STAT5a promotes the transcription of mature mmu-miR-135a in 3T3-L1 cells by binding to both miR-135a-1 and miR-135a-2 promoter elements.
Despite extensive research on the role of miR-135a in biological processes, very little attention has been paid to the regulation of its transcription. We have previously reported that miR-135a suppresses 3T3-L1 preadipocyte differentiation and adipogenesis by directly targeting the adenomatous polyposis coli (APC) gene and activating the canonical Wnt/β-catenin signaling pathway, but the regulatory elements that regulate the expression of the two isoforms of miR-135a (miR-135a-1 and miR-135a-2) remain poorly understood. Here, by using deletion analysis, we predicted two binding sites (-874/-856 and -2020/-2002) for the transcription factor Signal Transducers and Activators of Transcription 5a (STAT5a) within the core promoters of miR-135a-1 and miR-135a-2 (-1128/-556 and -2264/-1773), and the subsequent site-directed mutagenesis indicated that the two STAT5a binding sites regulated the activity of the miR-135a-1 and miR-135a-2 promoters. The binding of STAT5a to the miR-135a-1/2 core promoters in vitro and in cell culture was identified by electrophoretic mobility shift assays (EMSA) and chromatin immunoprecipitation (ChIP) assays. Overexpression and RNAi knockdown of STAT5a showed that the transcription factor regulated the endogenous miR-135a expression. Additionally, The expression time frame of STAT5a and APC indicated a potential negative feedback between them. In sum, the overall results from this study indicate that STAT5a regulates miR-135a transcription by binding to both miR-135a-1 and miR135a-2 promoter elements and the findings provide novel insights into the molecular regulatory mechanisms of miR-135a during adipogenesis.
Key Laboratory of Swine Genetics and Breeding of Agricultural Ministry, College of Animal Science and Technology, Huazhong Agricultural University, Wuhan, China; The Cooperative Innovation Center for Sustainable Pig Production, Wuhan, China.
Xiajie
X
Wei
College of Fisheries, Key Lab of Freshwater Animal Breeding, Ministry of Agriculture, Huazhong Agricultural University, Freshwater Aquaculture Collaborative Innovation Center of Hubei Province, Wuhan, China.
Xiaoyan
X
Cheng
Key Laboratory of Swine Genetics and Breeding of Agricultural Ministry, College of Animal Science and Technology, Huazhong Agricultural University, Wuhan, China; The Cooperative Innovation Center for Sustainable Pig Production, Wuhan, China; College of Animal Science and Technology, Hebei Normal University of Science and Technology, Qinhuangdao, 066004 Hebei, China.
Yongdong
Y
Peng
Key Laboratory of Swine Genetics and Breeding of Agricultural Ministry, College of Animal Science and Technology, Huazhong Agricultural University, Wuhan, China; The Cooperative Innovation Center for Sustainable Pig Production, Wuhan, China.
Rong
R
Zheng
Key Laboratory of Swine Genetics and Breeding of Agricultural Ministry, College of Animal Science and Technology, Huazhong Agricultural University, Wuhan, China; The Cooperative Innovation Center for Sustainable Pig Production, Wuhan, China.
Jin
J
Chai
Key Laboratory of Swine Genetics and Breeding of Agricultural Ministry, College of Animal Science and Technology, Huazhong Agricultural University, Wuhan, China; The Cooperative Innovation Center for Sustainable Pig Production, Wuhan, China. Electronic address: jiangsiwen@mail.hzau.edu.cn.
Siwen
S
Jiang
D041721:3T3-L1 Cells; D025601:Adenomatous Polyposis Coli Protein / Q000235:genetics; D000818:Animals; D051379:Mice; D035683:MicroRNAs / Q000235:genetics*; D009154:Mutation; D011401:Promoter Regions, Genetic / Q000235:genetics*; D050799:STAT5 Transcription Factor / Q000378:metabolism*; D014158:Transcription, Genetic; D015854:Up-Regulation
D025601:Adenomatous Polyposis Coli Protein; D035683:MicroRNAs; C549289:Mirn135 microRNA, mouse; D050799:STAT5 Transcription Factor; C000613192:adenomatous polyposis coli protein, mouse
3T3-L1 cells; Adipogenesis; STAT5a; Transcription; miR-135a-1/2