title: Distinct enhancers regulate skeletal and cardiac muscle-specific expression programs of the cardiac alpha-actin gene in Xenopus embryos.
abstract: During vertebrate embryonic development, cardiac and skeletal muscle originates from distinct precursor populations. Despite the profound structural and functional differences in the striated muscle tissue they eventually form, such progenitors share many features such as components of contractile apparatus. In vertebrate embryos, the alpha-cardiac actin gene encodes a major component of the myofibril in both skeletal and cardiac muscle. Here, we show that expression of Xenopus cardiac alpha-actin in the myotomes and developing heart tube of the tadpole requires distinct enhancers within its proximal promoter. Using transgenic embryos, we find that mutations in the promoter-proximal CArG box and 5 bp downstream of it specifically eliminate expression of a GFP transgene within the developing heart, while high levels of expression in somitic muscle are maintained. This sequence is insufficient on its own to limit expression solely to the myocardium, such restriction requiring multiple elements within the proximal promoter. Two additional enhancers are active in skeletal muscle of the embryo, either one of which has to interact with the proximal CArG box for correct expression to be established. Transgenic reporters containing multimerised copies of CArG box 1 faithfully detect most sites of SRF expression in the developing embryo as do equivalent reporters containing the SRF binding site from the c-fos promoter. Significantly, while these motifs possess a different A/T core within the CC(A/T)(6)GG consensus and show no similarity in flanking sequence, each can interact with a myotome-specific distal enhancer of cardiac alpha-actin promoter, to confer appropriate cardiac alpha-actin-specific regulation of transgene expression. Together, these results suggest that the role of CArG box 1 in the cardiac alpha-actin gene promoter is to act solely as a high-affinity SRF binding site.
authors.affiliation: Division of Developmental Biology, National Institute for Medical Research, The Ridgeway, London NW7 1AA, UK.
authors.forename: Branko V
authors.initials: BV
authors.lastname: Latinkić
authors.forename: Brian
authors.initials: B
authors.lastname: Cooper
authors.forename: Norma
authors.initials: N
authors.lastname: Towers
authors.forename: Duncan
authors.initials: D
authors.lastname: Sparrow
authors.forename: Surendra
authors.initials: S
authors.lastname: Kotecha
authors.forename: Timothy J
authors.initials: TJ
authors.lastname: Mohun
mesh_terms: D000199:Actins / Q000235:genetics*; D000818:Animals; D001483:Base Sequence; D017931:DNA Primers; D024202:Electrophoretic Mobility Shift Assay; D004742:Enhancer Elements, Genetic; D018507:Gene Expression Regulation, Developmental / Q000235:genetics*; D016762:Genes, fos; D006321:Heart / Q000196:embryology*; D018482:Muscle, Skeletal / Q000196:embryology / Q000378:metabolism*; D009206:Myocardium / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D014981:Xenopus / Q000196:embryology
chemical_list: D000199:Actins; D017931:DNA Primers