Skeletal Muscle Actin Gene Research Articles

Desmoglein 2 mutation provokes skeletal muscle actin expression and accumulation at intercalated discs in murine hearts.

Arrhythmogenic cardiomyopathy (AC) is an incurable progressive disease that is linked to mutations in genes coding for components of desmosomal adhesions that are localized to the intercalated disc region, which electromechanically couples adjacent cardiomyocytes. To date, the underlying molecular dysfunctions are not well characterized. In two murine AC models, we find an upregulation of the skeletal muscle actin gene (Acta1), which is known to be a compensatory reaction to compromised heart function. Expression of this gene is elevated prior to visible morphological alterations and clinical symptoms, and persists throughout pathogenesis with an additional major rise during the chronic disease stage. We provide evidence that the increased Acta1 transcription is initiated through nuclear activation of the serum response transcription factor (SRF) by its transcriptional co-activator megakaryoblastic leukemia 1 protein (MKL1, also known as MRTFA). Our data further suggest that perturbed desmosomal adhesion causes Acta1 overexpression during the early stages of the disease, which is amplified by transforming growth factor β (TGFβ) release from fibrotic lesions and surrounding cardiomyocytes during later disease stages. These observations highlight a hitherto unknown molecular AC pathomechanism.

Zebra body myopathy is caused by a mutation in the skeletal muscle actin gene (ACTA1)

We present follow up data on the original case of ‘zebra body myopathy’ published by Lake and Wilson in 1975. Pathological features in a second biopsy performed at the age of 29 years included a wide variation in fibre size, multiple split fibres, excess internal nuclei and endomysial connective tissue, rimmed vacuoles, accumulation of myofibrillar material and large ‘wiped out’ areas lacking stain for oxidative enzymes. The presence of nemaline rods and actin-like filaments in addition to small zebra bodies suggested ACTA1 as a candidate gene. This has been confirmed by the identification of a novel c.1043T.p.Leu348Gln mutation, which probably occurred de novo. This case illustrates that the myopathy associated with zebra bodies is part of the spectrum of myopathies associated with the ACTA1 gene. It also highlights that accumulation of actin filaments is not confined to severe neonatal ACTA1 cases and that progression of weakness can occur in congenital myopathies, as the patient is now wheelchair bound and can only stand with the aid of a walking frame.

C.O.3 Efficacy of cardiac actin over-expression therapy for ACTA1 disease seems mutation specific

Around 200 different mutations in the skeletal muscle actin gene (ACTA1) are known to cause myopathies with various pathologies (e.g. actin aggregates, nemaline bodies, intranuclear rods, cores, caps, and zebra bodies). Developing therapeutic approaches targeting specific mutations is therefore daunting. Cardiac actin (ACTC; the major actin isoform in adult heart) is 99% identical to ACTA1 protein, and plays an important role in fetal skeletal muscle. Thus we hypothesized ACTC might effectively substitute for ACTA1 in postnatal skeletal muscle. Confirming this, we previously showed that the early lethality of a recessive ACTA1 disease model (Acta1 knock-out mice) could be rescued by transgenic ACTC over-expression in post-natal skeletal muscles. Experimental and anecdotal evidence indicates that dominant ACTA1 disease is modulated by the relative amount of mutant ACTA1 protein. We thus investigated harnessing over-expression of ACTC in an attempt to decrease the proportion of mutant ACTA1 in the total striated actin pool, as a candidate therapy. We bred ACTC over-expressing mice with a severe mouse model of dominant ACTA1 disease (ACTA1(D286G).Acta1+/−). Usually ∼67% of mice in this line present with paralysis of the hind limbs, dying before 30days of age, whilst only ∼11% showed the same phenotype when also expressing ACTC. Thus ACTC over-expression produces a significant amelioration of this disease phenotype. We also bred ACTC over-expressing mice with the dominant Acta1(H40Y) mouse model. Male mice in this line exhibit a decreased lifespan, with ∼50% dying by 9weeks of age, and a further 30% dying by 5months of age. Transgenic over-expression of ACTC did not significantly alter their mortality rate. These data indicate that in addition to being effective for recessive ACTA1 mutations, cardiac actin over-expression might be therapeutic for some dominant ACTA1 mutations, but not others.

D.P.14 Mining modifier genes for skeletal muscle actin diseases

Actin and myosin are the most important muscle proteins: they generate the force of muscle contraction. Mutations of the skeletal muscle actin gene (ACTA1) cause congenital myopathies, whose commonest presentation is severe, with death within the first year of birth. However, some cases have a mild adult-onset disease suggesting other, modifier genes may be protective. In order to discover such genes, we are using The Collaborative Cross (CC). The CC is a revolutionary genetic resource, part developed at the Western Australian Institute for Medical Research. The CC is a genetic reference population of hundreds of mouse lines descended from eight very diverse founder strains allowing high resolution mapping of genes for traits of interest. Our approach is to cross Acta1 null mice, which usually die by 9days post-natal and are models of human patients with recessive ACTA1 disease, with a large panel of CC mouse strains. We will determine if any of the (CC×Acta1 null) mice are protected from the lethal myopathic phenotype, and will then use the dense genetic map (>600,000 polymorphisms) of the strains to determine the locations of modifier gene (s). In addition, we are using The CC to “mine” genetic loci associated with cardiac actin (Actc) expression in postnatal skeletal muscle. ACTC is the fetal homologue of ACTA1 in skeletal muscle, but is switched off in this tissue at birth. We previously found that transgenic ACTC expression in postnatal skeletal muscle rescues Acat1 knockout mice, suggesting ACTC could have a therapeutic role for ACTA1 diseases. The CC offers unparalleled ability to find modifier genes, as to do so in humans it would take large populations of patients with the same mutation. If we are successful in this project, we will set a precedent for applying The CC to discover disease modifier genes in any disease with a genetic basis.

Fetal akinesia caused by a novel actin filament aggregate myopathy skeletal muscle actin gene ( ACTA1) mutation

We report a female newborn, diagnosed with fetal akinesia in utero, who died one hour after birth. Post-mortem muscle biopsy demonstrated actin-filament myopathy based on immunolabelling for sarcomeric actin, and large areas of filaments, without rod formation, ultrastructurally. Analysis of DNA extracted from the muscle disclosed a novel de novo heterozygous c.44G>A, GGC>GAC, ‘p.Gly15Asp’ mutation in the ACTA1 gene. Analysis of the location of the mutated amino-acid in the actin molecule suggests the mutation most likely causes abnormal nucleotide binding, and consequent pathological actin polymerization. This case emphasizes the association of fetal akinesia with actin-filament myopathy.

Drosophila indirect flight muscle specific Act88F actin mutants as a model system for studying congenital myopathies of the human ACTA1 skeletal muscle actin gene

Most human ACTA1 skeletal actin gene mutations cause dominant, congenital myopathies often with severely reduced muscle function and neonatal mortality. High sequence conservation of actin means many mutated ACTA1 residues are identical to those in the Drosophila Act88F, an indirect flight muscle specific sarcomeric actin. Four known Act88F mutations occur at the same actin residues mutated in ten ACTA1 nemaline mutations, A138D/P, R256H/L, G268C/D/R/S and R372C/S. These Act88F mutants were examined for similar muscle phenotypes. Mutant homozygotes show phenotypes ranging from a lack of myofibrils to almost normal sarcomeres at eclosion. Aberrant Z-disc-like structures and serial Z-disc arrays, ‘zebra bodies’, are observed in homozygotes and heterozygotes of all four Act88F mutants. These electron-dense structures show homologies to human nemaline bodies/rods, but are much smaller than those typically found in the human myopathy. We conclude that the Drosophila indirect flight muscles provide a good model system for studying ACTA1 mutations.

Myopathies resulting from mutations in sarcomeric proteins.

The past decade has seen the discovery of the major role that mutations in the protein components of the sarcomere plays as a cause of human muscle disease. An overview of the more precise molecular definitions of these diseases is timely. Recent findings include: the beginnings of an understanding of the role of the sarcomere in controlling muscle gene expression; the theoretical analysis of the increasing number of mutations identified in the skeletal muscle actin gene; the identification of mutations in myosin causing hereditary inclusion body myopathy and hyaline body myopathy and the identification of mutations in myotilin in myofibrillar myopathy. An increasing spectrum of human muscle diseases is being shown to be caused by mutations in proteins of all the major components of the sarcomere. Molecular analysis is providing a more accurate delineation of these diseases, but for the giant nebulin and titin genes, molecular diagnosis remains difficult. Treatment options for these disorders will only come through a deeper understanding of the sarcomere and of the pathogenesis of its disorders.

Gene knock-down in rainbow trout embryos using antisense morpholino phosphorodiamidate oligonucleotides.

Gene knock-down technology using antisense molecules has many applications for studying gene function, disrupting undesirable genetic traits, as well as providing effective therapy for a number of viral diseases. Encouraged by these applications, we developed a gene knock-down technique to interfere with gene expression using transgenic rainbow trout expressing the green fluorescent protein (GFP) gene as a model. One of the antisense morpholino phosphorodiamidate oligonucleotides (AMOs) used in this study (AtGFP-1) was 25 nucleotides in length and localized against codons 2 to 8 of GFP messenger RNA. Microinjection of AtGFP-1 into the blastodisc of fertilized eggs decreased the level of GFP gene expression in a dose-dependent manner, A comparison of the effects of various doses of AtGFP-1 suggested that 10 ng of AtGFP-1 was the optimal concentration in that it interfered with specific gene expression without being strongly toxic to trout embryos. Conversely, morpholino phosphorodiamidate oligonucleotides with the inverted AtGFP-1 sequence, which cannot bind to the target mRNA, did not inhibit GFP gene expression. AtGFP-1 did not affect the expression of nontargeted genes such as the skeletal muscle actin and foreign lacZ genes. These results also indicate that AtGFP-1 interfered with the expression of only the targeted gene. Western blot and reverse transcriptase polymerase chain reaction analyses revealed that the amount of GFP protein drastically decreased whereas the mRNA level was not affected by AtGFP-1, suggesting that AtGFP-1 blocked specific gene function at the translational level. Further, this gene inhibition persisted until the hatching stage. Another AMO, which was localized against the junction region between the 5? untranslated region and the starting codon of GFP mRNA (AtGFP-2), also caused inhibition effects. Thus AMOs can have potent and specific gene knock-down effects in trout embryos. This technology may be useful for examining the roles of selected genes and disrupting their expression during embryonic development of salmonid fish.

In vivo analysis of two striated muscle actin promoters reveals combinations of multiple regulatory modules required for skeletal and cardiac muscle-specific gene expression.

We isolated two striated muscle actin genes from medaka Oryzias latipes. OIMA1 is a skeletal muscle actin gene expressed in somitic muscle and head muscle and OIMA2 is probably a cardiac muscle actin gene expressed in both somitic and cardiac muscle. The differential transcription mechanisms for these two genes were examined in embryos by introducing fusion genes in which the OIMA1 or OIMA2 upstream region was connected to the green fluorescent protein gene. Embryos were injected with these fusion genes at the 2-cell stage. A fusion gene containing the region up to -949 of OIMA1 exhibited strong expression in somitic muscle. The coexistence of two regions, -949/-662 and -421/-201, is necessary for skeletal muscle-specific expression of OIMA1. Two E boxes and other unidentified sequences cooperatively function to achieve the full activity of the enhancer -949/-662. As for OIMA2, the region up to -520 is sufficient for strong muscle-specific expression. The region between -520 and -174 of OIMA2 is necessary for specific expression in both skeletal and cardiac muscles. In addition to the CArG box located at -140, an E-box at -430 is important for the expression in cardiac muscle as well as skeletal muscle. When the enhancers for the two muscle actin genes were switched and combined with each other's promoter, they were able to upregulate tissue-specific expression according to their origin. These results suggest that distinct expression patterns of OIMA land OIMA2 are regulated by combination of regulatory modules, each of which contains multiple regulatory elements.

A Leuconostoc lactis protein with homology to ribosomal protein S1 shares common epitopes and common DNA binding properties with a mammalian DNA binding nuclear factor.

A mouse testis cDNA expression library (Clontech) was screened with a synthetic oligonucleotide ligand containing CT-rich motifs derived from the rat skeletal muscle actin gene promoter. These motifs bind nuclear proteins, and seem to be involved in the regulation of the gene. Analysis of isolated clones, which expressed proteins that specifically bind the oligonucleotide, indicated that they were derived from a single gene. This gene was identified as a contaminant of bacterial origin (Leuconostoc lactis). The cloned gene from L. lactis encodes a protein with significant homology to bacterial ribosomal protein S1, which we designated LrpS1-L. Band shift analysis and competition experiments indicated that both the bacterial protein and a mouse nuclear protein specifically bind to the same CT-rich motif of the skeletal muscle actin promoter. Furthermore, antibodies against the recombinant bacterial protein interfered with the formation of complex between the CT-rich element and the mouse nuclear protein. These results indicate that the bacterial LrpS1-L protein and the mammalian protein bind the same CT-rich motif and share common antigenic epitopes.

Isolation, Characterization and Evolution of Nine Pufferfish ( Fugu rubripes) Actin Genes

The Japanese pufferfish Fugu rubripes( Fugu) has a small genome of about 400 Mb. Nine different actin genes have been isolated and sequenced from a genomic library constructed from this teleost. The six muscle-type actin genes include two α-skeletal actins, three α-cardiac actins and an α-anomalous (testis type) actin, and the three cytoplasmic actins include two β-cytoplasmic actins and a β-cytoplasmic (vascular type) actin. The two skeletal muscle actin genes have identical genomic organization, but differ by five amino acid residues. The three cardiac actin genes code for the same protein but differ in their nucleotide sequences and genomic organization. β-Cytoplasmic actin1 differs by three amino acids from β-cytoplasmic actin2. The α-anomalous (testis type) and β-cytoplasmic (vascular type) actins are novel vertebrate actins. The amino acid sequence of α-anomalous (testis type) actin is the most divergent of all the known vertebrate actins and transcripts of this gene are abundant in the testis. The β-cytoplasmic (vascular type) actin gene has eight introns, similar to mammalian smooth muscle actins, and is expressed in vascular tissues such as the gills, kidney and skin. Several known regulatory elements are found in the 5′ flanking sequences and the first intron of various Fuguactin genes. The intron patterns of the various Fuguactins seem to be the result of loss of certain introns from a common ancestral gene.

The amount of the endogenous and exogenous skeletal muscle actin mRNA in the heart of transgenic mice is affected by the genotype of the cardiac actin gene

Abstract Both skeletal muscle and cardiac actins are co-expressed in the newborn heart. However, the amount of the skeletal muscle actin and its mRNA rapidly decreases during early development and the cardiac actin predominates in the adult heart. In BALB/c and DBA mice there is a mutation in the cardiac actin gene which is associated with decreased levels of cardiac actin mRNA and high levels of the skeletal muscle actin transcript in the adult heart. To examine the possibility that the amount of cardiac actin gene product modulates the expression of the skeletal muscle actin gene in the heart, transgenic mice carrying a tagged skeletal muscle actin gene were produced, and the expression of the endogenous and exogenous actin gene was analyzed in offspring carrying different combinations of the cardiac actin alleles. It was found that both the endogenous and exogenous skeletal muscle actin genes were expressed at low levels in the heart of adult mice homozygous for the wild-type cardiac actin gene allele, at abnormally high levels in mice homozygous for the mutated cardiac actin allele, and at intermediate levels in heterozygous mice. This shows that the level of expression of the cardiac actin gene has a trans effect on the expression of the skeletal muscle actin gene.

Regulated expression of muscle-specific genes introduced into mouse embryonal stem cells: inverse correlation with DNA methylation

Abstract Pluripotent embryonal stem cell lines (ES) were isolated from cultured normal mouse blastocysts. These cells retained their capacity to differentiate into a great variety of cell types in cell cultures or in tumors formed after subcutaneous injection of the cells into nude mice. A chimeric actin/globin gene containing about two-thirds of the rat skeletal muscle actin gene and 730 bp of its 5′flanking region fused to the 3′end of the human embryonic ε-globin gene, was inserted into a plasmid containing a neomycin resistance gene ( neo r) whose transcription is regulated by the SV40 early control elements. The prokaryotic vector DNA sequences of this plasmid ( pAG-Neo) were deleted and the two linked genes were introduced into the ES cells by electroporation. G418-resistant clones were isolated, amplified and injected subcutaneously into nude mice. From the teratocarcinoma-like tumors which developed we isolated myogenic as well as nonmyogenic cell lines. In cell lines derived from three independent transfected ES clones, expression of the actin/globin gene was developmentally regulated in myogenic cells. In contrast, in a number of experiments in which the actin/globin gene or other musclespecific genes were introduced into the ES cells without the removal of the pBR sequences, no expression could be detected at any stage. Moreover, in the differentiated lines derived from these clones, G418 resistance was lost, and no neo r transcripts could be detected. Southern-blot analysis of MSPI- or HpaII-digested DNA revealed extensive methylation in the clones that did not express the foreign DNA, whereas no significant methylation of the inserted DNA was observed in clones which expressed the transfected genes. Examination of the DNA extracted from transgenic mice carrying the same actin/globin gene revealed an inverse correlation between methylation of the exogenous gene and its potential to be expressed in the transgenic strain. However, no tissue-specific differences in methylation, related to the tissue specificity of expression of the exogenous gene, could be detected in these experiments. These results suggest that the process of methylation reported here is causally related to constitutive inactivation of the exogenous genes.

Different regulatory elements are required for cell-type and stage specific expression of the Xenopus laevis skeletal muscle actin gene upon injection in X.laevis oocytes and embryos.

In the present study, we demonstrate by transcript mapping that the injected Xenopus skeletal muscle alpha-actin gene is transcribed and spliced in Xenopus oocytes but not correctly initiated at the alpha-actin promoter. This leads to correctly spliced transcripts even if constructs without putative promoter sequences are injected. On the other hand, alpha-actin transcripts are translated in injected oocytes as shown by the detection of alpha-actin protein. By contrast, correctly initiated alpha-actin transcripts can be found in neurula embryos when the injected clone contains 5' flanking sequences extending from +27 to -680. alpha-actin gene fragments without the 680 nucleotides 5' flanking region are activated unspecifically after midblastula transition, whereas the clones carrying this region are activated correctly at the end of gastrulation. Cell type specific expression seems to be modulated by sequences within the transcribed region.

Expression in transgenic mice of two genes of different tissue specificity integrated into a single chromosomal site.

Transgenic mice were used to study the expression of pairs of genes with distinctly different tissue specificities, covalently linked and integrated into the same chromosomal site. A transgenic strain carrying, in close proximity and in the same orientation, the rat fast skeletal muscle myosin light-chain 2 (MLC2) gene and the mouse rearranged immunoglobulin kappa light-chain gene expressed the immunoglobulin gene specifically in the lymphoid tissues, whereas rat MLC2 transcripts were found in skeletal muscle but not in the spleen or the other tissues that were tested. In another transgenic strain, carrying the rat MLC2 gene and a modified rat skeletal muscle actin gene (actin-globin chimeric gene), transcripts of the rat MLC2 gene were detected in skeletal muscle only, whereas the actin-globin transcripts were detected in skeletal muscle as well as in the heart. Moreover, the expression of the chimeric gene was also developmentally regulated. Expression was higher in cardiac muscle than in the skeletal muscle of neonatal mice, whereas expression was higher in skeletal muscle in adult mice. This pattern is consistent with the regulation of the expression of the endogenous skeletal muscle actin gene. Thus, in those transgenic strains that expressed both genes, each gene retained its tissue specificity, in spite of their close proximity. These results indicate a high degree of autonomy of the control elements included in the cloned genomic DNA fragment and demonstrate that a single chromosomal site can be permissive for the proper expression of two genes with different tissue specificities.

Tissue-specific and developmentally regulated expression of a chimeric actin-globin gene in transgenic mice.

A chimeric plasmid containing about 2/3 of the rat skeletal muscle actin gene plus 730 base pairs of its 5' flanking sequences fused to the 3' end of a human embryonic globin gene (D. Melloul, B. Aloni, J. Calvo, D. Yaffe, and U. Nudel, EMBO J. 3:983-990, 1984) was inserted into mice by microinjection into fertilized eggs. Eleven transgenic mice carrying the chimeric gene with or without plasmid pBR322 DNA sequences were identified. The majority of these mice transmitted the injected DNA to about 50% of their progeny. However, in transgenic mouse CV1, transmission to progeny was associated with amplification or deletion of the injected DNA sequences, while in transgenic mouse CV4 transmission was distorted, probably as a result of insertional mutagenesis. Tissue-specific expression was dependent on the removal of the vector DNA sequences from the chimeric gene sequences prior to microinjection. None of the transgenic mice carrying the chimeric gene together with plasmid pBR322 sequences expressed the introduced gene in striated muscles. In contrast, the six transgenic mice carrying the chimeric gene sequences alone expressed the inserted gene specifically in skeletal and cardiac muscles. Moreover, expression of the chimeric gene was not only tissue specific, but also developmentally regulated. Similar to the endogenous skeletal muscle actin gene, the chimeric gene was expressed at a relatively high level in cardiac muscle of neonatal mice and at a significantly lower level in adult cardiac muscle. These results indicate that the injected DNA included sufficient cis-acting control elements for its tissue-specific and developmentally regulated expression in transgenic mice.

Tissue-Specific and Developmentally Regulated Expression of a Chimeric Actin-Globin Gene in Transgenic Mice

A chimeric plasmid containing about 2/3 of the rat skeletal muscle actin gene plus 730 base pairs of its 5' flanking sequences fused to the 3' end of a human embryonic globin gene (D. Melloul, B. Aloni, J. Calvo, D. Yaffe, and U. Nudel, EMBO J. 3:983-990, 1984) was inserted into mice by microinjection into fertilized eggs. Eleven transgenic mice carrying the chimeric gene with or without plasmid pBR322 DNA sequences were identified. The majority of these mice transmitted the injected DNA to about 50% of their progeny. However, in transgenic mouse CV1, transmission to progeny was associated with amplification or deletion of the injected DNA sequences, while in transgenic mouse CV4 transmission was distorted, probably as a result of insertional mutagenesis. Tissue-specific expression was dependent on the removal of the vector DNA sequences from the chimeric gene sequences prior to microinjection. None of the transgenic mice carrying the chimeric gene together with plasmid pBR322 sequences expressed the introduced gene in striated muscles. In contrast, the six transgenic mice carrying the chimeric gene sequences alone expressed the inserted gene specifically in skeletal and cardiac muscles. Moreover, expression of the chimeric gene was not only tissue specific, but also developmentally regulated. Similar to the endogenous skeletal muscle actin gene, the chimeric gene was expressed at a relatively high level in cardiac muscle of neonatal mice and at a significantly lower level in adult cardiac muscle. These results indicate that the injected DNA included sufficient cis-acting control elements for its tissue-specific and developmentally regulated expression in transgenic mice.

Isolation and Characterization of cDNA Clones from Mouse Skeletal Muscle Actin mRNA

The sequence corresponding to approximately 98% of mouse skeletal muscle actin mRNA was determined from cDNA clones isolated from a library of recombinants in pBR322. One of these clones contains DNA corresponding to the complete amino acid coding region and a large part of the 5' and 3' untranslated regions of the mRNA. Comparison of the mouse coding region (conserved at the amino acid level) and noncoding regions with the corresponding regions of the rat skeletal muscle actin gene indicates that the noncoding regions have also been under selective pressure during evolution.

Isolation and characterization of sarcomeric actin genes expressed in Xenopus laevis embryos

A Xenopus laevis complementary DNA (cDNA) library prepared from messenger RNAs extracted from embryos has been screened for actin-coding sequences. Two cDNA clones corresponding to an α cardiac and an α skeletal muscle actin mRNA have been identified and characterized. From a genomic library, we have furthermore isolated the genes that correspond to the characterized cDNAs. In addition we have identified an actin processed gene which seems to be derived from a second type of skeletal muscle actin gene. Southern blot analysis of X. laevis DNA reveals that each of the three genes is present in at least two copies. In Xenopus tropicalis, a similar Southern blot analysis demonstrates that the three α actin genes exist as single copy. This result correlates with the genome duplication that has been proposed to have occurred recently in a X. laevis ancestor. A sequence comparison of the X. laevis cardiac and skeletal muscle actin cDNAs shows that the encoded peptides are highly conserved. Nevertheless, the numerous nucleotide changes at silent mutation sites suggest that the genes originated before the amphibia/reptile-bird divergence, more than 350 million years ago. Comparison of the promoters of the cardiac and skeletal actin genes, which are co-expressed in embryos, reveals a few common structural sequence elements.

Actin genes in Xenopus and their developmental control

ABSTRACT The results summarized here have established the temporal and regional activation of three kinds of Xenopus actin genes. The cardiac and skeletal muscle actin genes are among the first cell-type-specific genes to be expressed in early development. The first transcripts to be synthesized by these genes appear to be correctly initiated, spliced, and at once translated into proteins. Both cardiac and skeletal actin genes are strongly transcribed in the axial skeletal muscle of embryos. The mechanism by which the cardiac actin gene is first transcribed in only the somite region of an embryo depends, at least in part, on materials already localized in the subequatorial region of a fertilized but uncleaved egg. Cells which acquire this material seem able to activate their cardiac actin genes without requiring normal contact with other cells.