Integrin-linked kinase is a positive mediator of L6 myoblast differentiation.
Overexpression of ILK in L6 myoblasts results in increased ILK kinase activity, stimulating myotube formation and induction of biochemical differentiation markers. Expression of a dominant negative ILK mutant, ILK(E359K), inhibits endogenous ILK activation and L6 differentiation. Cell cycle analysis of ILK(E359K) cells cultured in serum-free conditions indicates significant apoptosis (11-19% sub-diploid peak) which is not seen in insulin treated cells. Expression of ILK variants does not have significant effects on S-phase transit, however. Known targets of ILK, PKB/Akt or glycogen synthase kinase 3beta are not obviously involved in ILK-induced L6 differentiation. Insulin-stimulated phosphorylation of PKB at Ser473 is unimpaired in the ILK(E359K) cells, suggesting that PKB is not a myogenic target of ILK. Inhibition of GSK3beta by LiCl blocks L6 myogenesis, indicating that ILK-mediated inhibition of GSK3beta is not sufficient for differentiation. Our data do suggest that a LiCl-sensitive interaction of ILK is important in L6 myoblast differentiation.
Miller
Mathew G
MG
Department of Laboratory Medicine and Pathobiology, University of Toronto, Ont., Canada.
Naruszewicz
Izabela
I
Kumar
Ashu S
AS
Ramlal
Toolsie
T
Hannigan
Gregory E
GE
D000818:Animals; D017209:Apoptosis; D015153:Blotting, Western; D000252:Calcium-Transporting ATPases / Q000378:metabolism; D002453:Cell Cycle; D002454:Cell Differentiation; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D002469:Cell Separation; D002478:Cells, Cultured; D016895:Culture Media, Serum-Free / Q000494:pharmacology; D018076:DNA, Complementary / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D000939:Epitopes; D005434:Flow Cytometry; D005799:Genes, Dominant; D005822:Genetic Vectors; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism; D000071679:Glycogen Synthase Kinase 3 beta; D007328:Insulin / Q000494:pharmacology; D018021:Lithium Chloride / Q000494:pharmacology; D009154:Mutation; D009206:Myocardium / Q000166:cytology*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000502:physiology*; D051381:Rats; D013997:Time Factors; D014162:Transfection; D000099200:Scaffold Protein ILK
D000252:Calcium-Transporting ATPases; D016895:Culture Media, Serum-Free; D018076:DNA, Complementary; D004791:Enzyme Inhibitors; D000939:Epitopes; D038362:Glycogen Synthase Kinase 3; D000071679:Glycogen Synthase Kinase 3 beta; D007328:Insulin; D018021:Lithium Chloride; D019869:Phosphatidylinositol 3-Kinases; D017346:Protein Serine-Threonine Kinases; D000099200:Scaffold Protein ILK; C000605508:Gsk3b protein, rat