Reactive oxygen species facilitate adipocyte differentiation by accelerating mitotic clonal expansion.
Growth-arrested 3T3-L1 preadipocytes rapidly express CCAAT/enhancer-binding protein-beta (C/EBPbeta) upon hormonal induction of differentiation. However, the DNA binding activity of C/EBPbeta is not activated until the cells synchronously reenter S phase during the mitotic clonal expansion (MCE) phase of differentiation. In this period, C/EBPbeta is sequentially phosphorylated by MAPK and glycogen synthase kinase-3beta, inducing C/EBPbeta DNA binding activity and transcription of its target genes. Because the DNA binding activity of C/EBPbeta is further enhanced by oxidation in vitro, we investigated how redox state affects C/EBPbeta DNA binding and MCE during adipogenesis. When 3T3-L1 cells were treated with H(2)O(2) and hormonal stimuli, differentiation was accelerated with increased expression of peroxisome proliferator-activated receptor gamma. Interestingly, cell cycle progression (S to G(2)/M phase) was markedly enhanced by H(2)O(2), whereas antioxidants caused an S phase arrest during the MCE. H(2)O(2) treatment resulted in the early appearance of a punctate pattern observed by immunofluorescent staining of C/EBPbeta, which is a hallmark for C/EBPbeta binding to regulatory elements, whereas a short antioxidant treatment rapidly dispersed the centromeric localization of C/EBPbeta. Consistently, reactive oxygen species production was increased during 3T3-L1 differentiation. Our results indicate that redox-induced C/EBPbeta DNA binding activity, along with the dual phosphorylation of C/EBPbeta, is required for the MCE and terminal differentiation of adipocytes.
Lee
Haemi
H
Department of Biochemistry and Molecular Biology, Center for Chronic Metabolic Disease Research, Institute of Genetic Science, Yonsei University College of Medicine, Brain Korea 21 Project for Medical Science, Yonsei University, Seoul 120-752, Korea.
Lee
Yoo Jeong
YJ
Choi
Hyeonjin
H
Ko
Eun Hee
EH
Kim
Jae-Woo
JW
D041721:3T3-L1 Cells; D017667:Adipocytes / Q000166:cytology / Q000502:physiology*; D000818:Animals; D000975:Antioxidants / Q000378:metabolism; D022782:CCAAT-Enhancer-Binding Protein-beta / Q000235:genetics / Q000378:metabolism; D002454:Cell Differentiation / Q000502:physiology*; D019925:Cyclin A / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000378:metabolism; D051379:Mice; D008938:Mitosis / Q000502:physiology*; D016877:Oxidants / Q000378:metabolism; D010084:Oxidation-Reduction; D011485:Protein Binding; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism*; D000077185:Resveratrol; D056564:Sirtuin 1; D037761:Sirtuins / Q000235:genetics / Q000378:metabolism; D013267:Stilbenes / Q000378:metabolism
D000975:Antioxidants; D022782:CCAAT-Enhancer-Binding Protein-beta; D019925:Cyclin A; D016877:Oxidants; D034741:RNA, Small Interfering; D017382:Reactive Oxygen Species; D013267:Stilbenes; D004247:DNA; D006861:Hydrogen Peroxide; C470545:Sirt1 protein, mouse; D056564:Sirtuin 1; D037761:Sirtuins; D000077185:Resveratrol