Borrelia burgdorferi, the causative agent of Lyme disease, is maintained in nature through an infectious cycle between wild mammals and ticks. Like many bacterial pathogens, B. burgdorferi must cope with an array of changing environmental conditions to successfully persist, proliferate and be transmitted between hosts. The bacterial outer surface represents the primary site for interactions with the host. The array of B. burgdorferi outer surface proteins (Osps) has been shown to vary with the infectious cycle. It is likely that these different Osps endow the spirochete with distinct properties relevant to the disparate environments in which it must survive. Our broad objective is to use a molecular genetic approach to elucidate the molecular mechanisms of adaptation in B. burgdorferi and their roles in the infectious cycle. We have previously demonstrated that temperature represents an important environmental variable, and that synthesis of a number of Osps is increased after a rise in culture temperature. Protein levels parallel transcript levels, consistent with regulation at the level of gene expression. The complete genomic sequence of B. burgdorferi identified homologs of three transcription factors, sigma 70, sigma 54, and RpoS. We are interested in the roles of RpoS and sigma 54 in the temperature induction and transcriptional regulation of the osp genes. We inactivated the chromosomally encoded sigma factor RpoS. The B. burgdorferi RpoS mutant exhibited an altered stationary phase response and multiple differences in protein composition relative to wild type. Mass spectroscopy of total bacerial proteins separated by 2-dimensional gels has been used to identify putative genes regulated by RpoS. Further characterization of the RpoS mutant will provide information about how B. burgdorferi adapts to variable environmental conditions. The genomic sequence of B. burgdorferi also identified a homolog of a transporter for the disaccharide chitobiose, which could be useful for nutrient acquisition during growth in the tick. We have begun a genetic analysis of the chitobiose transporter genes (chbA, chbB and chbC) to determine their role and function in B. burgdorferi. We inactivated the chbB gene, encoding the putative membrane-spanning protein, and have compared growth of the mutant in various media to that of its isogenic parent. The mutant is unable to utiize chitobiose, whereas neither the mutant nor wild type bacteria can utilize cellobiose. We have also found that the chbB gene is regulated by growth temperature and medium composition. Studies with wild type B. burgdorferi demonstrated that chitobiose can replace N-acetyl glucosamine, which was previously considered essential, at a fraction of the molar concentration. Since chitobiose is a component of tick cuticle, we predict that the chbB genes should be important for bacterial growth in the host.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Intramural Research (Z01)
Project #
1Z01AI000802-04
Application #
6431686
Study Section
(LHBP)
Project Start
Project End
Budget Start
Budget End
Support Year
4
Fiscal Year
2000
Total Cost
Indirect Cost
Name
Niaid Extramural Activities
Department
Type
DUNS #
City
State
Country
United States
Zip Code
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Motaleb, M A; Corum, L; Bono, J L et al. (2000) Borrelia burgdorferi periplasmic flagella have both skeletal and motility functions. Proc Natl Acad Sci U S A 97:10899-904

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