Role of PPARγ in Regulating a Cascade Expression of Cyclin-dependent Kinase Inhibitors, p18(INK4c) and p21(Waf1/Cip1), during Adipogenesis

A role for PPARγ is demonstrated in mediating the differentiation-dependent cascade expression of cyclin-dependent kinase inhibitors, thereby providing a molecular mechanism coupling growth arrest and adipocyte differentiation.

Abstract

Molecular mechanisms coupling growth arrest and cell differentiation were examined during adipogenesis. Data are presented that document a cascade expression of members of two independent families of cyclin-dependent kinase inhibitors that define distinct states of growth arrest during 3T3-L1 preadipocyte differentiation. Exit from the cell cycle into a pre-differentiation state of post-mitotic growth arrest was characterized by significant increases in p21 and p27. During onset of irreversible growth arrest associated with terminal differentiation, the level of p21 declined with a concomitant, dramatic increase in p18 and a sustained level of p27. The expression of p18 and p21, regulated at the level of protein and mRNA accumulation, was directly coupled to differentiation. Stable cell lines were engineered to express adipogenic transcription factors to examine the active role of trans-acting elements in regulating these cell cycle inhibitors. Ectopic expression of peroxisome proliferator-activated receptor (PPAR) γ in non-precursor fibroblastic cell lines resulted in conversion to adipocytes and a coordinated increase in p18 and p21 mRNA and protein expression in a PPARγ ligand-associated manner. These data demonstrate a role for PPARγ in mediating the differentiation-dependent cascade expression of cyclin-dependent kinase inhibitors, thereby providing a molecular mechanism coupling growth arrest and adipocyte differentiation. Molecular mechanisms coupling growth arrest and cell differentiation were examined during adipogenesis. Data are presented that document a cascade expression of members of two independent families of cyclin-dependent kinase inhibitors that define distinct states of growth arrest during 3T3-L1 preadipocyte differentiation. Exit from the cell cycle into a pre-differentiation state of post-mitotic growth arrest was characterized by significant increases in p21 and p27. During onset of irreversible growth arrest associated with terminal differentiation, the level of p21 declined with a concomitant, dramatic increase in p18 and a sustained level of p27. The expression of p18 and p21, regulated at the level of protein and mRNA accumulation, was directly coupled to differentiation. Stable cell lines were engineered to express adipogenic transcription factors to examine the active role of trans-acting elements in regulating these cell cycle inhibitors. Ectopic expression of peroxisome proliferator-activated receptor (PPAR) γ in non-precursor fibroblastic cell lines resulted in conversion to adipocytes and a coordinated increase in p18 and p21 mRNA and protein expression in a PPARγ ligand-associated manner. These data demonstrate a role for PPARγ in mediating the differentiation-dependent cascade expression of cyclin-dependent kinase inhibitors, thereby providing a molecular mechanism coupling growth arrest and adipocyte differentiation. Adipocytes of white adipose tissue, as well as myocytes from heart and skeletal muscle, represent examples of terminal differentiation whereby the expression of a specialized phenotype is marked by cessation of cell proliferation and the accumulation of cells in the G1 phase of the cell cycle. In mammalian cells, phase transition is regulated by the phosphorylated states of various substrates including the retinoblastoma family proteins which mediate S phase progression (1Weinberg R.A. Cell. 1995; 81: 323-330Abstract Full Text PDF PubMed Scopus (4309) Google Scholar). These substrates are phosphorylated by a dimer complex comprising a regulatory “cyclin” subunit and a catalytic cyclin-dependent kinase (cdk). 1The abbreviations used are: cdk, cyclin-dependent kinase; CKI, cyclin-dependent kinase inhibitor; C/EBP, CCAAT/enhancer-binding protein; PPAR, peroxisome proliferator-activated receptor; TZD, thiazolidinedione; DMEM, Dulbecco's modified Eagle's medium; FBS, fetal bovine serum; MDI, 3-isobutyl-1-methylxanthine, dexamethasone, and insulin; kb, kilobase pair(s); TNFα, tumor necrosis factor α; PPA, proliferating preadipocyte 1The abbreviations used are: cdk, cyclin-dependent kinase; CKI, cyclin-dependent kinase inhibitor; C/EBP, CCAAT/enhancer-binding protein; PPAR, peroxisome proliferator-activated receptor; TZD, thiazolidinedione; DMEM, Dulbecco's modified Eagle's medium; FBS, fetal bovine serum; MDI, 3-isobutyl-1-methylxanthine, dexamethasone, and insulin; kb, kilobase pair(s); TNFα, tumor necrosis factor α; PPA, proliferating preadipocyte Phosphorylating activity of cyclin/cdk complexes is further modulated by cyclin-dependent kinase inhibitors (CKIs), which are grouped into two distinct families based on sequence homology and targets of inhibition (2Sherr C.J. Roberts J.M. Genes Dev. 1995; 9: 1149-1163Crossref PubMed Scopus (3209) Google Scholar). To date, seven CKIs have been identified, including p15INK4b, p16INK4a, p18INK4c, and p19INK4d defining the INK4 family, and p21Cip1, p27Kip1, and p57Kip2, representing the CIP/KIP family. Recent reports have demonstrated that CKI expression is up-regulated during cell differentiation in vitro and in vivo (3Harper J.W. Elledge S.J. Curr. Biol. 1996; 6: 56-64Google Scholar, 4Chellappan S.P. Giordano A. Fisher P.B. Curr. Top. Microbiol. Immunol. 1998; 227: 57-103Crossref PubMed Google Scholar), suggesting that these cell cycle inhibitors may play a universal role in exit from the cell cycle and/or maintenance of the irreversible growth arrest which defines terminal differentiation. Adipocyte differentiation is largely controlled by two families of transcription factors: the CCAAT/enhancer-binding proteins (C/EBPs) and peroxisome proliferator-activated receptors (PPARs) (5Cornelius P. MacDougald O.A. Lane M.D. Annu. Rev. Nutr. 1994; 14: 99-129Crossref PubMed Scopus (572) Google Scholar, 6Brun R.P. Kim J.B. Hu E. Altiok S. Spiegelman B.M. Curr. Opin. Cell Biol. 1996; 8: 826-832Crossref PubMed Scopus (154) Google Scholar, 7Gregoire F.M. Smas C.M. Sul H.S. Physiol. Rev. 1998; 78: 783-809Crossref PubMed Scopus (1846) Google Scholar). Members of the C/EBP family (C/EBPα, C/EBPβ, and CEBPδ) form heterodimers and homodimers via a leucine zipper motif with dimers binding to regulatory elements within target genes via basic DNA binding domains. Ectopic expression of various C/EBPs has been shown to convert non-precursor fibroblastic cell lines into fully differentiated adipocytes (8Freytag S.O. Paielli D.L. Gilbert J.D. Genes Dev. 1994; 8: 1654-1663Crossref PubMed Scopus (392) Google Scholar, 9Wu Z. Xie Y. Bucher N.L.R. Farmer S.R. Genes Dev. 1995; 9: 2350-2363Crossref PubMed Scopus (478) Google Scholar, 10Yeh W.-C. Cao Z. Classon M. McKnight S.L. Genes Dev. 1995; 9: 168-181Crossref PubMed Scopus (809) Google Scholar), whereas genetic knockouts in vitro andin vivo block adipocyte differentiation (11Lin F. Lane M.D. Genes Dev. 1992; 6: 533-544Crossref PubMed Scopus (274) Google Scholar, 12Wang N.D. Finegold M.J. Bradley A. Ou C.N. Abdelsayed S.V. Wilde M.D. Taylor LR Wilson D.R. Darlington G.J. Science. 1995; 269: 1108-1112Crossref PubMed Scopus (835) Google Scholar, 13Moitra J. Mason M.M. Olive M. Krylov D. Gavrilova O. MarcusSamuels B. Feigelman L. Lee E. Aoyama T. Eckhaus M. Reitman M.L. Vinson C. Genes Dev. 1998; 12: 3168-3181Crossref PubMed Scopus (665) Google Scholar). The PPARs (PPARα, PPARδ, and PPARγ) define a family of ligand-activated nuclear hormone receptors that heterodimerize with the retinoid X receptor and bind to specific peroxisome proliferator-responsive elements located within the promoters of target genes. Through utilization of different start sites and alternate splicing, the PPARγ gene gives rise to two isoforms, γ1 and γ2. Tissue distribution of PPARγ2 is highly enriched in adipose tissue, and ectopic expression in various non-precursor cell lines also gives rise to adipocyte differentiation (14Tontonoz P. Graves R.A. Budavari A.I. Erdjument-Bromage H. Lui M. Hu E. Tempst P. Spiegelman B.M. Nucleic Acids Res. 1994; 22: 5628-5634Crossref PubMed Scopus (330) Google Scholar). Although the natural ligand for PPARγ is still under investigation, a synthetic class of specific ligands, called thiazolidinediones (TZDs), greatly enhance transcriptional activity (15Lehmann J.M. Moore L.B. Smith-Oliver Wilson J. Biol. 1995; Full Text Full Text PDF PubMed Scopus Google Scholar). of the molecular of a cascade expression of and by the expression of and which and the expression of genes of the of of the these and elements that adipocyte differentiation has been with the of preadipocyte cell lines 3T3-L1 and that from cells into adipocytes and of white adipose H. O. Cell. Full Text PDF Scopus Google Scholar). a state of growth preadipocyte cell lines to with to a of and to these the cells the cell cycle for a of cell to as is by the of a state of post-mitotic growth to as which has been to for differentiation D.L. S. A. PubMed Scopus Google Scholar). the onset of differentiation, the cells a state of growth arrest that is are to Although reports have the transcriptional of adipocyte specific gene is the molecular in the progression of and the of distinct states of growth arrest that the progression terminal differentiation. In investigation, demonstrate that of differentiation of 3T3-L1 in gene expression representing cell cycle progression that to adipogenic gene expression with exit from the cell cycle. In the data presented document a cascade expression of members of two independent families of CKIs that define distinct states of growth arrest associated with adipogenesis. the expression of p18 and p21 is shown to regulated during the conversion of non-precursor into adipocytes by ectopic expression of the adipogenic transcription providing a molecular mechanism coupling growth arrest and adipocyte differentiation. The cell and under of a was and Z. Xie Y. Bucher N.L.R. Farmer S.R. J. 1998; PubMed Scopus Google Scholar). Stable cell lines PPARγ were by as R.P. P. B.M. J. Spiegelman B.M. Genes Dev. 1996; PubMed Scopus Google Scholar). cells were in Dulbecco's modified Eagle's fetal bovine the cells were with PPARγ2 expression by with as M.L. D. S. A. PubMed Scopus Google Scholar). were and with to proliferating and for was to cells were and to for cells were in 3T3-L1 and cell lines adipogenic transcription factors were to into adipocytes as Lane M.D. J. Biol. Full Text PDF PubMed Google Scholar). cells were in the was to with 3-isobutyl-1-methylxanthine, dexamethasone, and differentiation cells were in and the of was the cells were for the to cells of differentiation with the of to the The to the the of to the was from cell lines with to with cells were in with a and with the phase was with of and The was in with of for and The phase was and with of was in and The was to and were by the of DNA and to the S subunit was used to and of adipocytes from was as D.R. E. L.B. 1998; Scopus (330) Google Scholar). cells were with in and inhibitors, and of the was a to proteins were by and to were with and with the p21 and proliferating cell nuclear and and PPARγ and J. PubMed Scopus Google Scholar). were with and to 3T3-L1 to are to phase of which the of a have molecular mechanisms that may play a role in exit from and/or maintenance of growth arrest associated with terminal differentiation. To the growth and differentiation, characterized in gene expression during cell cycle progression that the of differentiation. were to and to as under was for a from growth to differentiation and to shown in cells to a state of growth as by the of and gene expression to proliferating to differentiation of with and resulted in cell cycle progression with of and gene The of gene and the of at that of these into the cell cycle the to differentiation The of gene expression presented are with in D. J. Biol. Full Text Full Text PDF PubMed Scopus Google and complexes J. Biol. Full Text Full Text PDF PubMed Scopus Google for 3T3-L1 on the of and gene and J.M. M.D. J. 1992; 9: PubMed Scopus Google Scholar, J. Cell. 1992; PubMed Scopus Google Scholar), that of differentiation of these resulted in of cell cycle gene expression that in the of and to cell also the of with adipogenic transcription factor gene of differentiation resulted in of and a that has been shown to independent of protein and by and dexamethasone, Z. McKnight S.L. Genes Dev. PubMed Scopus Google Scholar). Although is still that and may play in the of cell cycle is that cell proliferation the expression of and The of and gene expression during of differentiation that proliferating at during the of adipogenesis. To the growth and differentiation was for the to differentiation and to Data presented in the of growth and differentiation with a in gene expression associated with these independent of differentiation. the onset of and PPARγ with the in gene suggesting that these transcription factors may play a role in coupling growth arrest and adipocyte differentiation. is also to that the for the of differentiation were also for of the to growth and differentiation in the of is at the level and by the is in to skeletal of differentiation to a state of growth and the of as have M.J. J. Cell Google Scholar), that differentiation of preadipocyte cell lines for the coupling of growth arrest and cell differentiation. is well that cell cycle progression is controlled by cyclin/cdk protein activity modulated by and distinct To the of CKIs in coupling of growth arrest and adipocyte differentiation, the gene expression of the seven members of the INK4 and CIP/KIP families of CKIs was examined by was that terminal differentiation was marked by p18 and p21 mRNA on p18 and p21 were further examined at the level of protein expression during the that the growth and differentiation. regulated at the level of gene was also examined to reports CKI to arrest and the well of protein expression M. P. M. Science. 1995; 269: PubMed Scopus Google Scholar, L. Science. 1996; PubMed Scopus Google Scholar). To the of CKI expression during exit from and the onset of irreversible growth cell proteins were a to differentiation and to during two independent The examined protein expression for and the during the and for was from the in and to for mRNA expression as a of expression of and PPARγ was also examined to document the onset of differentiation. shown in protein was in during the of differentiation, to by and the of differentiation. the increase in mRNA and the in protein in a as cells and the S phase of In the protein expression of p21 was in proliferating a to differentiation p21 during of during S phase to with the expression of and and during of differentiation Although the with cell cycle the of p21 protein accumulation as cells the cell cycle as by a significant in the of cells S phase the protein expression of proliferating cell nuclear which to during was during suggesting that the cells a state of growth arrest that was from that during arrest terminal differentiation The expression of p18 to exit from the cell cycle and the onset of adipocyte gene expression with significant protein during of terminal differentiation. p21 and p18 protein was in proliferating reports have that CKI in p21, directly modulated by various and To that the cascade of CKI expression was to molecular of differentiation and to to the were to in the and of tumor necrosis factor has been shown to block the of the phenotype and associated gene expression to cells during the of preadipocyte differentiation. cell were on and in the and of and to for CKI expression of was also examined to the state of adipocyte differentiation. shown in the increase in p21 and p18 protein on and was in the of TNFα, suggesting that the expression of these CKIs was adipocyte differentiation and to of the and to differentiation. was further by the of of on suggesting that the on differentiation was to of To further the of CKI expression terminal differentiation, protein expression was examined in adipocytes and from as under The adipocytes were into and cell to that protein expression in adipocytes was and by protein cell were from 3T3-L1 adipocytes at and of differentiation and to shown in p18 and proteins were in cell to a to that for differentiated 3T3-L1 adipocytes p21 protein was to significant in fully differentiated 3T3-L1 adipocytes in of differentiated C/EBPs and PPARγ as transcription factors for of the The was to these adipogenic transcription factors play a role in regulating the cascade expression of CKIs during adipogenesis. the mRNA accumulation of p21, and was with the expression of and PPARγ the of 3T3-L1 preadipocyte differentiation. was for and to shown in the mRNA expression of in declined and at the of differentiation. Although the in protein that S phase of was by a in mRNA accumulation and the of protein to cell cycle progression to independent of gene expression and to via controlled state of p21 mRNA with protein expression and as cells the state of post-mitotic growth arrest the onset of terminal differentiation was marked by a in p21 protein the mRNA suggesting transcriptional and were to in p21 of p18 demonstrated a and that to with protein expression on and differentiation. Although the of p18 were the onset of terminal differentiation was marked by dramatic in the with have two p18 in the during cell differentiation Y. Hu E. Lee Y. Cell. Biol. 1998; PubMed Scopus Google Scholar, Y. Cell 1998; 9: Google Scholar). these data that differentiation-dependent increases in p18 and p21 protein expression were by in mRNA the accumulation of p18 and p21 with the onset of and PPARγ gene To the of adipogenic transcription factors in the of CKI gene a fibroblastic cell engineered to express and under the of a expression Z. Xie Y. Bucher N.L.R. Farmer S.R. J. 1998; PubMed Scopus Google Scholar). These cells, were in the of which has been shown to the ectopic expression of was from the growth at and at the growth was with differentiation with the TZD, the of the were to used for 3T3-L1 differentiation under was at and of differentiation and to in ectopic expression of and in the of and TZD, to the expression of PPARγ and genes with the onset of adipocyte differentiation was the accumulation of p18 and p21 to to in fully differentiated 3T3-L1 CKI mRNA accumulation in the of which has been shown to in Z. Xie Y. Bucher N.L.R. Farmer S.R. Genes Dev. 1995; 9: 2350-2363Crossref PubMed Scopus (478) Google Scholar, 10Yeh W.-C. Cao Z. Classon M. McKnight S.L. Genes Dev. 1995; 9: 168-181Crossref PubMed Scopus (809) Google Scholar). is to that the role of as adipogenic transcription factor in by the ectopic expression of and/or To to the molecular mechanism for CKI mRNA accumulation, these engineered were under various to gene expression and to and was of and the of in B. the ectopic and in the of to and resulted in a increase in p21 mRNA that was in cells the C/EBPs in the of differentiation Although of cells the C/EBPs resulted in PPARγ and gene expression significant p18 and p21 mRNA accumulation and of the adipocyte phenotype was cells under to PPARγ gene expression in the of ligand specific for PPARγ accumulation of p18 and p21 with ligand-activated PPARγ and C/EBP to the the adipogenic transcription factor and CKI gene expression by a to fibroblastic cell lines the of The expression the for PPARγ used in by B. M. was characterized for in active protein in P. Hu E. J. Spiegelman B.M. Cell. 1994; Full Text PDF PubMed Scopus Google Scholar). and PPARγ were into that were used to and as under the cell lines were to and to with in the of the TZD, of from and PPARγ at and of differentiation is in A. from differentiated 3T3-L1 adipocytes was for The was from the as the differentiation including fibroblastic cell lines PPARγ gene expression and cells data of the cells lines demonstrated a significant increase in p18 and p21 with the of the adipocyte in to cells, was in these and was regulated in a with adipocyte differentiation. and are as conversion cells are to of differentiation. To the of PPARγ adipogenic the PPARγ the were differentiated for in the and of was and the is in B. to the differentiation to cells resulted in a increase in and p21 and that was with to cells the and that cells to the differentiation the expression of p18 and p21 independent of level of expression was to a of the to differentiation and/or the adipocyte conversion of cells that the differentiation In to the PPARγ to as marked by independent of and the mRNA for p18 and p21 was and in cells in with expression and adipocyte differentiation. The increase in mRNA was cells with and cells differentiated in the of PPARγ ligand a was for p18 and p21 These data the that the increase in CKI mRNA was to to in cell as and cells were to differentiation The increase in p18 and p21 with of cells and to of which under these transcriptional and mechanisms were in the of p21 and during 3T3-L1 adipocyte differentiation. To adipocyte conversion by PPARγ also to CKI protein PPARγ and the were to with in the and of cell proteins were the of differentiation, and the of are in C. and of p18 and p21 protein expression were to that for 3T3-L1 adipocytes with p21 p18 expression by The of expression was in cells, which to adipocyte and cells, which as the of on adipocyte differentiation for protein was also that protein in cells, which of p18 and p21, suggesting that adipocyte differentiation resulted in the accumulation of cells in the G1 phase of the cell cycle. Although the specific of these CKIs during has to to that of is to cell proliferation to S phase transition during the onset of terminal differentiation. a molecular mechanism coupling growth arrest and adipocyte differentiation. demonstrate a in gene expression mediating the of growth and differentiation and that growth arrest with the expression of adipogenic transcription and data are presented a cascade of CKI expression that distinct states of growth arrest associated with adipogenesis. the differentiation-dependent of p18 and p21 is regulated at the level of mRNA and protein expression non-precursor are to adipocytes by the expression of PPARγ in a ligand-associated these data demonstrate that transcription factors that mediate also the expression of cell cycle inhibitors providing a molecular mechanism coupling these during exit from the cell cycle and the irreversible growth arrest of terminal differentiation. Data presented in and in a Y. Cell 1998; 9: Google a cascade expression of members of two independent families of CKIs during the of adipogenesis. in the protein expression of p21, and p18 defines states of growth arrest associated with post-mitotic growth and the onset of terminal differentiation, The and of expression that CKIs may play specific and in coupling growth arrest and cell differentiation. is to the of the protein and gene expression during reports have that protein under of and with the onset of cell cycle during cell and to with the onset of growth arrest C.J. Science. 1996; PubMed Scopus Google Scholar). The of to presented in to a of cell cycle and to the of into and exit from the cell cycle associated with Although the increase in is with of a the expression of with p21 during exit from the cell cycle and with p18 during terminal differentiation may represent a role for CKIs during distinct states of growth In the expression of p21 and may to the and of growth arrest The also that of p21 and may play a and/or regulatory role for adipocyte differentiation. In of the of cell differentiation has been shown to independent of p21 and were suggesting that CKI expression may for cell differentiation M. Lee M. M. Genes Dev. 1996; PubMed Scopus Google Scholar). the that ectopic expression of p21 of to differentiation the of CKI in regulating cell differentiation independent of growth arrest J. Y. T. H. Y. Y. Cell. Biol. PubMed Scopus Google Scholar). the and of CKI expression during may and specific specific to distinct states of growth arrest and different of adipocyte differentiation. in fully differentiated adipocytes in in that the expression of p21 increases during the of adipocyte differentiation. The of p21, which with the G1 phase of is with and of p21 during of cell cycle progression J. M.D. J.M. C. H.S. A. E. Genes Dev. PubMed Scopus Google Scholar). In the of p21 directly with from and is on the differentiation for that of and in the of differentiation two independent of p21 protein suggesting the of two independent regulatory mechanisms based on the progression of proliferation differentiation. The in p21 in the of CKIs during of is with reports a expression of p21 during differentiation in Y. Biol. Cell. 1996; PubMed Scopus Google and during in vivo M.J. A. Kim Lee J. Cell. 1998; Full Text PDF PubMed Scopus Google Scholar). The of p21, and with proteins mediating growth and differentiation is to specific of these CKIs during adipogenesis. also from fibroblastic cell lines regulating the expression of PPARγ by two independent mechanisms and with two different for that for a role of adipogenic transcription factors in regulating CKIs at the level of mRNA and protein in cell lines that were PPARγ for adipogenic gene expression were also for of p18 and p21, suggesting a regulatory role for PPARγ at during the of adipogenesis. The of PPARγ in the differentiation to the of p18 and p21 has to is that of p18 during at the level of gene as in protein expression with in mRNA In inhibition of protein by in cells PPARγ p18 mRNA accumulation suggesting that proteins of PPARγ are to in mediating p18 gene The of transcription factor is with the the expression of PPARγ and with p21 were as inhibition of protein independent of PPARγ to a dramatic increase in p21 mRNA based on a sequence in the of p21 and the of p21 and PPARγ is that CKI is directly regulated at the level of gene expression by of p21 protein expression during is to complex with in the and of protein expression in mRNA Although the data presented demonstrate regulatory role for PPARγ in the of p21 during adipocyte differentiation, is to that p21 protein expression presented in with the expression of a role for has been shown in the of p21 in and cells at the level of protein Wilde M. Finegold M.J. Darlington G.J. Cell. Biol. PubMed Google Scholar, Wilde M. M. Darlington G.J. Genes Dev. 1996; PubMed Scopus Google Scholar). the that of p21 protein in regulated by during adipogenesis. The that ectopic expression of PPARγ also in the expression of shown and D. J. Biol. Full Text Full Text PDF PubMed Scopus Google the of a cascade of these transcription factors in the of p21 at the level of gene expression and protein reports have a of these transcription factors in the of of the to that the complex of CKI expression during also the of PPARγ and the and mechanisms of CKI expression by these and transcription factors during are under have that McKnight S.L. Science. PubMed Scopus (572) Google and PPARγ S. M. Spiegelman B.M. Genes Dev. PubMed Scopus Google Scholar), the growth of various proliferating fibroblastic cell Although the data presented that growth arrest is coupled to adipocyte differentiation the expression of that growth arrest mechanisms independent of these cell cycle inhibitors may also by adipogenic transcription factors under that may adipogenesis. has been that the C.M. Cell. Biol. 1996; PubMed Google Scholar), and that PPARγ cell cycle by inhibition of binding activity via of the protein S. M. Spiegelman B.M. Genes Dev. PubMed Scopus Google Scholar). is that various of adipocyte gene expression may independent and/or growth arrest mechanisms as a to terminal differentiation. the of CKIs during and the complex of adipogenic transcription factors in regulating expression a of the of adipocyte proliferation coupling of growth arrest and cell differentiation. are to p18 and and Elledge Lee and Spiegelman PPARγ and for with also for and on 3T3-L1 and for with