title: FOXO1 modulates osteoblast differentiation.
abstract: Forkhead box O1 (FOXO1) is upregulated during bone formation and in response to stimulation by bone morphogenetic proteins. Studies presented here examined the functional role of FOXO1 in a well defined culture system in which pre-osteoblastic cells undergo terminal differentiation in vitro. Mineralizing cultures of MC3T3-E1 cells were examined with or without FOXO1 knockdown by RNAi. Normal cells show the upregulation of FOXO1 and RUNX2 DNA binding activity, alkaline phosphatase activity, and mRNA levels of FOXO1, RUNX2, type 1 collagen, osteocalcin and MMP13 during formation of mineralizing nodules. In FOXO1 depleted cells each of these measurements was significantly reduced compared to values in control cells transfected with scrambled siRNA (P<0.05). Depletion of FOXO1 also reduced the number of mineralized nodules formed. Moreover, chromatin immunoprecipitation assays revealed a direct interaction of FOXO1 with the RUNX2 promoter. Overexpression of FOXO1 reduced the MC3T3-E1 cell number and the number of PCNA positive cells with little effect on apoptosis. These findings indicate that FOXO1 plays an important role in promoting osteoblast differentiation and suppressing proliferation in differentiating cells.
authors.affiliation: Schulich School of Medicine & Dentistry, University of Western Ontario, London, ON, Canada. msiqueir@uwo.ca
authors.forename: Michelle F
authors.initials: MF
authors.lastname: Siqueira
authors.forename: Stephen
authors.initials: S
authors.lastname: Flowers
authors.forename: Rupa
authors.initials: R
authors.lastname: Bhattacharya
authors.forename: Dan
authors.initials: D
authors.lastname: Faibish
authors.forename: Yugal
authors.initials: Y
authors.lastname: Behl
authors.forename: Darrell N
authors.initials: DN
authors.lastname: Kotton
authors.forename: Lou
authors.initials: L
authors.lastname: Gerstenfeld
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Moran
authors.forename: Dana T
authors.initials: DT
authors.lastname: Graves
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D002113:Calcification, Physiologic; D002454:Cell Differentiation; D002460:Cell Line; D049109:Cell Proliferation; D050659:Core Binding Factor Alpha 1 Subunit / Q000235:genetics / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D000071161:Forkhead Box Protein O1; D051858:Forkhead Transcription Factors / Q000235:genetics / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D020287:In Situ Nick-End Labeling; D016086:Lentivirus / Q000235:genetics; D051379:Mice; D020411:Oligonucleotide Array Sequence Analysis; D010006:Osteoblasts / Q000166:cytology* / Q000378:metabolism; D015675:Osteocalcin / Q000235:genetics / Q000378:metabolism; D010012:Osteogenesis / Q000235:genetics; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D015854:Up-Regulation / Q000235:genetics
chemical_list: D015415:Biomarkers; D050659:Core Binding Factor Alpha 1 Subunit; D000071161:Forkhead Box Protein O1; D051858:Forkhead Transcription Factors; C467200:Foxo1 protein, mouse; D018809:Proliferating Cell Nuclear Antigen; D012333:RNA, Messenger; D034741:RNA, Small Interfering; C493716:Runx2 protein, mouse; D015675:Osteocalcin; D004247:DNA