Biology subjects

Prithviraj, M.

Publications and source records attributed to Prithviraj, M..

3 recordsLinked to original sources

bioRxiv2026

The phage shock protein A (PspA) maintains membrane potential and supports NADH dehydrogenase function in mycobacteria

Maintenance of membrane integrity and proton motive force (PMF) is critical for bacterial survival. The phage shock protein (Psp) system, conserved across bacterial species, stabilizes the membrane, maintains PMF, and protects against envelope damage. However, how the conserved effector PspA contributes to PMF maintenance remains unclear. Here, using the mycobacterial Psp system as a genetically tractable model, we provide mechanistic insight into this process. We show that PspA and the accessory protein PspM jointly sustain membrane potential, with PspM required to maintain a ~70 kDa PspA isoform at the membrane during envelope stress. Loss of PspA increases susceptibility to thioridazine, which targets type II NADH dehydrogenase (NDH-2), and to Ro 48-8071, an inhibitor of menaquinone biosynthesis. Notably, hypersusceptibility to thioridazine is rescued by exogenous menaquinone. Consistent with these phenotypes, a pspA-deficient mutant exhibits impaired NADH dehydrogenase activity despite unchanged abundance of NDH-2 and menaquinone (MK-9). Together, these findings identify a functional link between PspA and NADH dehydrogenase-dependent respiration and suggest that PspA contributes to PMF maintenance by supporting respiratory electron transfer.

molecular biology↗

bioRxiv2022

Genetic and lipidomic identification of tuberculostearic acid as a controller of mycobacterial membrane compartmentalization

Mycobacteria diverge in a basic way from other bacterial and eukaryotic cells based on their distinct membrane structures. Here we report genome-wide transposon sequencing to discover the controllers of membrane compartmentalization in Mycobacterium smegmatis. cfa, a gene that encodes a putative cyclopropane-fatty-acyl-phospholipid synthase, shows the most significant effect on recovery from a membrane destabilizer, dibucaine. Lipidomic analysis of cfa deletion mutants demonstrates an essential role of Cfa in the synthesis of specific membrane lipids containing a C19:0 monomethyl-branched stearic acid. This molecule, also known as tuberculostearic acid (TBSA), has been intensively studied for decades due to its high level and genus-specific expression in mycobacteria. The proposed Cfa-mediated conversion of an unsaturation to a methylation matched well with its proposed role in lateral membrane organization, so we used new tools to determine the non-redundant effects of Cfa and TBSA in mycobacterial cells. cfa expression regulated major classes of membrane lipids including phosphatidylinositols, phosphatidylethanolamines and phosphatidylinositol mannosides. Cfa localized within the intracellular membrane domain (IMD), where it controls both cellular growth and recovery from membrane fluidization by facilitating subpolar localization of the IMD. Overall, cfa controls lateral membrane partitioning but does not detectably alter orthogonal transmembrane permeability. More generally, these results support the proposed role of the subpolar IMD as a subcellular site of mycobacterial control of membrane function. SignificanceMycobacteria remain major causes of disease worldwide based in part on their unusual membrane structures, which interface with the host. Here we discover the long sought biosynthetic origin of tuberculostearic acid (TBSA), a major fatty acid found selectively in mycobacteria, as well as its role in mycobacterial cells. The lipid is produced by an enzyme called Cfa, whose loss causes a growth defect and slow reformation of a membrane domain near the pole of the rod-shaped cell. Thus, our study offers mechanistic insights to the intrinsic molecular factors critical for mycobacterial plasma membrane partitioning.

microbiology↗

bioRxiv2022

Inositol acylation of phosphatidylinositol mannosides: a rapid mass response to membrane fluidization in mycobacteria

Mycobacteria share an unusually complex, multilayered cell envelope, which contributes to adaptation to changing environments. The plasma membrane is the deepest layer of the cell envelope and acts as the final permeability barrier against outside molecules. There is an obvious need to maintain the plasma membrane integrity, but the adaptive responses of plasma membrane to stress exposure remain poorly understood. Using chemical treatment and heat stress to fluidize the membrane, we show here that phosphatidylinositol (PI)-anchored plasma membrane glycolipids known as PI mannosides (PIMs) rapidly remodel their structures upon membrane fluidization in Mycobacterium smegmatis. Without membrane stress, PIMs are predominantly in a tri-acylated form: two acyl chains of PI moiety plus one acyl chain modified at one of the mannose residues. Upon membrane fluidization, the fourth fatty acid is added to the inositol moiety of PIMs, making them tetra-acylated variants. PIM inositol acylation is a rapid response independent of de novo protein synthesis, representing one of the fastest mass conversions of lipid molecules found in nature. Strikingly, we found that M. smegmatis is more resistant to the bactericidal effect of a cationic detergent after benzyl alcohol preexposure. We further demonstrate that fluidization-induced PIM inositol acylation is conserved in pathogens such as Mycobacterium tuberculosis and Mycobacterium abscessus. Our results demonstrate that mycobacteria possess a mechanism to sense plasma membrane fluidity change. We suggest that inositol acylation of PIMs is a novel membrane stress response that enables mycobacterial cells to resist membrane fluidization.

biochemistry↗