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ORCID

http://orcid.org/0000-0003-0612-6280

Date of Award

Spring 5-15-2020

Author's School

Graduate School of Arts and Sciences

Author's Department

Biology & Biomedical Sciences (Molecular Cell Biology)

Degree Name

Doctor of Philosophy (PhD)

Degree Type

Dissertation

Abstract

Nonsense mediated RNA decay (NMD) is an RNA surveillance pathway present in all eukaryotes that detects and degrades nonsense mRNAs, which contain pre-mature translation termination codons. Nonsense mRNAs are prevalent when pre-mRNA splicing is altered or defective. Interestingly, defective pre-mRNA splicing is emerging as a major driver of cancer development, including development of myelodysplastic syndrome (MDS), leukemia, and some solid tumors. Moreover, pre-mRNA splicing is also thought to enhance NMD in human cells, although itճ still unclear whether and how splicing or splicing factors promote NMD. The role of NMD in regulating mis-spliced mRNA and the link between NMD and RNA splicing, suggest that understanding the process of NMD in the context of normal and defective splicing may hold some clues on developing therapies to treat cancers with dysregulated splicing. To better understand the process of NMD, we have developed a novel NMD reporter system to measure NMD activity in individual human cells and used it to perform a genome-wide CRISPR/Cas9 KO screen to identify genes that promote NMD. We found that the SF3B spliceosome complex promotes NMD without splicing of the target mRNA, suggesting that recruitment of certain spliceosome factors, but not pre-mRNA splicing per se, promotes NMD. In the context of defective splicing, we found that expression of cancer-associated spliceosome mutants (including mutant SF3B1) attenuate NMD. Importantly, cancer cells harboring spliceosome mutations were remarkably sensitive to inhibition of NMD. Therefore, inhibition of NMD is a novel potential therapeutic strategy to treat cancers with defective splicing. This finding suggests that small molecule inhibitors of NMD are needed to facilitate development of therapies that target the NMD pathway. In this dissertation, we have evaluated the use of two different compounds to inhibit NMD. SMG1i directly targets SMG1, the only kinase in the NMD pathway, while Compound C, a commonly used AMPK inhibitor, inhibits NMD indirectly probably by down-regulating NMD factors. Compound C is, however, non-specific, but its derivatives may generate specific NMD inhibitors. Collectively, our studies shed some new light on the process of NMD in the context of normal or defective splicing, uncover NMD as a novel vulnerability of cancers with defective splicing, and provide promising lead compounds for developing therapies that target NMD for cancer treatment.

Language

English (en)

Chair and Committee

Zhongsheng You

Committee Members

Sergej Djuranovic, Jean Schaffer, Amber Stratman, Heather True-Krob,

Available for download on Saturday, March 27, 2021

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