Abstract 1709: STRAP mediates the stemness of human colorectal cancer cells by epigenetic regulation of Notch pathway
Authors: Lin Jin, Trung Tam Vu, Pran K. Datta
Journal: Cancer Research (2016)

Abstract Human colon cancer-initiating cells (CCICs) are responsible for the unrestrained cell proliferation and chemoresistance of colorectal tumors. Notch signaling is critical for the maintenance and self-renewal of cancer-initiating cells in colon cancer. We have shown that the serine-threonine kinase receptor-associated protein (STRAP), a ubiquitous WD40 protein, is involved in promoting tumorigenesis through activated MAPK/ERK pathway and pRb phosphorylation. In addition, our previous studies uncovered that STRAP is up-regulated in 60% colon carcinomas and behaves as an oncogene in a TGF-β-dependent or -independent manner. Based on our new finding that STRAP stabilizes intracellular fragment of Notch3 (ICN3) via post-translational modification, we sought to explore the cross-talk between STRAP and Notch pathway in regulating stemness of CCICs. Here, we have generated STRAP knock down (KD) colorectal cancer cell lines (RKO, DLD1, HCT116, HT29, WiDr, SW480, SW620, and LOVO) to show how STRAP abrogates apoptosis by up-regulating CD44+/CD133+ subpopulations of CCICs. Heterozygous Strap KO inhibits tumorigenesis accompanied by the reduction of stemness phenotype (CD44+, Lgr5+) in a colitis-associated mouse model. Western blot assays and immunofluorescene staining demonstrated that the expression of stemness biomarkers (Nanog, Oct4 and Sox2) is decreased in STRAP KD cells. Furthermore, the transcriptional activation of Notch pathway related genes is downregulated in the STRAP KD cells. EpiTect Chip qPCR Arrays profiled that 17 out of 84 genes key to Notch signal transduction had higher activating (H3K4me3) and lower repressive (H3K27me3) histone modifications in WT cells compared to STRAP KD cells. Among these genes, we identified two transcription factors, Hes1 and Hes5, that are transcriptionally regulated by STRAP as demonstrated by luciferase assays and qPCR. We further demonstrated that Hes1 and Hes5 were transcriptionally activated by STRAP through Polycomb Repressive Complex 2 (PRC2)-dependent mechanism. Immunoprecipitation experiments showed that STRAP interacts with the key components, EZH2 and SUZ12 of PRC2 complex in vivo, which antagonizes the histone methyltransferase activity in H3K27 on the promoters of Hes1 and Hes5, resulting in activating these two genes. Of interest, ectopic expression of either Hes1 or Hes5 can rescue the stemness phenotype in STRAP KD cells. Together, our findings provide a novel mechanistic insight into the regulation of CCICs via STRAP-PRC2-Hes1/5 regulatory axis, which represents a potential target for CCICs, thus overcoming cancer chemoresistance and relapse. Citation Format: Lin Jin, Trung Tam Vu, Pran K. Datta. STRAP mediates the stemness of human colorectal cancer cells by epigenetic regulation of Notch pathway. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1709.