Effect of an arginine-specific ADP-ribosyltransferase inhibitor on differentiation of embryonic chick skeletal muscle cells in culture.
Primary cultures of embryonic chick skeletal myogenic cells were used as an experimental model to examine the possible role of mono(ADP-ribosyl)ation reactions in myogenic differentiation. Initial studies demonstrated arginine-specific mono(ADP-ribosyl)transferase activity in the myogenic cell cultures. We then examined the effect of a novel inhibitor of cellular arginine-specific mono(ADP-ribosyl)transferases, meta-iodobenzylguanidine (MIBG), on differentiation of cultured embryonic chick skeletal myoblasts. MIBG reversibly inhibited both proliferation and differentiation of embryonic chick myoblasts grown in culture. Micromolar (15-60 microM) concentrations of MIBG blocked myoblast fusion, the differentiation-specific increase in creatine phosphokinase activity, and both DNA and protein accumulation in myogenic cell cultures. Meta-iodobenzylamine, an analog of MIBG missing the guanidine group, had no effect. Low concentrations of methylglyoxal bis-guanylhydrazone, a substrate for cholera toxin with a higher Km than MIBG, also had no effect, but higher concentrations reversibly inhibited fusion. These findings suggest a possible role for mono(ADP-ribosyl)ation reactions in myogenesis. In addition, the total arginine-specific mono(ADP-ribosyl)transferase activity increased with differentiation in the myogenic cell cultures, and this increase was also blocked by MIBG treatment. Because high levels of activity were found in the membrane fraction derived from later, myotube cultures, the membrane fraction from 96-h cultures was incubated with [32P]NAD+ and subjected to electrophoresis and autoradiography. Three proteins, migrating at 21, 20, and 17 kDa, that were ADP-ribosylated in the absence, but not the presence, of MIBG were identified. These proteins may be endogenous substrates for this enzyme.
Kharadia
S V
SV
Department of Biochemistry and Biophysics, Iowa State University, Ames 50011.
Huiatt
T W
TW
Huang
H Y
HY
Peterson
J E
JE
Graves
D J
DJ
D019797:3-Iodobenzylguanidine; D036002:ADP Ribose Transferases; D000818:Animals; D001120:Arginine; D002454:Cell Differentiation / Q000187:drug effects; D002462:Cell Membrane / Q000187:drug effects; D002478:Cells, Cultured / Q000187:drug effects; D002642:Chick Embryo; D007462:Iodobenzenes / Q000494:pharmacology*; D007700:Kinetics; D009132:Muscles / Q000187:drug effects* / Q000196:embryology / Q000201:enzymology; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D011817:Rabbits
D007462:Iodobenzenes; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D019797:3-Iodobenzylguanidine; C008138:3-iodobenzylamine; D001120:Arginine; D036002:ADP Ribose Transferases