Characterization of Novel Post-Transcriptional Events Misregulated In Disease: Implications for the Development of Future Therapies

Title: Characterization of Novel Post-Transcriptional Events Misregulated In Disease: Implications for the Development of Future Therapies
Authors: Bondy-Chorney, Emma
Date: 2017
Abstract: The misregulation of post-transcriptional mechanisms has been linked to the development and progression of numerous human diseases, in particular neurological disorders and cancer. Investigating these misregulated RNA pathways is essential to fully understand the disease mechanisms, identify novel biomarkers, and to develop effective therapies. In this thesis, I present three manuscripts that investigate the mechanisms behind the post-transcriptional misregulation of RNA in human disease, with a focus on pre-mRNA splicing. In the first manuscript (Bondy-Chorney et al., 2016a), we investigated the role of Staufen1 (Stau1) in splicing regulation in the neuromuscular disorder Myotonic Dystrophy Type 1 (DM1). Here we report the first insights into the mechanism that Stau1 uses to regulate the alternative splicing of INSR exon 11 through an interaction with Alu elements located in intron 10. Moreover, using a high-throughput RT-PCR screen, we uncovered a number of additional Stau1-regulated alternative splicing events in both wild-type and DM1 myoblast cell lines. As Stau1 is known to be aberrantly upregulated in DM1 skeletal muscle, our findings suggest that Stau1 acts as a disease modifier in this disorder. The second manuscript (Sanchez, Bondy-Chorney et al., 2015), describes a novel role of the protein methyltransferase Coactivator-Associated Methyltransferase-1 (CARM1), a protein found to be overexpressed in Spinal Muscular Atrophy (SMA). We found that CARM1 can act as a mediator in the nonsense-mediated decay pathway (NMD) and associated UPF1 to promoted its occupancy on PTC-containing transcripts. We identified a subset of natural non-PTC containing NMD targets that were dependent on CARM1, a number of which were misregulated in SMA. This work uncovered a novel role for CARM1 in the NMD pathway and revealed that defective targeting of PTC-containing mRNAs should be included in the complex array of molecular defects associated with SMA. Finally, the third manuscript (Bondy-Chorney et al., – in prep) examines the alternative iv splicing regulation of the Protein Arginine Methyltransferase PRMT1 exon 2, an event shown to alter the growth, survival, and invasion of breast cancer cells. Here, we used an RNA interference (RNAi) RT-PCR screen to uncover several splicing proteins that regulate the inclusion of exon 2, several of which we found to be misregulated in a panel of breast cancer cell lines and patient tumours. These findings confirmed that the inclusion of PRMT1 exon 2 was regulated by alternative splicing via splicing factors that are altered in breast cancer. Moreover, depletion of one of these splicing factors, RALY, resulted in a decrease in the motility and invasive potential of an aggressive breast cancer cell line. These three manuscripts represent a collection of work focused on elucidating the mechanisms involved in post-transcriptional misregulation of RNA in three diverse human diseases. Taken together, the data presented here highlight the broad impact that proteins, such as Stau1 and CARM1, can have in neuromuscular disorders. Moreover, we also uncovered novel misregulation of splicing proteins that alter alternative splicing patterns in breast cancer. Elucidating these mechanisms is of the highest importance in order to identify potential new and effective treatment avenues.
CollectionThèses, 2011 - // Theses, 2011 -