title: Involvement of a plasmid-encoded type IV secretion system in the plant tissue watersoaking phenotype of Burkholderia cenocepacia.
abstract: Burkholderia cenocepacia strain K56-2, a representative of the Burkholderia cepacia complex, is part of the epidemic and clinically problematic ET12 lineage. The strain produced plant tissue watersoaking (ptw) on onion tissue, which is a plant disease-associated trait. Using plasposon mutagenesis, mutants in the ptw phenotype were generated. The translated sequence of a disrupted gene (ptwD4) from a ptw-negative mutant showed homology to VirD4-like proteins. Analysis of the region proximal to the transfer gene homolog identified a gene cluster located on the 92-kb resident plasmid that showed homology to type IV secretion systems. The role of ptwD4, ptwC, ptwB4, and ptwB10 in the expression of ptw activity was determined by conducting site-directed mutagenesis. The ptw phenotype was not expressed by K56-2 derivatives with a disruption in ptwD4, ptwB4, or ptwB10 but was observed in a derivative with a disruption in ptwC. Complementation of ptw-negative K56-2 derivatives in trans resulted in complete restoration of the ptw phenotype. In addition, analysis of culture supernatants revealed that the putative ptw effector(s) was a secreted, heat-stable protein(s) that caused plasmolysis of plant protoplasts. A second chromosomally encoded type IV secretion system with complete homology to the VirB-VirD system was identified in K56-2. Site-directed mutagenesis of key secretory genes in the VirB-VirD system did not affect expression of the ptw phenotype. Our findings indicate that in strain K56-2, the plasmid-encoded Ptw type IV secretion system is responsible for the secretion of a plant cytotoxic protein(s).
authors.affiliation: Department of Plant Pathology & Microbiology, Texas A&M University, 2132 TAMU, College Station, TX 77843, USA.
authors.forename: Amanda S
authors.initials: AS
authors.lastname: Engledow
authors.forename: Enrique G
authors.initials: EG
authors.lastname: Medrano
authors.forename: Eshwar
authors.initials: E
authors.lastname: Mahenthiralingam
authors.forename: John J
authors.initials: JJ
authors.lastname: LiPuma
authors.forename: Carlos F
authors.initials: CF
authors.lastname: Gonzalez
mesh_terms: D001426:Bacterial Proteins / Q000378:metabolism / Q000633:toxicity; D001692:Biological Transport / Q000235:genetics; D019121:Burkholderia Infections; D042602:Burkholderia cepacia complex / Q000235:genetics* / Q000378:metabolism / Q000472:pathogenicity*; D003227:Conjugation, Genetic; D003603:Cytotoxins / Q000378:metabolism* / Q000633:toxicity; D004269:DNA, Bacterial / Q000737:chemistry; D023061:Gene Order; D005798:Genes, Bacterial; D005816:Genetic Complementation Test; D005810:Multigene Family; D016254:Mutagenesis, Insertional; D016297:Mutagenesis, Site-Directed; D019697:Onions / Q000382:microbiology; D010935:Plant Diseases / Q000382:microbiology*; D010957:Plasmids; D017422:Sequence Analysis, DNA; D017385:Sequence Homology; D037521:Virulence Factors / Q000378:metabolism / Q000633:toxicity
chemical_list: D001426:Bacterial Proteins; D003603:Cytotoxins; D004269:DNA, Bacterial; D037521:Virulence Factors