Stable stem cell commitment to the adipocyte lineage by inhibition of DNA methylation: role of the BMP-4 gene.
Previous studies showed that exposure of C3H10T1/2 stem cells to bone morphogenetic protein-4 (BMP-4) produced cells that convert into adipocytes at high frequency when treated with differentiation inducers. In the present investigation, an independent approach shows that BMP-4 is required for stable commitment of pluripotent stem cells to the adipocyte lineage. Exposure of proliferating 10T1/2 stem cells to 5-azacytidine, a potent DNA methylation inhibitor, gave rise to a subpopulation of cells that can be cloned and that have the capacity to undergo conversion into adipocytes upon treatment with terminal differentiation inducers. Detailed studies performed with a cloned committed subline, the A33 line, verified stable adipocyte lineage determination in the absence of exogenous BMP-4. Remarkably, this cell line expresses and secretes BMP-4 during proliferation in the same time window that exogenous BMP-4 must be added to naïve 10T1/2 cells to induce maximal adipocyte commitment. Furthermore, exposure of A33 cells to noggin, a naturally occurring BMP-4-binding antagonist, during this critical time window blocks subsequent differentiation. The role of BMP-4 in adipocyte lineage commitment is further strengthened by gene expression profiling of proliferating 10T1/2 stem cells and A33 preadipocytes. These findings revealed changes in the molecular circuitry, specifically coordinated changes in the expression of members of the BMP-4 signaling pathway, that distinguish A33 preadipocytes from uncommitted parental 10T1/2 stem cells. Together, these studies provide compelling evidence for the participation of BMP-4 in adipocyte lineage determination.
Bowers
Robert R
RR
Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA. rbowers4@jhmi.edu
Kim
Jae Woo
JW
Otto
Tamara C
TC
Lane
M Daniel
MD
D017667:Adipocytes / Q000166:cytology* / Q000187:drug effects; D000818:Animals; D001374:Azacitidine / Q000494:pharmacology; D055415:Bone Morphogenetic Protein 4; D019485:Bone Morphogenetic Proteins / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism; D002452:Cell Count; D019070:Cell Lineage / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D019175:DNA Methylation / Q000187:drug effects; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D008648:Mesoderm / Q000166:cytology; D051379:Mice; D010641:Phenotype; D011401:Promoter Regions, Genetic / Q000187:drug effects; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013234:Stem Cells / Q000166:cytology* / Q000187:drug effects; D000099176:Noggin Protein
D001374:Azacitidine; D055415:Bone Morphogenetic Protein 4; D019485:Bone Morphogenetic Proteins; D002352:Carrier Proteins; D012333:RNA, Messenger; D000099176:Noggin Protein; C525809:BMP4 protein, human; C525810:Bmp4 protein, mouse