TY - JOUR TI - Insights into mechanisms of metal homeostasis in Staphylococcus aureus DO - https://doi.org/doi:10.7282/T3VT1V8F PY - 2016 AB - Staphylococcus aureus is a major public health concern worldwide. The human immune system employs oxidative- and copper-dependent mechanisms that disrupt metal homeostasis within the invading microorganism. This thesis describes new factors involved in iron-sulfur cluster biogenesis and copper detoxification in S. aureus, representing additional mechanisms of metal homeostasis that might aid this pathogen during host infection. Strains lacking the low-molecular-weight thiol bacillithiol (bshA mutants) display growth defects that are alleviated by the addition of exogenous Fe or the amino acids leucine and isoleucine. The bshA strain has decreased activities of Fe-S proteins, such as LeuCD, IlvD, AcnA, and GltBD, suggesting that the absence of BSH leads to a general defect in Fe-S cluster biogenesis. The growth defects and decreased enzymatic activities of the bshA strain are exacerbated in strains lacking other factors involved in Fe-S cluster biogenesis, such as the Fe-S cluster carriers Nfu and SufA, and partially suppressed by their overexpression, suggesting a functional overlap between BSH and Fe-S cluster carriers in Fe-S cluster biogenesis. We also found a two-gene operon involved in preventing copper (Cu) intoxication. These genes encode an ATPase Cu transporter (copB) and a putative lipoprotein (cbl). Mutational inactivation of copB or cbl resulted in Cu sensitivity, and their phenotypes are exacerbated in strains also lacking the highly conserved Cu exporter CopA, suggesting that CopB and Cbl are additional mechanisms that prevent Cu intoxication in S. aureus. Overexpression of either copB or cbl leads to increased Cu resistance in other S. aureus clinical isolates lacking these genes. We show that copB and cbl are co-transcribed, up-regulated under Cu stress, and repressed by CsoR. Genetic and biochemical evidence show that Cbl is a membrane-associated, surface-exposed lipoprotein that binds up to four Cu+ ions. Collectively, the research presented in this thesis describes new roles for BSH and Cbl in metal homeostasis in the human pathogen S. aureus. KW - Microbial Biology KW - Staphylococcus aureus KW - Homeostasis LA - eng ER -