Nucleotide sequence and expression of the human skeletal α-actin gene: Evolution of functional regulatory domains

doi:10.1016/0888-7543(88)90123-1

Genomics

Volume 3, Issue 4, November 1988, Pages 323-336

https://doi.org/10.1016/0888-7543(88)90123-1 Get rights and content

Abstract

Regulation of the actin multigene family involves the recognition of regulatory sequences that specify the tissue type and developmental program of expression for each actin isotype. In order to investigate the underlying regulatory mechanisms, the human skeletal α-actin gene and its 5′ regulatory region have been cloned and sequenced. This actin gene has seven exons; there is one large intron in the 5′ untranslated region which is characteristic of the actins and many muscle-specific genes. The 5′ flanking sequences are sufficient to direct tissue-specific and differentiation-regulated expression when transfected into the heterologous rat L8 myogenic cells, indicating a highly conserved regulatory system. The DNA sequence was compared to that of other actin genes, and several regions of sequence similarity were identified, particularly within regions known to be important for gene expression. Most notable among the conserved sequences are the $CC(A T rich)_{6} GG$ (CArG box) motifs which have demonstrated interactions with trans-acting transcriptional factors. This same motif has been identified in several other genes and in some also serves as a binding site for transcription regulatory factors.

References (89)

A. Barberis et al.
Mutually exclusive interaction of the CCAAT-binding factor and of a displacement protein with overlapping sequences of a histone gene promoter
Cell
(1987)
A. Bender et al.
MATα1 protein, a yeast transcription activator, binds synergistically with a second protein to a set of cell-type-specific genes
Cell
(1987)
M. Bienz et al.
Heat shock regulatory elements function as an inducible enhancer in the Xenopus hsp 70 gene and when linked to a heterologous promoter
Cell
(1986)
S. Carroll et al.
Structure and complete nucleotide sequence of the chicken α-smooth muscle (aortic) actin gene
J. Biol. Chem
(1986)
L. Chodosh et al.
Human CCAAT-binding proteins have heterologous subunits
Cell
(1988)
T. Cooper et al.
A single cardiac troponin T gene generates embryonic and adult isoforms via developmentally regulated alternate splicing
J. Biol. Chem
(1985)
J. Eldridge et al.
Nucleotide sequence of the chicken cardiac α-actin gene: Absence of strong homologies in the promoter and 3′-untranslated regions with the skeletal α-actin sequence
Gene
(1985)
R. Gahlmann et al.
Alternative splicing generates variants in important functional domains of human slow skeletal troponin T
J. Biol. Chem
(1987)
K. Jones et al.
A cellular DNA-binding protein that activates eukaryotic transcription and DNA replication
Cell
(1987)
S. McKnight et al.
The distal transcription signals of the herpes virus gene share a common hexanucleotide control sequence
Cell
(1984)

S. Pearson-White et al.

A novel hybrid α-tropomyosin in fibroblasts is produced by alternative splicing of transcripts from the skeletal muscle α-tropomyosin gene

J. Biol. Chem

(1987)

M. Periasamy et al.

Fast skeletal muscle myosin light chains 1 and 3 are produced from a single gene by a combined process of differential RNA transcription and splicing

J. Biol. Chem

(1984)

R. Prywes et al.

Inducible binding of a factor to the c-fos enhancer

Cell

(1986)

E. Strehler et al.

Complete nucleotide and encoded amino acid sequence of a mammalian myosin heavy chain gene: Evidence against intron-dependent evolution of the rod

J. Mol. Biol

(1986)

P. Venta et al.

Structure and exon to protein domain relationships of the mouse carbonic anhydrase II gene

J. Biol. Chem

(1985)

B. Winter et al.

Isolation and characterization of the chicken cardiac myosin light chain (L-2A) gene

J. Biol. Chem

(1985)

R. Akhurst et al.

Structure and organization of the CyIII actin gene subfamily of the sea urchin, Strongylocentrotus purpuratus

J. Mol. Biol

(1987)

S. Alonso et al.

Comparison of three actin-coding sequences in the mouse; Evolutionary relationships between the actin genes of warm-blooded vertebrates

J. Mol. Evol

(1986)

W. Bains

MULTAN: A program to align multiple DNA sequences

Nucleic Acids Res

(1986)

W. Bains et al.

Cardiac actin is the major actin gene product in skeletal muscle cell differentiation in vitro

Mol. Cell. Biol

(1984)

A. Baldwin et al.

Structure, evolution, and regulation of a fast skeletal muscle troponin I gene

D. Bergsma et al.

Delimitation and characterization of cis-acting DNA sequences required for the regulated expression and transcriptional control of the chicken skeletal α-actin gene

Mol. Cell. Biol

(1986)

L. Boxer et al.

Identification and characterization of a factor that binds to the human sarcomeric α-actin promoter

(1988)

R. Breathnach et al.

Organization and expression of eucaryotic split genes coding for proteins

Annu. Rev. Biochem

(1981)

R. Brietbart et al.

Alternative splicing: A ubiquitous mechanism for the generation of multiple protein isoforms from single genes

Annu. Rev. Biochem

(1987)

M. Buckingham et al.

Contractile protein genes

M. Buckingham et al.

Actin and myosin genes and their expression during myogenesis in the mouse

F. Calzone et al.

Protein-DNA interactions within regulatory regions required for embryonic activation of the sea urchin CyIIIa actin gene

K. Chang et al.

The complete sequence of the chicken α-cardiac actin gene: A highly conserved vertebrate gene

Nucleic Acids Res

(1985)

A. Cooper et al.

Complete nucleotide sequence of a sea urchin actin gene

Nucleic Acids Res

(1982)

J. Czelusniak et al.

Phylogenetic origins and adaptive evolution of avian and mammalian haemoglobin genes

Nature (London)

(1982)

J. Engel et al.

Isolation and characterization of human actin genes

H. Erba et al.

Structure, chromosome location and expression of the human γ-actin gene: Differential evolution, location and expression of the cytoskeletal β- and γ-actin genes

Mol. Cell. Biol

(1988)

J. Fornwald et al.

The complete nucleotide sequence of the chick α-actin gene and its evolutionary relationship to the actin gene family

Nucleic Acids Res

(1982)

E. Fyrberg et al.

The actin genes of Drosophila: Protein coding regions are highly conserved but intron positions are not

Cell

(1981)

I. Garner et al.

A 5′ duplication of the α-cardiac actin gene in $BALB c$ mice is associated with abnormal levels of α-cardiac and α-skeletal actin mRNAs in adult tissue

EMBO J

(1986)

A. Gil et al.

A sequence downstream of AAUAAA is required for rabbit β-globin mRNA 3′-end formation

Nature (London)

(1984)

M. Gilman et al.

Multiple protein-binding sites in the 5′-flanking region regulate c-fos expression

Mol. Cell. Biol

(1986)

M. Goldberg

C. Gorman et al.

Recombinant genomes which express chloramphenicol acetyl-transferase in mammalian cell

Mol. Cell. Biol

(1982)

F. Graham et al.

A new technique for the assay of infectivity of human adenovirus 5 DNA

Virology

(1973)

J. Grichnik et al.

Tissue restricted and stage specific transcription is maintained within 411 nucleotides flanking the 5′ end of the chicken α-skeletal gene

Nucleic Acids Res

(1986)

R. Grosschedl et al.

Spacer DNA sequences upstream of the TATAAATA sequence are essential for promotion of H2A histone gene transcription in vivo

P. Gunning et al.

Differential patterns of transcript accumulation during human myogenesis

Mol. Cell. Biol

(1987)

Cited by (75)

Development of a polymerase chain reaction assay for species identification of goose and mule duck in foie gras products
2003, Meat Science
Polymerase chain reaction amplification of a conserved region of the α-actin gene has been used for the specific identification of goose (Anser anser) and mule duck (Anas platyrhynchos×Cairina moschata) foie gras. Universal primers were used for the amplification of a DNA fragment containing three introns and four exons of the α-actin gene in goose and mule duck. Sequence analysis of the amplified fragments was necessary for the design of forward species-specific primers in the goose and mule duck α-actin genes. The use of species-specific forward primers, together with a reverse universal primer, produced amplicons of different length, allowing clear identification of goose and mule duck foie gras samples. Analysis of experimental mixtures demonstrated that 1% of duck can be easily detected in goose foie gras using the PCR method developed here. This genetic marker can be very useful for the accurate identification of these two species in foie gras products.
Combinatorial expression of GATA4, Nkx2-5, and serum response factor directs early cardiac gene activity
2002, Journal of Biological Chemistry
Citation Excerpt :
SRF binds to a variety of SREs including the four avian αCA SREs and to the greatest extent to the first (SRE1) and fourth (SRE4) SREs. Non-consensus second (SRE2) and third (SRE3) SREs contain a single GC substitution in the six AT-rich SRE core, and all of these SREs are highly conserved across the evolution of Aves and mammals (35, 36). SRE2 and SRE3 are weak SRF binding sites that resemble preferred Nkx2-5 binding elements (2) and also contain overlapping inhibitory YY1 binding sites (3).
Herein, the restricted expression of serum response factors (SRF) closely overlapped withNkx2-5 and GATA4 transcripts in early chick embryos coinciding with the earliest appearance of cardiac α-actin (αCA) transcripts and nascent myocardial cells. The combinatorial expression of SRF, a MADS box factor Nkx2-5 (a NK4 homeodomain), and/or GATA4, a dual C4 zinc finger protein, in heterologous CV1 fibroblasts and Schneider 2 insect cells demonstrated synergistic induction of αCA promoter activity. These three factors induced endogenous αCA mRNA over a 100-fold in murine embryonic stem cells. In addition, the DNA-binding defective mutant Nkx2-5pm efficiently coactivated the αCA promoter in the presence of SRF and GATA4 in the presence of all four SREs and was substantially weakened when individual SREs were mutated and or serially deleted. In contrast, the introduction of SRFpm, a SRF DNA-binding mutant, blocked the activation with all of the αCA promoter constructions. These assays indicated a dependence upon cooperative SRF binding for facilitating the recruitment of Nkx2-5 and GATA4 to the αCA promoter. Furthermore, the recruitment of Nkx2-5 and GATA4 by SRF was observed to strongly enhance SRF DNA binding affinity. This mechanism allowed for the formation of higher ordered αCA promoter DNA binding complexes, led to a model of SRF physical association with Nkx2-5 and GATA4.
Control of Cardiac-specific Transcription by p300 through Myocyte Enhancer Factor-2D
2001, Journal of Biological Chemistry
The transcriptional integrator p300 regulates gene expression by interaction with sequence-specific DNA-binding proteins and local remodeling of chromatin. p300 is required for cardiac-specific gene transcription, but the molecular basis of this requirement is unknown. Here we report that the MADS (MCM-1, agamous, deficiens, serum response factor) box transcription factor myocyte enhancer factor-2D (MEF-2D) acts as the principal conduit for cardiac transcriptional activation by p300. p300 activation of the native 2130-base pair human skeletal α-actin promoter required a single hybrid MEF-2/GATA-4 DNA motif centered at −1256 base pairs. Maximal expression of the promoter in cultured myocytes and in vivo correlated with binding of both MEF-2 and p300, but not GATA-4, to this AT-rich motif. p300 and MEF-2 were coprecipitated from cardiac nuclear extracts by an oligomer containing this element. p300 was found exclusively in a complex with MEF-2D at this and related sites in other cardiac-restricted promoters. MEF-2D, but not other MEFs, significantly potentiated cardiac-specific transcription by p300. No physical or functional interaction was observed between p300 and other factors implicated in skeletal actin transcription, including GATA-4, TEF-1, or SRF. These results show that, in the intact cell, p300 interactions with its protein targets are highly selective and that MEF-2D is the preferred channel for p300-mediated transcriptional control in the heart.AF288779
DNA annealing and DNA-protein interactions by capillary electrophoresis
2000, Biochemical and Biophysical Research Communications
This work deals with annealing of single-stranded DNA and the binding of a serum respond factor to a DNA probe containing specific binding site. Capillary electrophoresis (CE) method is explored and compared with the mobility-shift gel electrophoresis (GE) procedure. The results indicate the CE method offers direct and rapid annealing of the DNA strands. It requires no prior incubation with additives (polynucleotides, proteins) to reduce nonspecific DNA–protein interactions. Unwanted nonspecific interactions are not observed in the CE method. The presence of a fluorescein tag to the DNA probe yields identical results to those with the radioactive label. A fluorescein tag in the CE work can be used without any adverse effects. The dissociation constant (K_d) of this protein–DNA complex by the CE method was similar to those determined by the GE method (≅10⁻⁶ M). The proposed method is extremely powerful, highly sensitive, quantitative, and fast. It can determine even very small conformational differences of the DNA probe.
Improved muscle-derived expression of human coagulation factor IX from a skeletal actin/CMV hybrid enhancer/promoter
2000, Blood
Citation Excerpt :
The region of bp position −153 to +239 is described as necessary and sufficient for muscle-specificity of the HSA promoter.17 The promoter contains at least 4 CArG boxes.18,37 The CArG box with the consensus sequence CC(A/T)6GG, also referred to as serum response element, binds to serum response factor and is found in several muscle-specific elements.
Hemophilia B is caused by the absence of functional coagulation factor IX (F.IX) and represents an important model for treatment of genetic diseases by gene therapy. Recent studies have shown that intramuscular injection of an adeno-associated viral (AAV) vector into mice and hemophilia B dogs results in vector dose–dependent, long-term expression of biologically active F.IX at therapeutic levels. In this study, we demonstrate that levels of expression of approximately 300 ng/mL (6% of normal human F.IX levels) can be reached by intramuscular injection of mice using a 2- to 4-fold lower vector dose (1 × 10¹¹ vector genomes/mouse, injected into 4 intramuscular sites) than previously described. This was accomplished through the use of an improved expression cassette that uses the cytomegalovirus (CMV) immediate early enhancer/promoter in combination with a 1.2-kilobase portion of human skeletal actin promoter. These results correlated with enhanced levels of F.IX transcript and secreted F.IX protein in transduced murine C2C12 myotubes. Systemic F.IX expression from constructs containing the CMV enhancer/promoter alone was 120 to 200 ng/mL in mice injected with 1 × 10¹¹vector genomes. Muscle-specific promoters performed poorly for F.IX transgene expression in vitro and in vivo. However, the incorporation of a sequence from the α-skeletal actin promoter containing at least 1 muscle-specific enhancer and 1 enhancer-like element further improved muscle-derived expression of F.IX from a CMV enhancer/promoter-driven expression cassette over previously published results. These findings will allow the design of a clinical protocol for therapeutic levels of F.IX expression with lower vector doses, thus enhancing efficacy and safety of the protocol.
Physical mapping of the rippling muscle disease locus
1999, Genomics
Rippling muscle disease (RMD) is an autosomal dominant disorder characterized by electrically silent, percussion-induced muscular contractions. We previously reported the localization of a gene for RMD to 1q41–q42 by genome-wide linkage analysis in a large family from Oregon. This RMD gene was initially found to be contained within a 12-cM interval with a maximum multipoint lod score of 3.56. A YAC/BAC contig was assembled by STS content mapping and database searches spanning the nonrecombinant interval containing the RMD gene (RMD1). The physical map, in conjunction with recent mapping information from various other sources, clarified the order of genetic markers in this region and necessitated redefinition of the RMD genetic interval by linkage analysis with the newly ordered markers. Polymorphisms that mapped to the YACs in this contig were genotyped in this family and used to provide statistical support for narrowing of the critical genetic interval to 3 cM, corresponding to a maximum possible physical distance of 4.0 Mb. In addition, recombination breakpoint mapping supported the evidence thatRMD1must reside within this interval between markers D1S446 and D1S2680. ESTs (82) were mapped to the YACs spanning the region known to contain theRMD1gene, and of these, 9 become strong positional candidates. The physical and refined genetic maps of this RMD locus set the stage for isolation of the responsible gene and elucidation of a novel patho-mechanism of calcium homeostasis in skeletal muscle.

View all citing articles on Scopus

^☆: Sequence data from this article have been deposited with the EMBL/GenBank Data Libraries under Accession No. M20543.

View full text

Nucleotide sequence and expression of the human skeletal α-actin gene: Evolution of functional regulatory domains☆

Abstract

Cell

Cell

Cell

J. Biol. Chem

Cell

J. Biol. Chem

Gene

J. Biol. Chem

Cell

Cell

J. Biol. Chem

J. Biol. Chem

Cell

J. Mol. Biol

J. Biol. Chem

J. Biol. Chem

Structure and organization of the CyIII actin gene subfamily of the sea urchin, Strongylocentrotus purpuratus

J. Mol. Biol

Comparison of three actin-coding sequences in the mouse; Evolutionary relationships between the actin genes of warm-blooded vertebrates

J. Mol. Evol

MULTAN: A program to align multiple DNA sequences

Nucleic Acids Res

Cardiac actin is the major actin gene product in skeletal muscle cell differentiation in vitro

Mol. Cell. Biol

Structure, evolution, and regulation of a fast skeletal muscle troponin I gene

Delimitation and characterization of cis-acting DNA sequences required for the regulated expression and transcriptional control of the chicken skeletal α-actin gene

Mol. Cell. Biol

Identification and characterization of a factor that binds to the human sarcomeric α-actin promoter

Organization and expression of eucaryotic split genes coding for proteins

Annu. Rev. Biochem

Alternative splicing: A ubiquitous mechanism for the generation of multiple protein isoforms from single genes

Annu. Rev. Biochem

Contractile protein genes

Actin and myosin genes and their expression during myogenesis in the mouse

Protein-DNA interactions within regulatory regions required for embryonic activation of the sea urchin CyIIIa actin gene

The complete sequence of the chicken α-cardiac actin gene: A highly conserved vertebrate gene

Nucleic Acids Res

Complete nucleotide sequence of a sea urchin actin gene

Nucleic Acids Res

Phylogenetic origins and adaptive evolution of avian and mammalian haemoglobin genes

Nature (London)

Isolation and characterization of human actin genes

Structure, chromosome location and expression of the human γ-actin gene: Differential evolution, location and expression of the cytoskeletal β- and γ-actin genes

Mol. Cell. Biol

The complete nucleotide sequence of the chick α-actin gene and its evolutionary relationship to the actin gene family

Nucleic Acids Res

The actin genes of Drosophila: Protein coding regions are highly conserved but intron positions are not

Cell

A 5′ duplication of the α-cardiac actin gene in BALBc mice is associated with abnormal levels of α-cardiac and α-skeletal actin mRNAs in adult tissue

EMBO J

A sequence downstream of AAUAAA is required for rabbit β-globin mRNA 3′-end formation

Nature (London)

Multiple protein-binding sites in the 5′-flanking region regulate c-fos expression

Mol. Cell. Biol

Recombinant genomes which express chloramphenicol acetyl-transferase in mammalian cell

Mol. Cell. Biol

A new technique for the assay of infectivity of human adenovirus 5 DNA

Virology

Tissue restricted and stage specific transcription is maintained within 411 nucleotides flanking the 5′ end of the chicken α-skeletal gene

Nucleic Acids Res

Spacer DNA sequences upstream of the TATAAATA sequence are essential for promotion of H2A histone gene transcription in vivo

Differential patterns of transcript accumulation during human myogenesis

Mol. Cell. Biol

A 5′ duplication of the α-cardiac actin gene in $BALB c$ mice is associated with abnormal levels of α-cardiac and α-skeletal actin mRNAs in adult tissue