title: MiR-499 regulates cell proliferation and apoptosis during late-stage cardiac differentiation via Sox6 and cyclin D1.
abstract: BACKGROUND
  MiR-499 is a cardiac-abundant miRNA. However, the biological functions of miR-499 in differentiated cardiomyocytes or in the cardiomyocyte differentiation process is not very clear. Sox6 is believed to be one of its targets, and is also believed to play a role in cardiac differentiation. Therefore, our aim was to investigate the association between Sox6 and miR-499 during cardiac differentiation.
  
  
  RESULTS
  Using a well-established in vitro cardiomyocyte differentiation system, mouse P19CL6 cells, we found that miR-499 was highly expressed in the late stage of cardiac differentiation. In cells stably transfected with miR-499 (P-499 cells), it was found that miR-499 could promote the differentiation into cardiomyocytes at the early stage of cardiac differentiation. Notably, cell viability assay, EdU incorporation assay, and cell cycle profile analysis all showed that the P-499 cells displayed the distinctive feature of hyperplastic growth. Further investigation confirmed that miR-499 could promote neonatal rat cardiomyocyte proliferation. MiR-499 knock-down enhanced apoptosis in the late differentiation stage in P19CL6 cells, but overexpression of miR-499 resulted in a decrease in the apoptosis rate. Sox6 was identified as a direct target of miR-499 and its expression was detected from day 8 or day 10 of cardiac differentiation of P19CL6 cells. Sox6 played a role in cell viability, inhibited cell proliferation and promoted cell apoptosis in P19CL6 cells and cardiomyocytes. The overexpression of Sox6 could reverse the proliferation and anti-apoptosis effects of miR-499. It was also found that miR-499 might exert its function by regulating cyclin D1 via its influence on Sox6.
  
  
  CONCLUSIONS
  miR-499 probably regulates the proliferation and apoptosis of P19CL6 cells in the late stage of cardiac differentiation via its effects on Sox6 and cyclin D1.
authors.affiliation: Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Key Laboratory of Molecular Cardiovascular Sciences, Ministry of Education of China, Peking University, Beijing, China.
authors.forename: Xianhui
authors.initials: X
authors.lastname: Li
authors.forename: Jiaji
authors.initials: J
authors.lastname: Wang
authors.forename: Zhuqing
authors.initials: Z
authors.lastname: Jia
authors.forename: Qinghua
authors.initials: Q
authors.lastname: Cui
authors.forename: Chenguang
authors.initials: C
authors.lastname: Zhang
authors.forename: Weiping
authors.initials: W
authors.lastname: Wang
authors.forename: Ping
authors.initials: P
authors.lastname: Chen
authors.forename: Kangtao
authors.initials: K
authors.lastname: Ma
authors.forename: Chunyan
authors.initials: C
authors.lastname: Zhou
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D017209:Apoptosis / Q000235:genetics; D002453:Cell Cycle / Q000235:genetics; D002454:Cell Differentiation; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D019938:Cyclin D1 / Q000235:genetics* / Q000378:metabolism; D053595:Embryonic Stem Cells / Q000166:cytology / Q000378:metabolism*; D018507:Gene Expression Regulation, Developmental; D051379:Mice; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism; D032383:Myocytes, Cardiac / Q000166:cytology / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D055755:SOXD Transcription Factors / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction
chemical_list: C530105:Ccnd1 protein, mouse; C573190:MIRN499 microRNA, mouse; D035683:MicroRNAs; D055755:SOXD Transcription Factors; C095939:Sox6 protein, mouse; D019938:Cyclin D1
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