Spatially distinct and metabolically active membrane domain in mycobacteria.
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.
Department of Microbiology, University of Massachusetts, Amherst, MA 01003;
Jennifer M
JM
Hayashi
Department of Microbiology, University of Massachusetts, Amherst, MA 01003;
Chu-Yuan
CY
Luo
Division of Rheumatology, Immunology and Allergy, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02445;
Jacob A
JA
Mayfield
Department of Microbiology, University of Massachusetts, Amherst, MA 01003;
Tsungda
T
Hsu
Department of Immunoregulation, Research Institute for Microbial Diseases, Osaka University, Osaka, 565-0871, Japan;
Takeshi
T
Fukuda
Department of Microbiology, University of Massachusetts, Amherst, MA 01003;
Andrew L
AL
Walfield
Department of Microbiology, University of Massachusetts, Amherst, MA 01003;
Samantha R
SR
Giffen
Department of Biochemistry and Molecular Pharmacology and Proteomics and Mass Spectrometry Facility, University of Massachusetts Medical School, Worcester, MA 01605;
John D
JD
Leszyk
Department of Microbiology and Physiological Systems and Howard Hughes Medical Institute, University of Massachusetts Medical School, Worcester, MA 01605;
Christina E
CE
Baer
Department of Microbiology & Molecular Biology, Tufts University School of Medicine, Boston, MA 02111;
Owen T
OT
Bennion
Division of Rheumatology, Immunology and Allergy, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02445;
Ashoka
A
Madduri
Department of Biochemistry and Molecular Pharmacology and Proteomics and Mass Spectrometry Facility, University of Massachusetts Medical School, Worcester, MA 01605;
Scott A
SA
Shaffer
Department of Microbiology & Molecular Biology, Tufts University School of Medicine, Boston, MA 02111; Department of Biomedical Engineering, Tufts University, Medford, MA 02155;
Bree B
BB
Aldridge
Department of Microbiology and Physiological Systems and Howard Hughes Medical Institute, University of Massachusetts Medical School, Worcester, MA 01605;
Christopher M
CM
Sassetti
Department of Microbiology, University of Massachusetts, Amherst, MA 01003;
Steven J
SJ
Sandler
Department of Immunoregulation, Research Institute for Microbial Diseases, Osaka University, Osaka, 565-0871, Japan; Laboratory of Immunoglycobiology, WPI Immunology Frontier Research Center, Osaka University, Osaka, 565-0871, Japan.
Taroh
T
Kinoshita
Division of Rheumatology, Immunology and Allergy, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02445;
D Branch
DB
Moody
Department of Microbiology, University of Massachusetts, Amherst, MA 01003; ymorita@microbio.umass.edu.
Yasu S
YS
Morita
D001426:Bacterial Proteins / Q000378:metabolism*; D050356:Lipid Metabolism / Q000502:physiology*; D021962:Membrane Microdomains / Q000378:metabolism* / Q000648:ultrastructure*; D008565:Membrane Proteins / Q000378:metabolism*; D009161:Mycobacterium / Q000378:metabolism* / Q000648:ultrastructure*
D001426:Bacterial Proteins; D008565:Membrane Proteins
cell envelope; lipid biosynthesis; membrane domain; mycobacteria; polar growth