Project description:Bacterial outer membrane vesicles (OMVs) have important biological roles in pathogenesis and intercellular interactions, but a general mechanism of OMV formation is lacking. Here we show that the VacJ/Yrb ABC transport system, a proposed phospholipid (PL) transporter, is involved in OMV formation. Deletion or repression of VacJ/Yrb increases OMV production in two distantly related Gram-negative bacteria, Haemophilus influenzae and Vibrio cholerae. Within our studies we also analyzed the proteome of the OMV and outer membrane (OM) and found no massive alteration in Haemophilus influenzae Rd KW20, Rd ∆yrbE and Rd ∆vacJ. Lipidome analyses demonstrate that OMVs from VacJ/Yrb-defective mutants in H. influenzae are enriched in PLs and certain fatty acids. Furthermore, we demonstrate that OMV production and regulation of the VacJ/Yrb ABC transport system respond to iron starvation. Our results suggest a new general mechanism of OMV biogenesis based on PL accumulation in the outer leaflet of the outer membrane. This mechanism is highly conserved among Gram-negative bacteria, provides a means for regulation, can account for OMV formation under all growth conditions, and might have important pathophysiological roles in vivo.
Project description:Investigation of whole genome gene expression level changes in a Vibrio cholerae O395N1 delta-nqrA-F mutant, compared to the wild-type strain.
Project description:Environmental isolates of Vibrio cholerae from California coastal water compared to reference strain N16961. A genotyping experiment design type classifies an individual or group of individuals on the basis of alleles, haplotypes, SNP's. Keywords: genotyping_design; array CGH
Project description:Investigation of whole genome gene expression level changes in a Vibrio cholerae O395N1 delta-nqrA-F mutant, compared to the wild-type strain. Total RNA recovered from wild-type cultures of VIbrio cholerae O395N1 and its nqrA-F mutant strain. Each chip measures the expression level of 3,835 genes from Vibrio cholerae O1 biovar eltor str. N16961 with twenty average probes/gene, with five-fold technical redundancy.
Project description:Vibrio campbellii BB120 (ATCC BAA-1116, previously designated as Vibrio harveyi) is a fundamental model strain for studying population density-based cell-to-cell communication, known as quorum sensing, among gram-negative bacteria. In V. campbellii BB120, sensing of autoinducers at high cell densities activates the expression of the master transcriptional regulator, LuxR, which controls the expression of genes involved in group behaviors. Unlike BB120, the Vibrio campbellii environmental isolate DS40M4 was recently shown to be capable of natural transformation, a process by which bacteria take up exogenous DNA and incorporate it into their genome via homologous recombination. Here, we compare other phenotypes between DS40M4 and BB120. We find that DS40M4 has a faster growth rate and stronger type VI secretion-mediated cell killing, whereas BB120 forms more robust biofilms and is bioluminescent. We exploited the power of natural transformation to rapidly generate >30 mutant strains to explore the function of DS40M4-encoded homologs of the BB120 quorum-sensing system. Our results show that DS40M4 has a similar quorum-sensing circuit to BB120 but with three distinct differences: 1) DS40M4 lacks the canonical HAI-1 autoinducer LuxM synthase but has an active LuxN receptor, 2) the quorum regulatory small RNAs (Qrrs) are not solely regulated by autoinducer signaling through the response regulator LuxO, and 3) the DS40M4 LuxR regulon is <100 genes, which is relatively small compared to the >400 genes regulated in BB120. This work illustrates that DS40M4 is a tractable and relevant model strain for studying quorum-sensing phenotypes in Vibrio campbellii.
