Mesquita, F. S. | Search bioRxiv

bioRxiv2021

S-acylation controls SARS-Cov-2 membrane lipid organization and enhances infectivity

SARS-CoV-2 virions are surrounded by a lipid bilayer which contains membrane proteins such as Spike, responsible for target-cell binding and virus fusion, the envelope protein E and the accessory protein Orf3a. Here, we show that during SARS-CoV-2 infection, all three proteins become lipid modified, through action of the S-acyltransferase ZDHHC20. Particularly striking is the rapid acylation of Spike on 10 cytosolic cysteines within the ER and Golgi. Using a combination of computational, lipidomics and biochemical approaches, we show that this massive lipidation controls Spike biogenesis and degradation, and drives the formation of localized ordered cholesterol and sphingolipid rich lipid nanodomains, in the early Golgi where viral budding occurs. ZDHHC20-mediated acylation allows the formation of viruses with enhanced fusion capacity and overall infectivity. Our study points towards S-acylating enzymes and lipid biosynthesis enzymes as novel therapeutic anti-viral targets.

cell biology↗

bioRxiv2020

Src-dependent NM2A tyrosine-phosphorylation regulates actomyosin dynamics

Non-muscle myosin 2A (NM2A) is a key cytoskeletal enzyme that along with actin assembles into actomyosin filaments inside cells. NM2A is fundamental in cellular processes requiring force generation such as cell adhesion, motility and cell division, and plays important functions in different stages of development and during the progression of viral and bacterial infections. We previously identified a novel tyrosine phosphorylation on residue 158 (pTyr158) in the motor domain of NM2A. This phosphorylation is dependent on Src kinase and is promoted by Listeria monocytogenes infection of epithelial cells, however its role is unknown. Here we show that Listeriolysin O (LLO), the pore-forming toxin (PFT) secreted by L. monocytogenes, is sufficient to trigger NM2A pTyr158 by activating Src, an upstream regulator of actomyosin remodeling. We further address the role of NM2A pTyr158 on the organization and dynamics of the actomyosin cytoskeleton and find that, by controlling the activation of the NM2A, the status of the pTyr158 alters cytoskeletal organization, dynamics of focal adhesions and cell motility. In vitro, we observe that non-phosphorylatable and phospho-mimetic versions of NM2A at Tyr158 display motor and ATPase activities similar to the wild-type NM2A, which indicates that the phenotype of these mutants in cells is independent of their ability to translocate actin filaments. Importantly, we find the regulation of this phosphorylation site to be of physiological relevance in Caenorhabditis elegans, in particular in response to intoxication by a PFT and to heat shock. We conclude that the control of the phosphorylation status at NM2A Tyr158 is a conserved trait that contributes to the regulation of actomyosin dynamics and the ability of cells to respond to bacterial infection. We propose Src-dependent NM2A pTyr158 as a novel layer of regulation of the actomyosin cytoskeleton.

cell biology↗