Serine/threonine kinase signaling in beta-lactam resistance of Staphylococcus aureus

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Project Abstract: We have identified novel roles of the eukaryotic-like serine/threonine kinase (eSTK) signaling pathway in mediating broad-spectrum β-lactam resistance in methicillin-resistant Staphylococcus aureus (MRSA). Broad-spectrum β-lactam resistance, which renders most β-lactam drugs therapeutically ineffective, is classically mediated through mecA, the gene that encodes penicillin-binding protein 2a. Broad-spectrum resistance to β-lactams in S. aureus also occurs through non-classical mediators not directly related to mecA. The role played by the non-classical mediators in β-lactam resistance is only superficially understood. Both Stk1 and Stp1 (effectors of eSTK signaling; a serine/threonine kinase and phosphatase), mediate β-lactam sensitivity by loss of function or overexpression respectively, whereas functional Stk1 and non-functional Stp1 favor drug resistance. Our results show that eSTK modulates β-lactam resistance via pathways controlling mecA expression and through unknown non-classical mediators independently regulated by Stk1 and Stp1. mecA expression in community MRSA strains (and many hospital strains as well) is regulated by the BlaR1- BlaI regulatory pathway. Expression of mecA is normally suppressed by the transcriptional repressor, BlaI. Presence of β-lactam drugs is sensed by BlaR1, an integral membrane protein. Subsequently, BlaR1 undergoes a site-specific auto-proteolysis releasing its intracellular zinc metalloprotease (ZnMP) domain into the bacterial cytosol. The released ZnMP degrades BlaI to de-repress mecA expression, leading to drug resistance. Our data show that eSTK mediated phosphorylation of BlaR1 is important for efficient mecA induction. Through passaging studies, we have identified compensatory mechanisms that enable the bacteria to overcome drug sensitivity due to Stk1 loss of function or Stp1 overexpression, mentioned above. Genome sequencing studies carried out to decipher the basis of resistance in passaged strains indicated involvement of pathways that are unrelated to mecA. Three aims are proposed: a) to decipher the mechanism through which eSTK controls mecA expression, b) to identify eSTK mediators that confer non-classical β-lactam resistance, and c) to investigate the compensatory basis of resistance among resistant passaged strains. Our study will help determine the mechanism/s through which Stk1 and Stp1 control β-lactam resistance and could help to identify novel and improved treatment options for S. aureus infections.