Cha, a basic helix-loop-helix transcription factor involved in the regulation of upstream stimulatory factor activity.
We report here the characterization of Cha, a transcription factor of the basic helix-loop-helix (bHLH) family. The basic region of Cha shares DNA-interacting amino acids with members of class C bHLH transcription factors. In addition, the HLH region of Cha presents a Myc-type dimerization domain signature required for heterodimer formation between members of this class. Cha protein and mRNA were ubiquitously expressed in many human tissues. Electrophoretic mobility shift assays showed that Cha and upstream stimulatory factor (USF)-1 formed a complex that specifically bound to E-box DNA elements. Moreover, pull-down and co-immunoprecipitation experiments showed an interaction between Cha and USF-1. Cha did not bind to E-box DNA elements and required USF-1 for protein-DNA complex formation. Moreover, Cha inhibited USF-1-stimulated transcription of CD2 (a USF-1-dependent gene) and E-box promoter reporter plasmids. Chromatin immunoprecipitation assays showed that Cha occupied the CD2 promoter in resting, but not in mitogen-stimulated, T cells. Finally, Cha mRNA and protein expression were high in resting T cells and absent in mitogen-activated T cells and inversely correlated with CD2 expression. Contrarily, overexpression of Cha in T cells significantly reduced CD2 expression. In summary, our results indicated that Cha is a new bHLH transcription factor that negatively regulates USF-dependent transcription.
Rodríguez
Clara I
CI
Centro de Biología Molecular, Consejo Superior de Investigaciones Científicas-Universidad Autónoma de Madrid, Universidad Autónoma de Madrid, Cantoblanco, Madrid 28049, Spain.
Gironès
Núria
N
Fresno
Manuel
M
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D051792:Basic Helix-Loop-Helix Proteins; D018801:CD2 Antigens / Q000096:biosynthesis; D019556:COS Cells; D002478:Cells, Cultured; D002843:Chromatin / Q000378:metabolism; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000378:metabolism; D004268:DNA-Binding Proteins; D019281:Dimerization; D005434:Flow Cytometry; D005786:Gene Expression Regulation; D018257:Helix-Loop-Helix Motifs; D006801:Humans; D019169:Jurkat Cells; D008957:Models, Genetic; D008969:Molecular Sequence Data; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013601:T-Lymphocytes / Q000378:metabolism; D013997:Time Factors; D014018:Tissue Distribution; D014157:Transcription Factors / Q000096:biosynthesis* / Q000737:chemistry* / Q000235:genetics* / Q000502:physiology*; D014158:Transcription, Genetic; D051720:Upstream Stimulatory Factors
D051792:Basic Helix-Loop-Helix Proteins; D018801:CD2 Antigens; D002843:Chromatin; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D012333:RNA, Messenger; D011994:Recombinant Proteins; D014157:Transcription Factors; D051720:Upstream Stimulatory Factors; C115162:TCFL5 protein, human; C497181:USF1 protein, human