Characterization of a TM-4 type tropomyosin that is essential for myofibrillogenesis and contractile activity in embryonic hearts of the Mexican axolotl.
A striated muscle isoform of a Tropomyosin (TM-4) gene was characterized and found to be necessary for contractile function in embryonic heart. The full-length clone of this isoform was isolated from the Mexican axolotl (Ambystoma mexicanum) and named Axolotl Tropomyosin Cardiac-3 (ATmC-3). The gene encoded a cardiac-specific tropomyosin protein with 284 amino acid residues that demonstrated high homology to the Xenopus cardiac TM-4 type tropomyosin. Northern blot analysis indicates a transcript of approximately 1.25 kb in size. RT-PCR and in situ hybridization demonstrated that this isoform is predominantly in cardiac tissue. Our laboratory uses an animal model that carries a cardiac lethal mutation (gene c), this mutation results in a greatly diminished level of tropomyosin protein in the ventricle. Transfection of ATmC-3 DNA into mutant hearts increased tropomyosin levels and promoted myofibrillogenesis. ATmC-3 expression was blocked in normal hearts by transfection of exon-specific anti-sense oligonucleotide (AS-ODN). RT-PCR confirmed lower transcript expression of ATmC-3 and in vitro analysis confirmed the specificity of the ATmC-3 exon 2 anti-sense oligonucleotide. These AS-ODN treated hearts also had a disruption of myofibril organization and disruption of synchronous contractions. These results demonstrated that a striated muscle isoform of the TM-4 gene was expressed embryonically and was necessary for normal structure and function of the ventricle.
Spinner
Belinda J
BJ
Department of Cell and Developmental Biology, Upstate Medical University, 750 East Adams Street, Syracuse, New York 13210, USA.
Zajdel
Robert W
RW
McLean
Matthew D
MD
Denz
Christopher R
CR
Dube
Syamalima
S
Mehta
Sonali
S
Choudhury
Aruna
A
Nakatsugawa
Masako
M
Dobbins
Nancy
N
Lemanski
Larry F
LF
Dube
Dipak K
DK
D000558:Ambystoma mexicanum / Q000196:embryology* / Q000235:genetics* / Q000502:physiology; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D018507:Gene Expression Regulation, Developmental; D006321:Heart / Q000196:embryology* / Q000502:physiology; D017403:In Situ Hybridization; D008969:Molecular Sequence Data; D009154:Mutation; D009200:Myocardial Contraction; D020319:Oligodeoxyribonucleotides, Antisense / Q000235:genetics / Q000494:pharmacology; D010802:Phylogeny; D014335:Tropomyosin / Q000235:genetics* / Q000502:physiology
D018076:DNA, Complementary; D020319:Oligodeoxyribonucleotides, Antisense; D014335:Tropomyosin