Abstract 4029: Sox9 functions downstream of Hedgehog-Gli1 in epithelial transformation

Abstract

Abstract The Sonic Hedgehog (Shh) signaling pathway plays critical roles in cell fate specification during embryonic and fetal development and in adult tissues. The extracellular morphogen, Shh, signals through Gli family zinc finger transcription factors to regulate the expression of downstream target genes. Constitutive Shh-Gli signaling is implicated in the pathogenesis of several human cancers, including cutaneous basal cell carcinoma (BCC), prostate cancer and medulloblastoma. How Gli can signal through downstream genes to induce malignant transformation remains poorly understood. In this work, we performed microarray analysis to reveal the gene expression profile after tetracycline-regulated induction of Gli1 for 3 or 6 hours in cultured epithelial cells, identifying multiple known Shh target genes. Among the most rapidly induced genes was Sox9, a member of SRY family of transcription factors. Sox9 is essential for male sex determination and is a master regulator of chondrogenesis. Sox9 is also well established as a Shh-Gli target gene, but how it functions in Shh-Gli signaling is not well understood. We found that Sox9 mRNA and protein levels are greatly increased after short-term or long-term Gli1 expression in a rat culture model of epithelial cells, termed RK3E, and in mouse skin. As shown by reporter assays, Sox9 transcriptional activity was increased by approximately 18-fold in RK3E cells transformed by Gli1. To enable knockdown studies in rat cells, we defined the full-length rat Sox9 coding sequence. Subsequently, suppression of rat Sox9 by siRNA indicated an essential role for this Gli-regulated gene in malignant transformation of RK3E in vitro. Specifically, shRNA plasmids that suppress Sox9 could dramatically reduced the transforming activity of Gli1. In contrast, there was little effect on other transforming oncoproteins including H-Ras and ErbB2. These results establish Sox9 induction as an early response to Gli1, and demonstrate that induction of Sox9 is essential for transformation by Gli1. Further characterization of the Sox9 knockdown phenotype is in progress and will be presented. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4029.