Biology subjects

Yadav, G. P.

Publications and source records attributed to Yadav, G. P..

3 recordsLinked to original sources

bioRxiv2025

Molecular mechanism by which SARS-CoV-2 Orf9b suppresses the Tom70-Hsp90 interaction to evade innate immunity

The Tom70-Hsp90 interaction is critical for activating MAVS-mediated interferon (IFN) production. Upon RNA virus infection, cytosolic Hsp90 recruits key innate immune signaling proteins to MAVS on mitochondria through its interaction with Tom70. To evade this innate immune response, the SARS-CoV-2 protein Orf9b binds to Tom70, thereby disrupting the Tom70-Hsp90 interaction and suppressing IFN production. Despite its importance, the molecular mechanism underlying Orf9b-mediated inhibition of IFN signaling remains unclear. Here, using an integrative approach combining cryo-electron microscopy, 19F NMR spectroscopy, and isothermal titration calorimetry (ITC), we show that Orf9b inhibits Hsp90 binding to Tom70 through a bipartite mechanism. The helix and intrinsically disordered tail of Orf9b sterically block the access of two distinct structural units of Hsp90 to Tom70. We also find that Orf9b-mediated allosteric conformational changes in Tom70 do not contribute to the inhibition of the Hsp90 binding. Comprehensive structural, thermodynamic, and kinetic analyses further reveal that Orf9b primarily slows the association kinetics between Hsp90 and Tom70. Collectively, our results provide a high-resolution mechanistic framework for understanding Orf9b-mediated suppression of the host innate immune response.

biophysics↗

bioRxiv2025

Harnessing Protein Unfolding for Thermosensing: Structural Insights from TRPV3

How proteins sense temperature with high precision is a fundamental biological challenge. Here, we structurally elucidate a dynamics-based mechanism for thermoTRPs, termed suicidal gating, in which extreme thermosensitivity arises from intrinsic instability and concerted protein dynamics, coupling channel opening to partial protein unfolding. Using cryo-EM, we directly capture heat-induced partial unfolding within the channels central pore domain, a structural core of gating. Analysis of mutant structures capturing intermediate activated states reveals no large domain shifts despite high temperature dependence, only subtle interfacial changes between domains. Both mutation and heat treatment consistently impact a critical network of thermolabile "latch" interactions along the S6-TRP helix-proximal N-terminus axis, driving receptor activation from a stable closed structure to progressively decoupled states and eventual disintegration. These findings provide direct structural evidence for a new receptor mechanism and establish a broader paradigm in which distributed structural flexibility, rather than localized dedicated sensors, drives extreme biological functions.

physiology↗

bioRxiv2024

Molecular requirements of chromogranin B for the long-sought anion shunter of regulated secretion

All eukaryotes utilize regulated secretion to release molecular signals packaged in secretory granules for local and remote signaling. An anion shunt conductance was first suggested in secretory granules of bovine chromaffin cells nearly five decades ago. Biochemical identity of this conductance remains undefined. CLC-3, an intracellular Cl-/H+ exchanger, was proposed as a candidate sixteen years ago, which, however, was contested experimentally. Here, we show that chromogranin B (CHGB) makes the kernel of the long-sought anion shunter in cultured and primary neuroendocrine cells and its channel functions are essential to proper granule maturation. Intragranular pH measurements and cargo maturation assays revealed that normal granular acidification, proinsulin-insulin conversion, and dopamine-loading in neuroendocrine cells all rely on functional CHGB+ channels. Primary {beta}-cells from Chgb-/- mice exhibited persistent granule deacidification, which suffices to uplift plasma proinsulin level, diminish glucose-induced 2nd-phase insulin secretion and dwindle monoamine content in chromaffin granules from the knockout mice. Data from targeted genetic manipulations, dominant negativity of a deletion mutant lacking channel-forming parts and tests of CLC-3/5 and ANO-1/2 all exclude CHGB-less channels from anion shunting in secretory granules. The highly conserved CHGB+ channels thus function in regulated secretory pathways in neuronal, endocrine, exocrine and stem cells of probably all vertebrates. HIGHLIGHTSO_LILoss of CHGB channel functions impairs secretory granule acidification in neuroendocrine cells, which necessitates anion shunt conduction. C_LIO_LICHGB{Delta}MIF, a mutant unable to form a functional Cl- channel, exerts negative dominance on endogenous CHGB and results in granule deacidification in cultured cells. C_LIO_LINeither CLC-3 & -5 nor ANO-1 & -2 participate in the CHGB-mediated granule acidification. Clcn3 knockout effects on regulated secretion can be attributed to its functions in endosomal and endolysosomal compartments. C_LIO_LIPrimary Chgb-/- {beta}-cells exhibit persistent granule deacidification, presenting a unifying mechanism for disparate mouse phenotypes: hyperproinsulinemia, near abrogation of 2nd phase insulin release after glucose challenge and diminution of monoamine contents in chromaffin granules. C_LI

cell biology↗