Methicillin-resistant Staphylococcus aureus (MRSA) is causing major healthcare problems worldwide and is becoming increasingly challenging to treat with current antibiotics. S. aureus is a common cause of skin infections and respiratory diseases that can be life-threatening. This emphasizes the importance of developing alternative therapeutic approaches to combat infections. S. aureus is an effective pathogen because it can rapidly adapt to environmental insults, such as attacks by the immune system and antibiotic stress. To achieve such rapid adaptation, S. aureus remodels its transcriptome within minutes of stress imposition. It was thought that transcription factors were mainly responsible for directing this process; however, post-transcriptional regulators, such as RNA-binding proteins (RBPs), are now recognized as key players in controlling adaptive responses by modulating mRNA translation and/or degradation rates.
Although RBPs are believed to play a fundamental role in regulating gene expression during stress adaptation, it remains unclear which RBPs are key players in this process and how they control rapid gene expression remodeling. Using novel proteomic approaches[1,2], we have recently unearthed the RNA-binding proteome (RBPome) in S. aureus cells. Strikingly, many peptidoglycan synthesis enzymes, including PBPs, were identified as putative RBPs in our RBPome. This included Pbp2a, a protein that offers S. aureus resistance to many β-lactam antibiotics. This was surprising since these enzymes primarily reside in the cell wall, where they are presumably less likely to encounter RNA. Our recent data imply that PBPs bind RNA in the cytosol, presumably co-translationally. Drawing on these results, and given that PBPs are prone to aggregation, we hypothesize that RNA may regulate the folding of PBPs and prevent them from becoming enzymatically active before they reach their final destination in the cell wall.
The Goals of This Project Are To:
- Understand how RNA impacts the folding and localization of MRSA PBPs.
- Determine how RNA regulates the enzymatic activity of PBPs.
- Investigate whether PBP RNA-binding activity can be exploited to develop new tools and RNA-based therapies.
The techniques that the student will be using include cross-linking and immunoprecipitation assays (CRAC; Chu et al., 2022), the production of recombinant proteins, testing of protein-RNA interactions in vitro using various biochemical approaches, genetic manipulation of S. aureus, in silico modeling of protein-RNA interactions, fluorescence microscopy, analysis of high-throughput datasets, and programming using Python and R. Therefore, the project offers an exceptional learning environment for students who wish to cultivate skills in these domains.
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