest on virus growth is well established, the role of SGs in the cellular defense against viruses is still unclear. The observation of specific interference with SG formation during various virus infections led to the hypothesis that SGs could serve as antiviral signaling platforms. In this study, we used mouse hepatitis virus (MHV) to characterize SGs formed during coronavirus infection. By applying APEX2-mediated proximity labelling in combination with quantitative proteomics, we dissected the proteome of MHV-induced granules and compared it to canonical SGs formed during oxidative stress. Our data revealed substantial differences in protein abundance and composition indicating stressor-specific SG characteristics. To assess if the observed differences are a general feature of virus-induced SGs or rather virus-specific, we extended our investigations to the Semliki Forest virus (SFV), a member of the alphavirus family known to induce SGs. An initial comparison of SG formation kinetics by live-cell imaging showed distinct time points of SG induction between both viruses. A comprehensive comparison of the SG protein compositions revealed profound differences in the SG proteome between SFV and MHV. A further subcellular localization of SG components by microscopy not only confirmed a reduced abundance of several translation initiation factors in MHV-induced granules, but surprisingly, revealed the presence of SFV RNA and the absence of MHV RNA in virus-induce SGs. The reduced connection to canonical SG themes observed for MHV-induced granules compared to SFV- and oxidative stress-induced ones indicates a different impact of these condensates on MHV replication and further raises the question whether they should be considered SGs. The surprising plasticity of SGs concerning induction kinetics, protein composition and abundance, and inclusion or exclusion of viral RNA provide a base for future investigations of the role(s) of SGs in the context of viral infection and how they may impact virus replication.

microbiology↗

bioRxiv2024

Subendothelial Calcification Increases Surface Roughness and Promotes Lipid Deposition into the Arterial Wall

ObjectiveWe have previously demonstrated that subendothelial calcification accelerates atherosclerosis in mice. This study addresses a mechanism by which subendothelial calcifications can increase low-density lipoprotein (LDL) uptake into the arterial wall. MethodsMice overexpressing tissue-nonspecific alkaline phosphatase (TNAP) in endothelial cells (eTNAP mice) were used as a model of calcification. Calcification and atherosclerosis were detected by micro-computed tomography (micro-CT) and histology. The endothelial roughness was characterized by surface metrology. A fluid-structure interaction model was used to calculate wall shear stress (WSS). The uptake of fluorescent LDL was traced in vitro and in vivo. Human arteries were assessed for the prevalence of internal elastic lamina (IEL) calcification. ResultseTNAP mice developed more severe aortic atherosclerosis than controls on the LDL receptor mutant background (p<0.01). Subendothelial calcifications in eTNAP mice were confirmed by micro-CT. An increase in aortic surface area roughness, including the height, volume, and steepness parameters, was observed in eTNAP mice compared to controls (p<0.01). Calcifications affected near-wall hemodynamics, creating pockets of reduced WSS. Endothelial cells cultured on rough surfaces showed increased LDL uptake compared to cells cultured on smooth collagen (p<0.0001). Fluorescent LDLs were traced to subendothelial calcifications in eTNAP mice but not in controls. In humans, IEL calcification was prevalent in older adults and inversely correlated with arterial diameter (p<0.05). ConclusionSubendothelial calcification is sufficient to perturb near-wall hemodynamics, creating localized areas of reduced WSS, consistent with increased LDL uptake near calcified lesions. Subendothelial calcification may represent an alternative or concurrent mechanism for the initiation of atherosclerosis. Research perspectiveO_ST_ABSWhat is new?C_ST_ABSO_LIWe tested a novel hypothesis that subendothelial microcalcification can initiate atherosclerosis. C_LIO_LIThe study demonstrated that micron-sized subendothelial calcifications, induced by the overexpression of tissue-nonspecific alkaline phosphatase in the endothelium, are sufficient to perturb local hemodynamics, creating pockets of low wall shear stress, consistent with an increase in low-density lipoprotein uptake and deposition into the arterial wall in juxtaposition to calcified lesions. C_LI What are the clinical implications?O_LIWe suggest that calcification of internal elastic lamina of medium-sized arteries may represent early lesions initiating atherosclerosis; however, the epidemiologic evidence for this theory is currently unavailable. C_LI

physiology↗

bioRxiv2024

Distinct phenotypic consequences of cholangiocarcinoma-associated FGFR2 alterations depend on biliary epithelial maturity

Epithelial cancers disrupt tissue architecture and are often driven by mutations in genes that play important roles in normal epithelial morphogenesis. The intrahepatic biliary system is an epithelial tubular network that forms within the developing liver via the de novo initiation and expansion of apical lumens. Intrahepatic biliary tumors (intrahepatic cholangiocarcinoma) commonly harbor activating genomic alterations in the FGFR2 receptor tyrosine kinase, which plays important roles in epithelial morphogenesis in other developmental settings. Using a physiologic and quantitative 3D model we demonstrate that FGFR signaling is important for biliary morphogenesis and that oncogenic FGFR2 fusions and in-frame deletions disrupt biliary architecture. Importantly, we show that the trafficking of and signaling from the FGFR2 mutants, as well as their phenotypic impacts, are governed by the epithelial state of the cell. Unexpectedly, we also found that distinct tumor-driving FGFR2 mutants disrupt biliary morphogenesis in completely different and clinically relevant ways, informing our understanding of morphogenesis and tumorigenesis and highlighting the importance of convergent studies of both.

cancer biology↗

bioRxiv2021

Single-molecule imaging reveals the coupling of translation and mRNA decay

The relationship between mRNA translation and decay is incompletely understood, with conflicting reports suggesting that translation can either promote decay or stabilize mRNAs. The effect of translation on mRNA decay has mainly been studied using ensemble measurements and global inhibitors of transcription and translation, which can mask the underlying mechanisms. We developed a single-molecule imaging approach to control the translation of a specific transcript that enabled simultaneous measurement of translation and mRNA decay. Our results demonstrate that mRNAs undergoing translation are degraded faster than non-translating ones, although with slower kinetics than translation-coupled degradation of transcripts targeted by NMD. Furthermore, our results indicate that miRNAs mediate efficient degradation of both translating and non-translating target mRNAs. Single-molecule measurements of translation and decay reveal a predominant role of mRNA decay in miRNA-mediated regulation. Simultaneous visualization of translation and decay on single mRNAs provides a framework to study how these processes are interconnected in cells.

molecular biology↗