Mint represses transactivation of the type II collagen gene enhancer through interaction with alpha A-crystallin-binding protein 1.
Collagen type II is an extracellular matrix protein important for cartilage and bone formation, and its expression is controlled by multiple cis- and trans-acting elements, including the zinc finger transcription factor alpha A-crystallin-binding protein 1 (CRYBP1). Here we show that MSX2-interacting nuclear target protein (MINT), a conserved transcriptional repressor, associates with CRYBP1 and negatively regulates the transactivation of the collagen type II gene (Col2a1) enhancer. We identified CRYBP1 as a binding partner of MINT by screening a mouse embryonic cDNA library using the yeast two-hybrid system. We demonstrated that the C terminus of MINT interacts with the C terminus of CRYBP1 using the mammalian cell two-hybrid assay, glutathione S-transferase pull-down, and co-immunoprecipitation analyses. Furthermore, MINT and CRYBP1 form a complex on the Col2a1 enhancer, as shown by chromatin immunoprecipitation and gel shift assays. In the presence of CRYBP1, overexpression of MINT or its C-terminal fragment in cells repressed a reporter construct driven by the Col2a1 enhancer elements. This transcription repression is dependent on histone deacetylase, the main co-repressor recruited by MINT. The present study shows that MINT is involved in CRYBP1-mediated Col2a1 gene repression and may play a role in regulation of cartilage development.
Yang
Xi
X
Department of Medical Genetics and Developmental Biology, State Key Laboratory of Cancer Biology, Fourth Military Medical University, Xian 710032, China.
Li
Junfeng
J
Qin
Hongyan
H
Yang
Hui
H
Li
Junlin
J
Zhou
Peng
P
Liang
Yingmin
Y
Han
Hua
H
D000818:Animals; D001842:Bone and Bones / Q000378:metabolism; D002356:Cartilage / Q000378:metabolism; D002460:Cell Line; D047369:Chromatin Immunoprecipitation; D024043:Collagen Type II / Q000737:chemistry / Q000235:genetics*; D018076:DNA, Complementary / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry*; D004742:Enhancer Elements, Genetic; D005786:Gene Expression Regulation; D015723:Gene Library; D017930:Genes, Reporter; D005982:Glutathione Transferase / Q000378:metabolism; D006657:Histones / Q000737:chemistry; D006801:Humans; D047468:Immunoprecipitation; D008156:Luciferases / Q000378:metabolism; D051379:Mice; D041681:NIH 3T3 Cells; D009687:Nuclear Proteins / Q000378:metabolism / Q000502:physiology*; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D016601:RNA-Binding Proteins; D014157:Transcription Factors / Q000737:chemistry*; D015533:Transcriptional Activation; D020798:Two-Hybrid System Techniques; D016335:Zinc Fingers; D038202:alpha-Crystallin A Chain / Q000737:chemistry*
C499177:COL2A1 protein, human; C447413:Col2a1 protein, mouse; D024043:Collagen Type II; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D006657:Histones; C091656:Hivep1 protein, mouse; D009687:Nuclear Proteins; D016601:RNA-Binding Proteins; C121253:Spen protein, mouse; D014157:Transcription Factors; D038202:alpha-Crystallin A Chain; D008156:Luciferases; D005982:Glutathione Transferase