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The homeobox transcription factor Irxl1 negatively regulates MyoD expression and myoblast differentiation
Author(s) -
Chuang HanNi,
Hsiao KuangMing,
Chang HuiYi,
Wu ChiaChi,
Pan Huichin
Publication year - 2014
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/febs.12837
Subject(s) - myod , myogenesis , pitx2 , homeobox , biology , transcription factor , microbiology and biotechnology , chromatin immunoprecipitation , gene knockdown , dlx5 , myod protein , myogenic regulatory factors , myf5 , myocyte , promoter , gene expression , gene , genetics
Irxl1 / Mkx ( Iroquois homeobox‐like 1 / Mohawk ) encodes a member of the TALE subfamily of homeodomain proteins. It is expressed in multiple mesoderm‐derived tissues and has recently been shown to regulate tendon differentiation during mouse embryonic development. Previously we showed that knockdown of Irxl1 in zebrafish caused a deficit in neural crest cells which consequently resulted in deformation of craniofacial muscles and arch cartilages. Here, we further demonstrate that loss of Irxl1 function results in deformed somites with disordered muscle fibers and myotendinous junctions. Because expression of myoD is increased in the somites of Irxl1 knockdown morphants, we test whether Irxl1 negatively regulates myoD expression. When stable C2C12 myoblasts overexpressing Irxl1/Mkx were induced to differentiate, myotube formation was inhibited and protein levels of myoD and myosin heavy chain were decreased accordingly. A series of deletion constructs of myoD promoter fragments were tested by luciferase reporter assays, which identified a promoter fragment that is necessary and sufficient for Irxl1‐mediated repression. Direct interaction of Irxl1 and myoD promoter was subsequently elucidated by yeast one‐hybrid assays, electrophoretic mobility shift assays and chromatin immunoprecipitation analysis. Furthermore, mouse Mkx also binds to and represses myoD promoter. These results indicate that Irxl1/Mkx can repress myoD expression through direct binding to its promoter and may thus play a negative regulatory role in muscle differentiation.