E2F1 and TFDP1 Regulate PITX1 Expression in Normal and Osteoarthritic Articular Chondrocytes.
We previously reported a loss-of-PITX1 expression in patients suffering of knee/hip osteoarthritis (OA). Search for the mechanism underlying this event led us to discover that PITX1 repression was triggered by the aberrant nuclear accumulation of Prohibitin (PHB1), an E2F1 co-repressor, in OA articular chondrocytes. In the current study, we assessed in details the involvement of E2F transcription factors in regulating PITX1 expression. We also analyzed other genes that are similarly regulated by E2F in regard to osteoarthritis. The transcriptional regulation of the PITX1 promoter by E2F1 was analyzed with the luciferase reporter assay, and chromatin immunoprecipitation assays, which confirmed direct E2F1-PITX1 interactions. The probable binding sites for E2F1 in the PITX1 promoter were identified by DNA pulldown experiments. In silico and in vitro analyses show that the PITX1 proximal promoter region contains 2 specific sequences that are bound by E2F1. Overexpression of E2F1 enhances PITX1 promoter activity and mRNA transcription. In primary control and osteoarthritis chondrocytes, real time RT-PCR was used to measure the mRNA expression levels of candidate genes under E2F1 transcriptional control. Transcription Factor Dp-1 (TFDP1) knockdown experiments confirmed that the E2F1-TFDP1 complex regulates PITX1. Knockdown of TFDP1, an E2F1 dimerization partner, inhibits the activating effect of E2F1 and reduces both PITX1 promoter activity and mRNA transcription. Real time RT-PCR results reveal reduced expression of TFDP1 and a similar downregulation of their targets PITX1, BRCA1, CDKN1A, and RAD51 in mid-stage OA chondrocytes. Collectively, our data define a previously uncharacterized role for E2F1 and TFDP1 in the transcriptional regulation of PITX1 in articular chondrocytes. Additional E2F1 targets may be affected in OA pathogenesis.
Viscogliosi Laboratory in Molecular Genetics of Musculoskeletal Diseases, Sainte-Justine University Hospital Research Center, Montréal, Québec, Canada.|Department of Biochemistry and Molecular Medicine, Faculty of Medicine, Université de Montréal, Montréal, Québec, Canada.
Martin
M
Pellicelli
Viscogliosi Laboratory in Molecular Genetics of Musculoskeletal Diseases, Sainte-Justine University Hospital Research Center, Montréal, Québec, Canada.|Department of Biochemistry and Molecular Medicine, Faculty of Medicine, Université de Montréal, Montréal, Québec, Canada.
Cynthia
C
Picard
Viscogliosi Laboratory in Molecular Genetics of Musculoskeletal Diseases, Sainte-Justine University Hospital Research Center, Montréal, Québec, Canada.
DaShen
D
Wang
Orthopedic Division, Maisonneuve-Rosemont Hospital, Montréal, Québec, Canada and Department of Surgery, Faculty of Medicine, Université de Montréal, Montréal, Québec, Canada.
Patrick
P
Lavigne
Viscogliosi Laboratory in Molecular Genetics of Musculoskeletal Diseases, Sainte-Justine University Hospital Research Center, Montréal, Québec, Canada.|Department of Biochemistry and Molecular Medicine, Faculty of Medicine, Université de Montréal, Montréal, Québec, Canada.|Department of Stomatology, Faculty of Dentistry, Université de Montréal, Montréal, Québec, Canada.
Alain
A
Moreau
D000328:Adult; D001483:Base Sequence; D019902:Chondrocytes / Q000378:metabolism*; D050687:E2F1 Transcription Factor / Q000378:metabolism*; D005260:Female; D005786:Gene Expression Regulation; D055785:Gene Knockdown Techniques; D006706:Homeostasis; D006801:Humans; D007719:Knee Joint / Q000473:pathology; D008297:Male; D008875:Middle Aged; D010003:Osteoarthritis / Q000378:metabolism*; D051761:Paired Box Transcription Factors / Q000235:genetics* / Q000378:metabolism*; D000091402:Prohibitins; D011401:Promoter Regions, Genetic / Q000235:genetics; D020218:Response Elements / Q000235:genetics; D050680:Transcription Factor DP1 / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D015854:Up-Regulation
D050687:E2F1 Transcription Factor; C493753:E2F1 protein, human; C000717328:PHB protein, human; D051761:Paired Box Transcription Factors; D000091402:Prohibitins; D050680:Transcription Factor DP1; C101485:homeobox protein PITX1