Gli1 protein regulates the S-phase checkpoint in tumor cells via Bid protein, and its inhibition sensitizes to DNA topoisomerase 1 inhibitors.

Abstract

Aberrant expression of hedgehog molecules, particularly Gli1, is common in cancers of many tissues and is responsible for their aggressive behavior and chemoresistance. Here we demonstrate a novel and tumor-specific role for aberrant Gli1 in the regulation of the S-phase checkpoint that suppresses replication stress and resistance to chemotherapy. Inhibition of Gli1 in tumor cells induced replication stress-mediated DNA damage response, attenuated their clonogenic potential, abrogated camptothecin (CPT)-induced Chk1 phosphorylation, and potentiated its cytotoxicity. However, in normal fibroblasts, Gli1 siRNAs showed no significant changes in CPT-induced Chk1 phosphorylation. Further analysis of ataxia telangiectasia and Rad3-related protein (ATR)/Chk1 signaling cascade genes in tumor cells revealed an unexpected mechanism whereby Gli1 regulates ATR-mediated Chk1 phosphorylation by transcriptional regulation of the BH3-only protein Bid. Consistent with its role in DNA damage response, Bid down-regulation in tumor cells abolished CPT-induced Chk1 phosphorylation and sensitized them to CPT. Correspondingly, Gli1 inhibition affected the expression of Bid and the association of replication protein A (RPA) with the ATR- interacting protein (ATRIP)-ATR complex, and this compromised the S-phase checkpoint. Conversely, complementation of Bid in Gli1-deficient cells restored CPT-induced Chk1 phosphorylation. An in silico analysis of the Bid promoter identified a putative Gli1 binding site, and further studies using luciferase reporter assays confirmed Gli1-dependent promoter activity. Collectively, our studies established a novel connection between aberrant Gli1 and Bid in the survival of tumor cells and their response to chemotherapy, at least in part, by regulating the S-phase checkpoint. Importantly, our data suggest a novel drug combination of Gli1 and Top1 inhibitors as an effective therapeutic strategy in treating tumors that expresses Gli1.

• Authors: Kaushlendra Tripathi, Chinnadurai Mani, Reagan Barnett, Sriram Nalluri, Lavanya Bachaboina, Rodney P Rocconi, Mohammed Athar, Laurie B Owen, Komaraiah Palle
• Journal: The Journal of biological chemistry (J Biol Chem) Vol. 289 Issue 45 Pg. 31513-25 (Nov 07 2014) ISSN: 1083-351X [Electronic] United States
• PMID: 25253693 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't)
• Copyright: © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

Chemical References

• Antineoplastic Agents
• BH3 Interacting Domain Death Agonist Protein
• BID protein, human
• GLI1 protein, human
• RNA, Small Interfering
• RPA1 protein, human
• Replication Protein A
• Topoisomerase I Inhibitors
• Transcription Factors
• Zinc Finger Protein GLI1
• Protein Kinases
• CHEK1 protein, human
• Checkpoint Kinase 1
• Camptothecin

Topics

• Antineoplastic Agents (chemistry)
• BH3 Interacting Domain Death Agonist Protein (metabolism)
• Binding Sites
• Camptothecin (chemistry)
• Cell Cycle
• Cell Line, Tumor
• Cell Survival
• Checkpoint Kinase 1
• DNA Repair
• Down-Regulation
• Drug Resistance
• Fibroblasts (metabolism)
• Gene Expression Regulation, Neoplastic
• Humans
• Phosphorylation
• Promoter Regions, Genetic
• Protein Kinases (metabolism)
• RNA Interference
• RNA, Small Interfering (metabolism)
• Replication Protein A (metabolism)
• S Phase
• Signal Transduction
• Topoisomerase I Inhibitors (chemistry, metabolism)
• Transcription Factors (metabolism)
• Zinc Finger Protein GLI1