title: 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.affiliation: From the Department of Oncologic Sciences, Mitchell Cancer Institute, University of South Alabama, Mobile, Alabama 36604 and.
authors.forename: Kaushlendra
authors.initials: K
authors.lastname: Tripathi
authors.affiliation: From the Department of Oncologic Sciences, Mitchell Cancer Institute, University of South Alabama, Mobile, Alabama 36604 and.
authors.forename: Chinnadurai
authors.initials: C
authors.lastname: Mani
authors.affiliation: From the Department of Oncologic Sciences, Mitchell Cancer Institute, University of South Alabama, Mobile, Alabama 36604 and.
authors.forename: Reagan
authors.initials: R
authors.lastname: Barnett
authors.affiliation: From the Department of Oncologic Sciences, Mitchell Cancer Institute, University of South Alabama, Mobile, Alabama 36604 and.
authors.forename: Sriram
authors.initials: S
authors.lastname: Nalluri
authors.affiliation: From the Department of Oncologic Sciences, Mitchell Cancer Institute, University of South Alabama, Mobile, Alabama 36604 and.
authors.forename: Lavanya
authors.initials: L
authors.lastname: Bachaboina
authors.affiliation: From the Department of Oncologic Sciences, Mitchell Cancer Institute, University of South Alabama, Mobile, Alabama 36604 and.
authors.forename: Rodney P
authors.initials: RP
authors.lastname: Rocconi
authors.affiliation: the Department of Dermatology, University of Alabama at Birmingham, Birmingham, Alabama 35294.
authors.forename: Mohammed
authors.initials: M
authors.lastname: Athar
authors.affiliation: From the Department of Oncologic Sciences, Mitchell Cancer Institute, University of South Alabama, Mobile, Alabama 36604 and.
authors.forename: Laurie B
authors.initials: LB
authors.lastname: Owen
authors.affiliation: From the Department of Oncologic Sciences, Mitchell Cancer Institute, University of South Alabama, Mobile, Alabama 36604 and kpalle@health.southalabama.edu.
authors.forename: Komaraiah
authors.initials: K
authors.lastname: Palle
mesh_terms: D000970:Antineoplastic Agents / Q000737:chemistry; D051023:BH3 Interacting Domain Death Agonist Protein / Q000378:metabolism*; D001665:Binding Sites; D002166:Camptothecin / Q000737:chemistry; D002453:Cell Cycle; D045744:Cell Line, Tumor; D002470:Cell Survival; D000071877:Checkpoint Kinase 1; D004260:DNA Repair; D015536:Down-Regulation; D004351:Drug Resistance; D005347:Fibroblasts / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D011494:Protein Kinases / Q000378:metabolism*; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D051716:Replication Protein A / Q000378:metabolism; D016196:S Phase; D015398:Signal Transduction; D059004:Topoisomerase I Inhibitors / Q000737:chemistry / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*; D000071676:Zinc Finger Protein GLI1
chemical_list: D000970:Antineoplastic Agents; D051023:BH3 Interacting Domain Death Agonist Protein; C494814:BID protein, human; C400551:GLI1 protein, human; D034741:RNA, Small Interfering; C497167:RPA1 protein, human; D051716:Replication Protein A; D059004:Topoisomerase I Inhibitors; D014157:Transcription Factors; D000071676:Zinc Finger Protein GLI1; D011494:Protein Kinases; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1; D002166:Camptothecin
keywords: Cancer Biology; Cell Cycle; Checkpoint Control; Chemoresistance; DNA Damage; DNA Damage Response; DNA Repair; DNA Replication; DNA Topoisomerase; Hedgehog Signaling Pathway