Ras Promotes Growth by Alternative Splicing-Mediated Inactivation of the KLF6 Tumor Suppressor in Hepatocellular Carcinoma

Steven Yea, Goutham Narla, Xiao Zhao, Rakhi Garg, Sigal Tal-Kremer, Eldad Hod, Augusto Villanueva, Johnny Loke, Mirko Tarocchi, Kunihara Akita, Senji Shirasawa, Takehiko Sasazuki, John A. Martignetti, Josep M. Llovet, Scott L. Friedman

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Abstract

Background & Aims: Hepatocellular carcinoma (HCC) is the fifth most prevalent cancer worldwide and the third most lethal. Dysregulation of alternative splicing underlies a number of human diseases, yet its contribution to liver cancer has not been explored fully. The Krüppel-like factor 6 (KLF6) gene is a zinc finger transcription factor that inhibits cellular growth in part by transcriptional activation of p21. KLF6 function is abrogated in human cancers owing to increased alternative splicing that yields a dominant-negative isoform, KLF6 splice variant 1 (SV1), which antagonizes full-length KLF6-mediated growth suppression. The molecular basis for stimulation of KLF6 splicing is unknown. Methods: In human HCC samples and cell lines, we functionally link oncogenic Ras signaling to increased alternative splicing of KLF6 through signaling by phosphatidylinositol-3 kinase and Akt, mediated by the splice regulatory protein ASF/SF2. Results: In 67 human HCCs, there is a significant correlation between activated Ras signaling and increased KLF6 alternative splicing. In cultured cells, Ras signaling increases the expression of KLF6 SV1, relative to full-length KLF6, thereby enhancing proliferation. Abrogation of oncogenic Ras signaling by small interfering RNA (siRNA) or a farnesyl-transferase inhibitor decreases KLF6 SV1 and suppresses growth. Growth inhibition by farnesyl-transferase inhibitor in transformed cell lines is overcome by ectopic expression of KLF6 SV1. Down-regulation of the splice factor ASF/SF2 by siRNA increases KLF6 SV1 messenger RNA levels. KLF6 alternative splicing is not coupled to its transcriptional regulation. Conclusions: Our findings expand the role of Ras in human HCC by identifying a novel mechanism of tumor-suppressor inactivation through increased alternative splicing mediated by an oncogenic signaling cascade.

Original languageEnglish (US)
Pages (from-to)1521-1531
Number of pages11
JournalGastroenterology
Volume134
Issue number5
DOIs
Publication statusPublished - May 2008
Externally publishedYes

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ASJC Scopus subject areas

  • Hepatology
  • Gastroenterology

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