Date of Award
5-1-2011
Degree Name
Master of Science
Department
Molecular Biology Microbiology and Biochemistry
First Advisor
Bender, Kelly
Abstract
Species in the genus Burkholderia are found in a wide variety of environments ranging from plant rhizospheres to the human respiratory tract. Even though they have great biotechnological and bioremediative potential in preventing some plant diseases, promoting crop production, and degrading toxic compounds, some species of Burkholderia can be serious pathogens to those who have compromised immune systems, particularly those with Cystic Fibrosis (CF). Many Burkholderia species have two distinct copies of the hfq gene, a rare phenomenon in bacteria studied to date. As a global regulatory protein, Hfq has been shown to act as an RNA chaperone involved in stress responses, survival, and virulence in a variety of other bacteria via riboregulation. To address the significance of possessing two distinct Hfq proteins, sequence and expression analyses of the two corresponding B. cenocepacia genes were performed. RT-PCR revealed that both hfq genes were expressed constitutively and that neither hfq1 nor hfq2 appear to be transcribed as part of an operon. Phylogenetic analysis indicated that Bc-Hfq2 clustered with Hfq proteins from other Beta-Proteobacteria, while Bc-Hfq1 shared a common ancestor with the Hfq from Delta-Proteobacteria. This incongruence with 16S rRNA gene phylogeny suggests that the B. cenocepacia hfq1 gene may have been acquired through horizontal transfer. Further analysis of the two B.cenocepacia proteins indicated that Bc-Hfq2 was able to partially complement an E. coli hfq mutant, while the effect of heterologously expressing hfq1 could not be determined. A corresponding B. cenocepacia hfq2 deletion strain was constructed utilizing homologous recombination. This mutant showed extended lag phases when grown at 37°C and 41°C and a slower growth rate at a pH of 5 compared to the wild type. These data suggest that Hfq plays an important and yet still not fully understood role in the stress response of Burkholderia species.
Access
This thesis is Open Access and may be downloaded by anyone.