Spatially distinct and metabolically active membrane domain in mycobacteria

Abstract

Protected from host immune attack and antibiotic penetration by their unique cell envelope, mycobacterial pathogens cause devastating human diseases such as tuberculosis. Seamless coordination of cell growth with cell envelope elongation at the pole maintains this barrier. Unraveling this spatiotemporal regulation is a potential strategy for controlling mycobacterial infections. Our biochemical analysis previously revealed two functionally distinct membrane fractions in Mycobacterium smegmatis cell lysates: plasma membrane tightly associated with the cell wall (PM-CW) and a distinct fraction of pure membrane free of cell wall components (PMf). To provide further insight into the functions of these membrane fractions, we took the approach of comparative proteomics and identified more than 300 proteins specifically associated with the PMf, including essential enzymes involved in cell envelope synthesis such as a mannosyltransferase, Ppm1, and a galactosyltransferase, GlfT2. Furthermore, comparative lipidomics revealed the distinct lipid composition of the PMf, with specific association of key cell envelope biosynthetic precursors. Live-imaging fluorescence microscopy visualized the PMf as patches of membrane spatially distinct from the PM-CW and notably enriched in the pole of the growing cells. Taken together, our study provides the basis for assigning the PMf as a spatiotemporally distinct and metabolically active membrane domain involved in cell envelope biogenesis.

DivIVA concentrates mycobacterial cell envelope assembly for initiation and stabilization of polar growth

“…In support, the cytoplasmic enzymes that mediate arabinogalactan and mycomembrane synthesis are enriched at the mycobacterial cell poles (Carel et al, 2014;Hayashi et al, 2016;Meniche et al, 2014), as is metabolic labeling by trehalose monomycolate precursors that incorporate into the mycomembrane (Foley et al, 2016). In support, the cytoplasmic enzymes that mediate arabinogalactan and mycomembrane synthesis are enriched at the mycobacterial cell poles (Carel et al, 2014;Hayashi et al, 2016;Meniche et al, 2014), as is metabolic labeling by trehalose monomycolate precursors that incorporate into the mycomembrane (Foley et al, 2016).…”

A Screen for Protein–Protein Interactions in Live Mycobacteria Reveals a Functional Link between the Virulence-Associated Lipid Transporter LprG and the Mycolyltransferase Antigen 85A

“…Cell growth and lipid biosynthesis have been previously linked via the physical association of Wag31 with fatty acid and lipid biosynthetic enzymes (FASII, AccA3, AccD5), and the inner membrane mycolate transporter MmpL3 20, 22, 45 . Also, enzymes in the phosphatidyl inositol mannoside lipid and phosphatidic acid biosynthetic pathways co-localize at the cell poles, pointing to analogous coordination of inner membrane synthesis 46 . Our results with LprG support these models and provide novel insights into the composition of protein complexes that we suggest are integral to the regulation of OM maintenance.…”

MAB_3551c encodes the primary triacylglycerol synthase involved in lipid accumulation in Mycobacterium abscessus

“…Once inside the cytosolic compartment, the fatty acids either enter β‐oxidation providing energy and carbon or they are rapidly coupled to a CoA molecule through the action of an acyl‐CoA synthetase and subsequently used to produce TAG through the action mainly of Tgs1, but also possibly to a much lesser extent Tgs4. Our subcellular fractionation data, in agreement with previous studies (Elamin et al ., ; Hayashi et al ., ), suggests that all of this occurs at the inner leaflet of the cell membrane. The TAG which is produced is either transported towards the outer cell wall through LprG/Rv1410 (Martinot et al ., ) or it is accumulated in the form of ILI in the cytosol where it may now act as a reservoir of a rich energy and carbon source during prolonged infection.…”