Activation of cardiac and smooth muscle-specific genes in primary human cells after forced expression of human myocardin.
OBJECTIVE
Myocardin is a recently discovered transcriptional regulator of cardiac and smooth muscle development. Its ability to transactivate smooth muscle-specific genes has been firmly established in animal cells but its effect on heart muscle genes has been investigated less extensively and the consequences of ectopic myocardin expression in human cells are unknown.


METHODS
In this study, primary human mesenchymal stem cells and foreskin fibroblasts were transduced with human adenovirus vectors expressing the longest splice variant of the human myocardin gene (hAd5/F50.CMV.myocL) or with control vectors. One week later, the expression of muscle-restricted genes in these cells was analyzed by reverse transcription-polymerase chain reactions and immunofluorescence microscopy.


RESULTS
Forced expression of myocardin induced transcription of cardiac and smooth muscle genes in both cell types but did not lead to activation of skeletal muscle-specific genes. Double labeling experiments using monoclonal antibodies directed against striated (i.e. sarcomeric alpha-actin and sarcomeric alpha-actinin) and cardiac (i.e. natriuretic peptide precursor A) muscle-specific proteins together with a polyclonal antiserum specific for smooth muscle myosin heavy chain revealed that hAd5/F50.CMV.myocL-transduced cells co-express heart and smooth muscle-specific genes.


CONCLUSIONS
These data indicate that the myocardin protein is a strong inducer of both smooth and cardiac muscle genes, but that additional factors are necessary to fully commit cells to either cardiac or smooth muscle cell fates.
van Tuyn
John
J
Department of Cardiology, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, P.O. Box 9600, 2300 RC Leiden, The Netherlands.
Knaän-Shanzer
Shoshan
S
van de Watering
Marloes J M
MJ
de Graaf
Michelle
M
van der Laarse
Arnoud
A
Schalij
Martin J
MJ
van der Wall
Ernst E
EE
de Vries
Antoine A F
AA
Atsma
Douwe E
DE
D000185:Actinin / Q000235:genetics; D000199:Actins / Q000235:genetics; D000256:Adenoviridae / Q000235:genetics; D009320:Atrial Natriuretic Factor / Q000235:genetics; D002454:Cell Differentiation; D002470:Cell Survival; D005347:Fibroblasts; D015870:Gene Expression; D005822:Genetic Vectors / Q000008:administration & dosage; D006801:Humans; D007106:Immune Sera / Q000494:pharmacology; D008297:Male; D059630:Mesenchymal Stem Cells / Q000166:cytology / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D009130:Muscle, Smooth / Q000166:cytology / Q000378:metabolism*; D009206:Myocardium / Q000166:cytology / Q000378:metabolism*; D018995:Myosin Heavy Chains / Q000276:immunology; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D014161:Transduction, Genetic; D000099251:Myocardin
D000185:Actinin; D000199:Actins; D009320:Atrial Natriuretic Factor; D007106:Immune Sera; D018995:Myosin Heavy Chains; D009687:Nuclear Proteins; D015534:Trans-Activators; D000099251:Myocardin; C472955:NPPA protein, human