BcsKC is an essential protein for the type VI secretion system activity in Burkholderia cenocepacia that forms an outer membrane complex with BcsLB.
The type VI secretion system (T6SS) contributes to the virulence of Burkholderia cenocepacia, an opportunistic pathogen causing serious chronic infections in patients with cystic fibrosis. BcsK(C) is a highly conserved protein among the T6SSs in Gram-negative bacteria. Here, we show that BcsK(C) is required for Hcp secretion and cytoskeletal redistribution in macrophages upon bacterial infection. These two phenotypes are associated with a functional T6SS in B. cenocepacia. Experiments employing a bacterial two-hybrid system and pulldown assays demonstrated that BcsK(C) interacts with BcsL(B), another conserved T6SS component. Internal deletions within BcsK(C) revealed that its N-terminal domain is necessary and sufficient for interaction with BcsL(B). Fractionation experiments showed that BcsK(C) can be in the cytosol or tightly associated with the outer membrane and that BcsK(C) and BcsL(B) form a high molecular weight complex anchored to the outer membrane that requires BcsF(H) (a ClpV homolog) to be assembled. Together, our data show that BcsK(C)/BcsL(B) interaction is essential for the T6SS activity in B. cenocepacia.
Aubert
Daniel
D
Department of Microbiology and Immunology, Siebens Drake Medical Research Institute, University of Western Ontario, London, Ontario N6A 5C1, Canada.
MacDonald
Douglas K
DK
Valvano
Miguel A
MA
D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015153:Blotting, Western; D019121:Burkholderia Infections / Q000382:microbiology; D057508:Burkholderia cenocepacia / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism*; D005110:Extracellular Space / Q000378:metabolism; D054884:Host-Pathogen Interactions; D006801:Humans; D047468:Immunoprecipitation; D008264:Macrophages / Q000378:metabolism / Q000382:microbiology; D013058:Mass Spectrometry; D051379:Mice; D008810:Mice, Inbred C57BL; D008970:Molecular Weight; D046912:Multiprotein Complexes / Q000737:chemistry / Q000378:metabolism*; D009154:Mutation; D011485:Protein Binding; D020798:Two-Hybrid System Techniques
D001426:Bacterial Proteins; D046912:Multiprotein Complexes