Date of Award

12-2009

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biology

First Advisor

Zehava Eichenbaum - Chair

Second Advisor

John Houghton

Third Advisor

Chung-Dar Lu

Fourth Advisor

Phang C. Tai

Abstract

Group A Streptococcus (GAS) is a common pathogen of the human skin and mucosal surfaces and is capable of producing a variety of diseases. This dissertation investigates the function of a metalloregulator named MtsR in GAS physiology and disease process. An mtsR mutant was constructed and analyzed. Consistent with MtsR role in iron uptake regulation, the mtsR mutant accumulates more iron (80 ± 22.5%) than the wild type strain. Inactivation of mtsR results in constitutive transcription of the sia (Streptococcal Iron Acquisition) operon, which is negatively regulated by iron in the parent strain. We identified the promoter that controls the expression of the sia operon (Pshr) and used it as a model to study MtsR interaction with DNA. Electrophoretic mobility gel shift assays (EMSAs) demonstrated that MtsR binds to the shr upstream region specifically and in an iron and manganese dependent manner. DNase I footprint analysis revealed that MtsR protects a 69 bp segment in Pshr that includes 2 inverted repeats, overlapping the core promoter elements. A global transcriptional analysis determined that MtsR modulates the expression of 64 genes, of which 44 were upregulated and 20 were downregulated in the mtsR mutant. MtsR controls genes with diverse functions including immune evasion, colonization, dissemination, metal homeostasis, nucleic acid and amino acid metabolism, and protein stability. MtsR functions as a dual regulator as it binds to the promoters of the repressed genes ska, aroE, and nrdF.2, as well as the upstream region of the positively regulated genes mga, emm49, and pyrF. A 16 bp MtsR-binding consensus region was identified in all of the promoters that are directly regulated by MtsR. In conclusion, we have demonstrated that MtsR is a global regulator in GAS that controls the expression of vital virulence factors and genes involved in metal transport, virulence and metabolic pathways.

DOI

https://doi.org/10.57709/1234542

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