Identification of the important pathways controlled by the splicing regulator Tra2β in lung cancer

Abstract

Transformer 2 proteins are ancient RNA binding proteins conserved across the animal kingdom, with two copies existing in vertebrates – Tra2α and Tra2β. Tra2β plays a crucial role in splicing regulation of targets important to spermatogenesis and various neurological conditions, as well as regulating splice switches important to the progression of several cancers. Recent work in our group has outlined Tra2β as an important regulator of splicing in breast cancer cells, with a particular involvement in the splicing of DNA damage related targets. A large-scale analysis of RNA binding protein expression in major TCGA patient cohorts highlighted Tra2β expression as being significantly downregulated in kidney and thyroid cancer, whilst its expression is significantly upregulated in lung squamous cell carcinoma. This upregulation of Tra2β expression, along with its regulatory role in splicing of key genes in breast cancer, led us to question what role Tra2β might have in the regulation of splicing in lung cancer. In this thesis, RNA interference was used to generate a knockdown model for Tra2β expression in NCI-H520 cells, a squamous lung cancer cell line, which was then analysed by RNA-Sequencing. This was used to search for splicing events that are controlled by Tra2β and might be clinically relevant. I have identified a panel of splicing patterns that have a reciprocal change when Tra2β is up-regulated in lung cancer tissue relative to its depletion in NCI-H520 cells. Preliminary integration of this data with lung cancer survival profiles revealed a functional convergence of target exons affected by Tra2β expression and gene pathways commonly mis-regulated in lung cancer cells. Tra2β targets involved in DNA damage and cell cycle regulation pathways were of particular interest. Alternatively spliced exons in DEPDC1, MKI67 and SGOL1 were studied further in the laboratory, as well as how these might interact to promote lung cancer progression.