Transcription factor LSF: interactions with protein partners leading to epigenetic regulation and microtubule modifications
Chin, Hang Gyeong
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Transcription factor LSF is an oncogene in Hepatocellular Carcinoma (HCC). HCC is the sixth most common cancer worldwide and the second highest cause of cancer-related death globally. LSF is overexpressed in human HCC cell lines, compared to normal hepatocytes, and expression levels show significant correlation with the stage and grades of the disease. Current treatments for HCC are insufficient, especially given the frequency of late stage diagnoses. Therefore, it is necessary to understand the molecular mechanism of HCC disease to aid in targeted and effective treatments. Most investigations of the regulation of LSF activity have focused on its post-translational modifications in response to cellular proliferation and signal transduction. Chromatin modifications and epigenetic mechanisms of LSF-mediated gene regulation had not been investigated. Given that alterations of epigenetic writers or readers have been demonstrated in a large fraction of HCC patient samples, I examined the connection between LSF and epigenetic regulators. In particular, LSF is shown to interact with DNA methyltransferase 1 (DNMT1) and Ubiquitin like with PHD and Ring Finger Domains (UHRF1), with consequences for global DNA methylation and transcription patterns. Additionally, I identified unexpected, pairwise associations between LSF, histone methyltransferase SET8, and tubulin, both in vitro and in vivo. The interactions were identified by proteomics analyses, co-localization, co-immunoprecipitation, and direct protein-protein interaction studies in vitro. Strikingly, both LSF and SET8 associate with microtubules, leading to the discovery that SET8 methylates α-tubulin at several novel, specific lysines. This suggests parallels between regulation of chromatin by the histone code and regulation of microtubule function by the tubulin code. Surprisingly, LSF enhances tubulin methylation by SET8 in vitro and FQI1, a specific LSF small molecule inhibitor, reduces tubulin methylation. Furthermore, LSF promotes, and FQI1 inhibits, tubulin polymerization in vitro. Taken together, these findings suggest that SET8 is a microtubule-associated methyltransferase that LSF recruits to microtubules to enhance tubulin modification. The results indicate that both LSF and SET8 have cellular implications beyond their roles in gene transcription and histone methylation. Finally, this discovery of the dual functions for LSF and SET8 set up the possibility for connections between epigenetic and cytoskeleton modifications in cancer.