UC Santa Cruz

While it's known that the dysregulation of splicing contributes to disease, identifying specific splicing alterations is a bottleneck for current research. On an individual scale, mutations in signaling cues disrupt canonical splicing and force the production of an aberrant splicing event. On a global scale, splicing factor mutations result in widespread splicing dysregulation and create a subset of isoforms that contribute to disease progression. In both cases, identifying specific splicing aberrations that drive disease is important in developing targeted therapies, and can greatly improve the lives of patients bearing these mutations. To study novel deleterious splicing events, scientists need the appropriate tools to identify and functionally characterize these isoforms. In chapter 2 of this dissertation, I found that using CRISPR/Cas9 to express aberrant exon skipping events from their endogenous promoter did not produce enough aberrant isoform to confirm the oncogenic potential in two functional assays. Because exon skipping events, alone, were not sufficient to confer an oncogenic phenotype, I investigated the level of a known oncogenic isoform, MET exon 14 skipping (METΔ14), expressed in lung adenocarcinoma primary samples in chapter 3. I found that METΔ14 is overexpressed in an allele-specific manner, and METΔ14 overexpression is required to activate the receptor. In chapter 4, through a collaboration with Dr. Esther Obeng’s lab, I performed long read sequencing in cell lines expressing mutant SF3B1, a common splicing factor mutation in Myelodysplastic Syndrome (MDS). This collaboration uncovered previously unannotated isoforms with potential implication in MDS. Overall, the work in this dissertation outlines the trials and tribulations of using the correct tools to functionally characterize aberrant splicing events, and describes triumphs of discovering how METΔ14 is expressed in lung cancer tumors and isoforms implicated in MDS-associated anemia requiring further characterization.