Postdoctoral Researcher University of Massachusetts Amherst Amherst, Massachusetts, United States
Takehiro Kado (University of Massachusetts Amherst)| Daisuke Motooka (Osaka University)| Alam García-Heredia (University of Massachusetts Amherst)| Shota Nakamura (Osaka University)| M. Sloan Siegrist (University of Massachusetts Amherst)| Yasu S. Morita (University of Massachusetts Amherst)
Plasma membrane is a heterogeneous mixture of lipids and proteins that includes laterally discrete domains. Membrane domains have been extensively described in eukaryotes, but little is known in prokaryotes. Mycobacterium, a genus that includes several notorious human pathogens, organizes its plasma membrane by creating the intracellular membrane domain (IMD), which is spatially and biochemically separable from the conventional plasma membrane (PM-CW). This membrane architecture is proposed to optimize the synthesis of cell envelope precursors such as peptide glycan (PG) and arabinogalactan (AG) (Fig. 1). However, the mechanisms that govern the formation and maintenance of the IMD is completely unknown. We recently found that membrane fluidizers such as benzyl alcohol and dibucaine can reversibly disrupt the formation of the IMD. To reveal the mechanism of this stress response, in this study, we carried out transposon sequencing (Tn-seq), comparing pools of mutants that were treated or mock-treated with either benzyl alcohol or dibucaine. We found genes involved in peptidoglycan biosynthesis and outer membrane biosynthesis important for withstanding membrane fluidizer treatments, suggesting that the physical pinning by the loadbearing exoskeleton structures is critical for the IMD maintenance and recovery. We also found genes involved in modulating fatty acid structures, indicating the role of membrane lipid composition in the membrane stress response. We are currently verifying the roles of these genes by making gene deletion mutants, and testing the sensitivities of these mutants to the membrane fluidizers.