title: An HF-1a/HF-1b/MEF-2 combinatorial element confers cardiac ventricular specificity and established an anterior-posterior gradient of expression.
abstract: The molecular determinants that direct gene expression to the ventricles of the heart are for the most part unknown. Additionally, little data is available on how the anterior/posterior axis of the heart tube is determined and whether the left and right atrial and ventricular chambers are assigned as part of this process. Utilizing myosin light chain-2 ventricular promoter/beta-galactosidase reporter transgenes, we have determined the minimal cis-acting sequences required for ventricular-specific gene expression. In multiple independent transgenic mouse lines, we found that both a 250 base pair myosin light chain-2 ventricular promoter fragment, as well as a dimerized 28 bp sub-element (HF-1) containing binding sites for HF1a and HF1b/MEF2 factors, directed ventricular-specific reporter expression from as early as the endogenous gene, at day 7.5-8.0 post coitum. While the endogenous gene is expressed uniformly throughout both ventricles, the transgenes were expressed in a right ventricular/conotruncal dominant fashion, suggesting that they contain only a subset of the elements which respond to positional information in the developing heart tube. Expression of the transgene was cell autonomous and its temporospatial characteristics not affected by mouse strain/methylation state of the genome. To determine whether ventricular-specific expression of the transgene was dependent upon regulatory genes required for correct ventricular differentiation, the 250 base pair transgene was bred into both retinoid X receptoralpha and Nkx2-5 null backgrounds. The transgene was expressed in both mutant backgrounds, despite the absence of endogenous myosin light chain-2 ventricular transcript in Nkx2-5 null embryos. Ventricular specification, as judged by transgene expression, appeared to occur normally in both mutants. Thus, the HF-1 element, directs chamber-specific transcription of a transgene reporter independently of retinoid X receptoralpha and Nkx2-5, and defines a minimal combinatorial pathway for ventricular chamber gene expression. The patterned expression of this transgene may provide a model system in which to investigate the cues that dictate anterior-posterior (right ventricle/left ventricle) gradients during mammalian heart development.
authors.affiliation: Department of Medicine, Center for Molecular Genetics, University of California, San Diego, School of Medicine, La Jolla 92093, USA.
authors.forename: R S
authors.initials: RS
authors.lastname: Ross
authors.forename: S
authors.initials: S
authors.lastname: Navankasattusas
authors.forename: R P
authors.initials: RP
authors.lastname: Harvey
authors.forename: K R
authors.initials: KR
authors.lastname: Chien
mesh_terms: D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005260:Female; D018507:Gene Expression Regulation, Developmental; D006321:Heart / Q000196:embryology*; D007763:Lac Operon; D064326:MEF2 Transcription Factors; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008811:Mice, Inbred DBA; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D018006:Myogenic Regulatory Factors; D018994:Myosin Light Chains / Q000235:genetics / Q000378:metabolism; D009838:Oligodeoxyribonucleotides; D017354:Point Mutation; D011401:Promoter Regions, Genetic; D051381:Rats; D012045:Regulatory Sequences, Nucleic Acid; D051706:Sp4 Transcription Factor; D014157:Transcription Factors / Q000378:metabolism*; D019076:Transgenes
chemical_list: D004268:DNA-Binding Proteins; D064326:MEF2 Transcription Factors; D018006:Myogenic Regulatory Factors; D018994:Myosin Light Chains; D009838:Oligodeoxyribonucleotides; D051706:Sp4 Transcription Factor; C081928:Sp4 protein, rat; D014157:Transcription Factors