Dok-5 is involved in cardiomyocyte differentiation through PKB/FOXO3a pathway.
The insulin receptor substrate (IRS) family plays important roles in cellular growth, signaling, and survival in the brain. We identified IRS6/Dok-5, a member of the IRS family, also expressed in heart. Dok-5 expression level significantly increased during cardiomyocyte differentiation of P19CL6 cells. To understand the mechanism of Dok-5 gene expression and regulation during cardiomyocyte differentiation, we first mapped the transcription start site of the mouse Dok-5 gene and characterized its promoter regions. Truncation and mutation analysis of the Dok-5 promoter identified the forkhead binding element responsible for the repression of Dok-5 promoter activation. The co-localization of FOXO3a and Dok-5 in the mouse heart allows FOXO3a to be a transcriptional regulator of Dok-5. Electrophoretic mobility shift assay and chromatin immunoprecipitation assay confirmed that FOXO3a could bind to the Dok-5 promoter, accompanied by FOXO3a translocation from the nucleus to cytoplasm. FOXO3a overexpression could inhibit Dok-5 promoter activity. Silencing FOXO3a expression by siRNA upregulated the expression of Dok-5 and enhanced cardiomyocyte differentiation. Moreover, Dok-5 siRNA attenuated cardiomyocyte differentiation. Our results provide the first evidence that FOXO3a, the PI3K/PKB downstream substrate, acts as a transcriptional repressor to inhibit the expression of Dok-5. Dok-5 is involved in cardiomyocyte differentiation by a PI3K/PKB/FOXO3a signaling pathway.
National Key Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100005, China.
Jianyan
J
Wen
Qing
Q
Xia
Cheng
C
Wang
Wei
W
Liu
Yang
Y
Chen
Jing
J
Gao
Yanhua
Y
Gong
Bin
B
Yin
Yuannan
Y
Ke
Boqin
B
Qiang
Jiangang
J
Yuan
Xiaozhong
X
Peng
D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002454:Cell Differentiation; D045744:Cell Line, Tumor; D024202:Electrophoretic Mobility Shift Assay; D000071316:Forkhead Box Protein O3; D051858:Forkhead Transcription Factors / Q000378:metabolism*; D018507:Gene Expression Regulation, Developmental; D020868:Gene Silencing; D051379:Mice; D008954:Models, Biological; D008969:Molecular Sequence Data; D009206:Myocardium / Q000378:metabolism; D032383:Myocytes, Cardiac / Q000166:cytology* / Q000201:enzymology*; D021141:Nucleic Acid Amplification Techniques; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000378:metabolism; D024363:Transcription Initiation Site; D014158:Transcription, Genetic
D048868:Adaptor Proteins, Signal Transducing; C499010:Dok5 protein, mouse; D000071316:Forkhead Box Protein O3; D051858:Forkhead Transcription Factors; C474033:FoxO3 protein, mouse; D010750:Phosphoproteins; D034741:RNA, Small Interfering; D051057:Proto-Oncogene Proteins c-akt