Exploring the roles and interplay of REST and USP15 in cancer

Repressor Element-1 (RE-1) Silencing Transcription Factor (REST), through the actions of associated co-repression complexes, modifies chromatin with repressive epigenetic modifications. REST can regulate a myriad of genes that in part, restricts access to proteins expressed in neuronal cells. During the cell cycle REST is acutely degraded at the onset of mitosis, which is required for effective cell division. As cells exit mitosis, re-expression of REST protein is supported by co-translational deubiquitylation of USP15. The overarching aim of this project was to explore the relationship between REST and USP15 in lung cancer and during the cell cycle. The expression of REST is differentially regulated in neuroendocrine cancers, with a known increase in a truncated isoform, REST4. In response to the finding of a novel protein coding REST transcript, I performed a quantitative assessment of REST expression in thoracic cancers. Simultaneously, I analysed the expression of USP15 and its isoforms. Interestingly, USP15 isoform 1 was markedly increased in neuroendocrine cancer cell lines. During the cell cycle, the truncated REST4 isoform was increased eight-fold during mitosis, which coincides with the degradation of full-length protein. The splicing factor proposed to regulate splicing of REST was not identified in the cell line. The regulation of gene expression by full-length REST underpins its tumour suppressive function. The loss of REST elicits broad transcriptional changes and is complicated by intricate feedback loops of opposing mRNA and miRNAs. As such, the consequences of REST dysregulation or loss on the cellular proteome and systems are not well studied. Here, I addressed the consequences of the loss of REST in lung cancer, using a proteomic approach in combination with siRNA knockdown. The proteins responsive to REST or USP15 knockdown, were used to determine cellular pathways that could influence cellular physiology. The aim of which is to determine whether the loss of REST protein promotes the neuroendocrine phenotype or metastatic properties in lung cancer. As USP15, is a known regulator of REST protein expression, parallel proteomic studies were performed and correlated to determine the interplay of USP15 and REST. USP15-mediated regulation of mitochondrial Complex I proteins, identified by proteomic analysis, suggested a potentially novel role for USP15. The downregulation of Complex I proteins and proposed dysfunction of mitochondrial Complex I was not apparent in high-glucose conditions. Future experiments are required to determine the extent of USP15 involvement.