title: Unexpected link between lipooligosaccharide biosynthesis and surface protein release in Mycobacterium marinum.
abstract: The mycobacterial cell envelope is characterized by the presence of a highly impermeable second membrane, which is composed of mycolic acids intercalated with different unusual free lipids, such as lipooligosaccharides (LOS). Transport across this cell envelope requires a dedicated secretion system for extracellular proteins, such as PE_PGRS proteins, which are specific mycobacterial proteins with polymorphic GC-rich sequence (PGRS). In this study, we set out to identify novel components involved in the secretion of PE_PGRS proteins by screening Mycobacterium marinum transposon mutants for secretion defects. Interestingly, most mutants were not affected in secretion but in the release of PE_PGRS proteins from the cell surface. These mutants had insertions in a gene cluster associated with LOS biosynthesis. Lipid analysis of these mutants revealed a role at different stages of LOS biosynthesis for 10 novel genes. Furthermore, we show that regulatory protein WhiB4 is involved in LOS biosynthesis. The absence of the most extended LOS molecule, i.e. LOS-IV, and a concomitant accumulation of LOS-III was already sufficient to reduce the release of PE_PGRS proteins from the mycobacterial cell surface. A similar effect was observed for major surface protein EspE. These results show that the attachment of surface proteins is strongly influenced by the glycolipid composition of the mycobacterial cell envelope. Finally, we tested the virulence of a LOS-IV-deficient mutant in our zebrafish embryo infection model. This mutant showed a marked increase in virulence as compared with the wild-type strain, suggesting that LOS-IV plays a role in the modulation of mycobacterial virulence.
authors.affiliation: Department of Medical Microbiology and Infection Control, VU University Medical Center, 1081 HV Amsterdam, The Netherlands.
authors.forename: Aniek D
authors.initials: AD
authors.lastname: van der Woude
authors.forename: Debasmita
authors.initials: D
authors.lastname: Sarkar
authors.forename: Apoorva
authors.initials: A
authors.lastname: Bhatt
authors.forename: Marion
authors.initials: M
authors.lastname: Sparrius
authors.forename: Susanne A
authors.initials: SA
authors.lastname: Raadsen
authors.forename: Louis
authors.initials: L
authors.lastname: Boon
authors.forename: Jeroen
authors.initials: J
authors.lastname: Geurtsen
authors.forename: Astrid M
authors.initials: AM
authors.lastname: van der Sar
authors.forename: Joen
authors.initials: J
authors.lastname: Luirink
authors.forename: Edith N G
authors.initials: EN
authors.lastname: Houben
authors.forename: Gurdyal S
authors.initials: GS
authors.lastname: Besra
authors.forename: Wilbert
authors.initials: W
authors.lastname: Bitter
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001693:Biological Transport, Active / Q000502:physiology; D004251:DNA Transposable Elements / Q000502:physiology; D005393:Fish Diseases / Q000235:genetics / Q000378:metabolism; D008070:Lipopolysaccharides / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D009165:Mycobacterium Infections, Nontuberculous / Q000235:genetics / Q000378:metabolism / Q000662:veterinary; D019910:Mycobacterium marinum / Q000235:genetics / Q000378:metabolism*; D015027:Zebrafish / Q000382:microbiology
chemical_list: D001426:Bacterial Proteins; D004251:DNA Transposable Elements; D008070:Lipopolysaccharides