Detection Of Diffusible Chemical Signals By Two Species Of Proteobacteria
This dissertation focuses on how two different species of Proteobacteria, Yersinia enterocolitica and Agrobacterium tumefaciens, detect chemical signals that are synthesized by the same species or by other organisms. Y. enterocolitica encodes proteins YenR and YenI, that resemble the LuxR and LuxI proteins of Vibrio fischeri. YenI synthesizes mainly 3-oxohexanoylhomoserine lactone (OHHL), while YenR is thought to be an OHHL-dependent transcriptional activator. Chromatin immunoprecipitation assays suggested that YenR might bind to a region upstream of the yenR gene. This was confirmed using purified YenR in electrophoretic mobility shift assays. Bound YenR activated a gene divergent from yenR, designated yenS, and did so only in the absence of OHHL. The yenS gene encodes two overlapping non-translated RNAs that share the same promoter. In cells expressing yenI, the yenS promoter was expressed preferentially at low cell densities. A bioinformatic analysis showed that a portion of YenS was complementary to the ribosome binding site and start codon of the yenI mRNA. The activity of yenI-lacZ translational fusions was inhibited by YenS. Earlier studies of the YenR/YenI system suggested a role in swimming and swarming motility. A yenI mutant was strongly motile on semisolid agar, while the wild type strain, the yenR mutant, and the yenS mutant were nonmotile. A yenI, yenR double mutant and a yenI, yenS double mutant were nonmotile, suggesting a direct positive role for YenR and YenS. A separate study focused on the OccR protein, which is encoded by the tumor inducing (Ti) plasmid of A. tumefaciens. OccR activates an operon that directs the uptake and catabolism of a tumor-released nutrient called octopine. In the absence of octopine, OccR binds to its operator and causes a high angle DNA bend. In contrast, OccR-octopine complexes bind DNA and cause a low angle DNA bend. mutagenesis. Eight positive control mutants were identified by random Of these, six mutants showed the same high angle bend as wild type in the absence of octopine, but retained this bend in the presence of octopine. These data support the idea that relaxation of the DNA bend by the inducing ligand is critical for transcriptional activation.
Yersinia enterocolitica; Quorum sensing; LysR protein
Winans, Stephen C
Roberts, Jeffrey Warren; Marquis, Helene
Ph.D. of Microbiology
Doctor of Philosophy
dissertation or thesis