The advent of deep sequencing has enabled the discovery of a multitude of novel cellular functions for RNA. Even in the face of all of this new knowledge, long-standing, fundamental questions remain about the classical messenger function of RNA: how it mediates gene expression into protein. My research group will use chemical biology to elucidate spatial and structural regulation of mRNA degradation inside living cells as a mechanism of gene expression regulation, as well as to identify sites and cellular roles of non-canonical translation initiation. We will also develop novel tools to study RNA trafficking via fluorescence microscopy. These methods will provide new insight into how cells turn off gene expression from both normal and disease-related mutant mRNAs, as well as how unexpected translation start sites in cellular mRNAs generate proteins that are essential for cell survival during (patho)physiological processes such as apoptosis, cell division, and mutagenic stress.
1. How is mRNA degradation spatiotemporally regulated in cells?
Cytoplasmic mRNA degradation regulates gene expression by controlling mRNA levels and half-life, and eliminates disease-related mRNAs containing premature stop codons. This process may be spatiotemporally regulated in cells by ribonucleoprotein particles called P-bodies. P-bodies contain mRNA degradation intermediates as well as proteins that function in mRNA decay, though all of their molecular components, internal structure, and function (do they enhance, or inhibit, degradation?) have not been determined. Understanding how mRNA decay is regulated by P-bodies will elucidate how mRNA levels are controlled, whether other metabolic pathways are regulated by similar assembly mechanisms, and whether this regulation is involved in eliminating mutant mRNAs. We will develop chemical tools to probe the structure and function of the P-body inside living cells with high spatial and temporal precision.
2. Non-canonical translation initiation: identifying protein products and cellular roles
The accepted model for eukaryotic translation initiation is that initiation factors recognize the mRNA 5’ cap, and then assemble small ribosomal subunits in initiation complexes that scan the mRNA until they reach the first ATG codon, where translation begins. In reality, ribosome profiling and proteomic studies have revealed that translation from non-canonical start sites, including internal sites and non-ATG start codons, is widespread. We believe that these non-canonical translation events are actually widespread and essential to cellular survival, rather than being rare and aberrant. We will develop methods to isolate and detect all products of initiation at non-canonical sites, as well as to explore a fundamental question: why would evolution select for such permissivity in translation initiation?
3. Engineering novel probes for RNA and ribonucleoprotein particle imaging