Abstract 4699A: Hedgehog signaling (HH/Gli) transcriptionally regulates hTERT gene expression in human cancer cells

Abstract

Abstract Tightly coordinated Hedgehog (HH) signaling is critical during normal embryonic development. Aberrant HH signaling is involved in driving cell proliferation in multiple human cancers. Classical HH signaling involves the interaction of the soluble HH ligands with their receptor, Patched (Ptch), thereby releasing a transmembrane protein Smoothened (Smo) from Ptch-mediated inhibition. Smo then activates the Gli family of transcription factors that regulate HH target genes. We have previously reported that HH signaling provides essential survival support to human colon cancer cells and inhibition of this signal induces DNA damage and extensive cell death. However the downstream targets that govern the regulation of cell survival by HH/Gli in cancer cells are not completely defined. This study demonstrates that the Gli proteins transcriptionally regulate the expression of human telomerase reverse transcriptase (hTERT) gene in human cancer cells. hTERT is a known regulator of telomere homeostasis, which determines the replicative potential and hence the life span of cells. Suppression of both Gli1 and Gli2 functions by exogenous expression of a C-terminus truncated Gli3 repressor mutant (Gli3R), or by GANT61, a pharmacological inhibitor of Gli1 and Gli2 activity, reduced hTERT protein expression over a period of 72 hr in human colon, prostate and brain cancer cell lines. Further, exogenously expressed Gli2 significantly increased hTERT protein expression in human colon cancer cell lines. Exposure to GANT61 also inhibited hTERT mRNA expression within 24 hr in human colon cancer cell lines. Insilico analysis of the hTERT promoter revealed 7 putative Gli binding sites suggesting a transcriptional mode of regulation of hTERT expression by Gli2. Chromatin immunoprecipitation with Gli2 antibody precipitated fragments of the hTERT promoter in human colon cancer cell lines, indicating a direct interaction between Gli2 and the hTERT promoter. The binding between Gli2 and hTERT promoter was significantly reduced upon exposure to GANT61. Further, overexpression of the wild type hTERT cDNA in cancer cells prevented the cytotoxic effects of blocking the HH signaling pathway with GANT61. These findings demonstrate hTERT to be a direct transcriptional target and a critical mediator of the HH/Gli signaling pathway, and identify a previously unknown role of the HH/Gli axis in regulating the replication potential of cancer cells. These findings are of significance in understanding important regulatory mechanisms that determine the role of HH/Gli signaling in cancer cell survival. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4699A. doi:1538-7445.AM2012-4699A