Intestine-specific homeobox (ISX) upregulates E2F1 expression and related oncogenic activities in HCC.

// Shen-Nien Wang 1, 2, * , Li-Ting Wang 2, * , Ding-Ping Sun 3, 4, * , Chee-Yin Chai 5 , Edward Hsi 6 , Hsing-Tao Kuo 7, 8 , Kazunari K. Yokoyama 2, 9, 10, 11, 12, 13 , Shih-Hsien Hsu 2, 13 1 Division of Hepatobiliary Surgery, Department of Surgery, Faculty of Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan 2 Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan 3 Division of General Surgery, Department of Surgery, Chi-Mei Medical Center, Tainan, Taiwan 4 Department of Food Science and Technology, Chia Nan University of Pharmacy and Science, Tainan, Taiwan 5 Department of Pathology, Faculty of Medicine, College of Medicine, Kaohsiung, Taiwan 6 Department of Genome Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan 7 Department of Internal Medicine, Division of Hepatogastroenterology, Chi-Mei Medical Center, Tainan, Taiwan 8 Department of Senior Citizen Service Management, Chia Nan University of Pharmacy & Science, Tainan, Taiwan 9 Research Center for Stem Cell Research, Kaohsiung Medical University, Kaohsiung, Taiwan 10 Center for Environmental Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan 11 Department of Molecular Preventive Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan 12 Faculty of Science and Engineering, Tokushima Bunri University, Sanuki, Japan 13 Center of Infectious Disease and Cancer Research, Kaohsiung Medical University, Kaohsiung, Taiwan * These authors have contributed equally to this work Correspondence to: Shih-Hsien Hsu, e-mail: jackhsu@kmu.edu.tw Kazunari K. Yokoyama, e-mail: kazu@kmu.edu.tw Hsing-Tao Kuo, e-mail: kuohsu2003@yahoo.com.tw Keywords: cyclin D1 , DP1, E2F1 , hepatocellular carcinoma (HCC), ISX Received: October 05, 2015     Accepted: April 16, 2016     Published: May 09, 2016 ABSTRACT Intestine-specific homeobox ( ISX ), a newly identified proto-oncogene, is involved in cell proliferation and progression of hepatocellular carcinoma (HCC). However, the underlying mechanisms linking gene expression and tumor formation remain unclear. In this study, we found that ISX transcriptionally activated E2F transcription factor 1 ( E2F1 ) and associated oncogenic activity by directly binding to the E2 site of its promoter. Forced expression of ISX increased the expression of and phosphorylated the serine residue at position 332 of E2F1 , which may be translocated into the nucleus to form the E2F1 –DP-1 complex, suggesting that the promotion of oncogenic activities of the ISX–E2F1 axis plays a critical role in hepatoma cells. Coexpression of ISX and E2F1 significantly promoted p53 and RB-mediated cell proliferation and anti-apoptosis, and repressed apoptosis and autophagy. In contrast, short hairpin RNAi-mediated attenuation of ISX and E2F1 decreased cell proliferation and malignant transformation, respectively, in hepatoma cells in vitro and in vivo . The mRNA expression of E2F1 and ISX in 238 paired specimens from human HCC patients, and the adjacent, normal tissues exhibited a tumor-specific expression pattern which was highly correlated with disease pathogenesis, patient survival time, progression stage, and poor prognosis. Therefore, our results indicate that E2F1 is an important downstream gene of ISX in hepatoma progression.