Characterization of the regulation of CD46 alternative splicing
Tang, Sze Jing
Date of Issue2016-02-15
School of Biological Sciences
Alternative splicing gives rise to multiple mRNA and protein isoforms. This process is catalyzed by the spliceosome and regulated by cis-acting elements, trans-acting factors, transcription and chromatin structure. CD46 is a membrane-bound complement control protein with several extracellular STP domain variants and two mutually exclusive cytoplasmic tails derived from the alternative splicing of cassette exons 7, 8 and 9, or cassette exon 13, respectively. The two cytoplasmic tails have different signaling capacities and regulatory functions in T helper 1 cells and epithelial cells. Besides, the association between aberrant CD46 alternative splicing and autoimmune diseases proposes a possible novel therapy by amending splicing. To understand the splicing regulatory mechanisms of CD46, serial deletion assays in splicing minigenes were performed to identify cis-acting elements while the loss- and gain-of-function assays as well as RNA pull-down were carried out to determine the roles of protein regulators. SRSF1 and PTBP1 repress exon 13 inclusion via exonic splicing silencers. SRSF1 is a common activator yet it inhibits exon 13 inclusion in this context and it appears to do so by direct binding within this exon. On the other hand, the splicing activators, TIA1 and TIAL1, strongly enhance exon 13 recognition via the poly-U rich sequences downstream of the 5'ss. The alternative splicing pattern of exon 13 was efficiently modulated by anti-sense oligonucleotides targeting splicing enhancers or silencers in a dose-dependent manner but not by T-cell signaling. From these studies, a better understanding for CD46 alternative splicing regulation was achieved yet much remains to be elucidated to fully decipher the CD46 alternative splicing mechanisms. In conclusion, splicing regulation is complicated and context-dependent, and thus intensive studies of alternative splicing events are necessary to improve the current knowledge.