Genome-Scale Analysis of Transcriptional Regulation by the Fbw7 Ubiquitin Ligase

Abstract

Protein turnover is tightly regulated by the ubiquitin proteasome system (UPS). Dysregulated UPS components lead to many diseases, including cancer. Fbw7 is the substrate receptor of the SCFFbw7 E3 ubiquitin ligase and is one of the most commonly mutated tumor suppressor genes. Fbw7 is mutated in a wide array of cancers, including T-cell leukemia, colorectal and endometrial cancers. Fbw7 targets more than two dozen proteins for degradation and the majority of them are transcription factors (TFs). Therefore, Fbw7 exerts a widespread influence on transcription and cellular mechanisms. A holistic molecular level overview of how Fbw7 regulates multiple TFs and global transcription is needed to unravel the complex regulatory network of Fbw7. Heterozygous arginine missense mutations (Fbw7Arg/+) that impair Fbw7’s function is favored by human cancers over complete loss of Fbw7. However, the tumorigenic mechanisms led by Fbw7Arg/+ are as of yet poorly understood. Therefore, I initiated an integrative study where I employed RNA-Seq and chromatin profiling tools (CUT&RUN) to understand context-specific transcriptional regulation in Fbw7 wild-type and mutant cells. I investigated the deregulation of two well-studied oncogenic Fbw7 substrates (cMyc, cJun) in colorectal cancer cell lines and neural stem cells. This work revealed that Fbw7 regulates substrates at specific genomic sites which may not always be reflected by analysis of mRNA. Fbw7 preferentially regulates TF occupancy at distal regulatory genomic regions; however, the specific targeted genes and genomic loci differ in a cell-type and mutation-type specific manner. Fbw7 also coordinately co-regulates both Jun and Myc at some genomic sites. I discovered that loss of Fbw7 upregulates class II HLA genes by controlling cMyc and cJun occupancy at one of the co-regulated sites, which is within the CIITA upstream regulatory region. Together, this body of work improves our understanding of context-specific transcriptional regulation by Fbw7 that may be useful in developing better targeted therapies. The identification of class II HLA regulation by Fbw7 may be incorporated in developing prognostic and diagnostic methods.