Biology subjects

Malvankar, S. R.

Publications and source records attributed to Malvankar, S. R..

3 recordsLinked to original sources

bioRxiv2026

Metformin Redirects Autophagy from Bulk Turnover to Mitochondrial Clearance

Metformin is the most widely prescribed antidiabetic drug and an active candidate for repurposing in oncology. How it engages autophagy - a pathway central to both its metabolic and its anti-tumor effects - has remained unresolved, with reports of induction, suppression, and no effect. Here we show that metformin reroutes rather than induces or inhibits autophagy in human cancer cells: at therapeutic concentrations, it suppresses bulk cytosolic turnover by selectively blocking WIPI2-mediated phagophore tethering, while the ULK1 initiation complex relocates toward mitochondria and engages selective mitochondrial clearance. We trace this redirection to mitochondrial complex I inhibition, registered as a shift in the NAD+/NADH ratio before any change in the adenylate pool, and to a non-canonical reprogramming of the ULK1 complex that operates independently of mTORC1 and of the proposed PEN2-lysosomal route. AMPK is engaged in a subunit-specific manner that restrains ATG13 at initiation and enables WIPI2 displacement at maturation. The ULK1 complex is therefore the node at which metformin sets autophagic substrate selection, with direct implications for combination therapy in diabetes and cancer.

cell biology↗

bioRxiv2025

γ-Secretase-mediated endoproteolysis of neuregulin-1 and E-cadherin

{gamma}-Secretase is an intramembrane protease complex, with nearly 150 substrates that are cleaved within their transmembrane domains (TMD). Amyloid Precursor Protein (APP) is the most widely studied, as processive proteolysis by {gamma}-secretase releases the amyloid-{beta}-peptide (A{beta}) implicated in the pathogenesis of Alzheimers disease. In contrast, proteolysis of other substrates has been little explored. The only known sequence specificity rule for {gamma}-secretase cleavage is for APP, in which phenylalanine is not tolerated at P2 with respect to any step in processive proteolysis. Recently, we found this specificity rule applies to the initial cleavage of Notch1 substrate as well. In this study, we examined the site of initial cleavage by {gamma}-secretase and explored the phenylalanine rule for two other {gamma}-secretase substrates: neuregulin1 (NRG1) and E-cadherin (CDH1). Upon incubation of recombinant substrates with purified protease complex, followed by mass spectroscopy (MS) and immunoblot analysis, initial cleavage products for NRG1 and CDH1 were identified. Two cleavage sites were observed in the NRG1 TMD, one of which matched that seen previously. However, the observed single CDH1 TMD cleavage site differed from the reported cytosolic cleavage site. Phenylalanine mutants of NRG1 and CDH1 in the P2 position relative to the first {gamma}-secretase cleavage site showed a shift in the cleavage site, along with reduction in total C-terminal and N-terminal products, compared to that seen with wild-type substrates. Taking together, these findings clarify the initial cleavage sites of NRG1 and CDH1 and support the intolerance of Phe at P2 position as a general rule for {gamma}-secretase substrates. For Table of Contents use only O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=109 SRC="FIGDIR/small/649652v1_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@1cfdda7org.highwire.dtl.DTLVardef@1e10813org.highwire.dtl.DTLVardef@d94ba6org.highwire.dtl.DTLVardef@1f58c08_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2023

Molecular Dynamics Activation of γ-Secretase for Cleavage of Notch1 Substrate

{gamma}-Secretase is an intramembrane aspartyl protease complex which cleaves the transmembrane domain of over 150 peptide substrates, including amyloid precursor protein (APP) and the Notch family of receptors, via two conserved aspartates D257 and D385 in the Presenilin-1 (PS1) catalytic subunit. However, while the activation of {gamma}-secretase for cleavage of APP has been widely studied, the cleavage of Notch by {gamma}-secretase remains poorly explored. Here, we combined Gaussian accelerated Molecular Dynamics (GaMD) simulations and mass spectrometry (MS) analysis of proteolytic products to present the first dynamic models for cleavage of Notch by {gamma}-secretase. MS showed that {gamma}-secretase cleaved the WT Notch at Notch residue G34, while cleavage of L36F mutant Notch occurred at Notch residue C33. Initially, we prepared our simulation systems starting from the cryoEM structure of Notch-bound {gamma}-secretase (PDB: 6IDF) and failed to capture the proper cleavages of WT and L36F Notch by {gamma}-secretase. We then discovered an incorrect registry of the Notch substrate in the PS1 active through alignment of the experimental structure of Notch-bound (PDB: 6IDF) and APP-bound {gamma}-secretase (PDB: 6IYC). Every residue of APP substrate was systematically mutated to the corresponding Notch residue to prepare a resolved model of Notch-bound {gamma}-secretase complexes. GaMD simulations of the resolved model successfully captured {gamma}-secretase activation for proper cleavages of both WT and L36F mutant Notch. Our findings here provided mechanistic insights into the structural dynamics and enzyme-substrate interactions required for {gamma}-secretase activation for cleavage of Notch and other substrates.

biophysics↗