Date on Master's Thesis/Doctoral Dissertation
Biochemistry and Molecular Biology
Biochemistry and Molecular Biology, PhD
Tooley, Christine Schaner
methylation; NRMT1; NRMT2; mutations; activity; DNA damage
Protein methylation is an established and critical posttranslational modification controlling multiple cellular events. Alterations in protein methylation have been implicated in many diseases, including cancer. My work focused on the N-terminal trimethylase NRMT1 and the N-terminal monomethylase NRMT2. Previous work proposed that NRMT2 assists NRMT1 by priming its substrates for trimethylation. Importantly, NRMT1 mutations have been found in cancers, and loss of NRMT1 has been shown to promote oncogenic phenotypes in cancer cells. Together, this suggests that altered activities of NRMT1/2 may play a role in cancer progression. Although NRMT1/2 are 50% identical, they differ in key aromatic residues in their active site. Interestingly, mutation of the corresponding aromatic residues in the methyltransferase EZH2 (B-cell lymphoma) changes its activity from a monomethylase to a trimethylase, conferring oncogenicity. Therefore, I hypothesized that the differences in these aromatic residues are responsible for the distinct catalytic activities of NRMT1/2. I also proposed that NRMT1 cancer mutations are responsible for oncogenic phenotypes. My work illustrates that while aromatic residue mutations had no catalytic effect, both NRMT1 cancer mutants N209I (endometrial) and P211S (lung) displayed decreased trimethylase and increased mono-/dimethylase activity. These mutations are located in the peptide-binding channel and suggest there may be a second structural region impacting enzyme specificity. The mutants also required greater time and substrate levels to be comparable to WT NRMT1. Furthermore, in a cellular context lacking endogenous NRMT1, the N209I and P211S mutants were incapable of rescuing trimethylation levels or proliferation. Additional preliminary studies suggested a potential role for NRMT1 in the DNA damage response pathway. However, further studies will be required to shed more light into its cellular function.
Shields, Kaitlyn Marie, "Investigating the impact of NRMT1 cancer mutants on catalytic specificity and the DNA damage response." (2018). Electronic Theses and Dissertations. Paper 3034.
Retrieved from https://ir.library.louisville.edu/etd/3034