Terminal differentiation of Sol 8 myoblasts is retarded by a transforming growth factor-beta autocrine regulatory loop.
In DM (differentiation medium), Sol 8 myoblasts spontaneously form myotubes and express the betaMHC (beta-myosin heavy chain), their main marker of terminal differentiation. This marker is detectable at 24 h, and increases up to 72 h. Our aim was to define temporal effects of TGFbeta (transforming growth factor beta) on betaMHC expression in Sol 8 cells. TGFbeta1 (1 ng/ml) added at time zero to DM decreased MyoD expression and completely inhibited betaMHC expression in Sol 8 cells. This inhibition of betaMHC expression was progressively lost when TGFbeta1 was added from 8 to 34 h. After 34 h, the cells were irreversibly differentiated, and TGFbeta1 did not inhibit betaMHC accumulation any longer. Two independent approaches showed that a TGFbeta autocrine regulatory loop retarded and partially impaired Sol 8 cell terminal differentiation. First, permanent immunoneutralization of the active TGFbetas released by the cells into DM increased betaMHC levels at 72 h compared with controls. Secondly, a dominant-negative mutant of the TGFbeta type II receptor was overexpressed in Sol 8 cells under the control of the betaMHC promoter. Both the dominant-negative receptor and the betaMHC gene were expressed after 24 h in DM. The delayed blocking of the TGFbeta signalling pathway by the dominant-negative receptor was as effective as permanent immunoneutralization to promote betaMHC expression. To conclude, TGFbeta inhibits Sol 8 cell terminal differentiation within a narrow time interval (24-34 h) that coincides with the onset of betaMHC expression.
Allegra
Séverine
S
UMR 369 INSERM/UCBL and IFR 62 Laënnec, Faculté de médecine, R.T.H. Laënnec, 7 rue G. Paradin, 69372 Lyon, Cedex 08, France.
Li
Jacques Yuan
JY
Saez
José Maria
JM
Langlois
Dominique
D
D000818:Animals; D019898:Autocrine Communication / Q000502:physiology*; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology*; D002460:Cell Line; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000502:physiology; D051379:Mice; D008809:Mice, Inbred C3H; D032446:Myoblasts / Q000187:drug effects / Q000378:metabolism / Q000502:physiology*; D018995:Myosin Heavy Chains / Q000096:biosynthesis; D012333:RNA, Messenger / Q000096:biosynthesis; D018125:Receptors, Transforming Growth Factor beta / Q000096:biosynthesis / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000502:physiology; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction / Q000502:physiology; D016212:Transforming Growth Factor beta / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000494:pharmacology / Q000502:physiology*; D053773:Transforming Growth Factor beta1
C000594638:MYH7 protein, rat; D012333:RNA, Messenger; D018125:Receptors, Transforming Growth Factor beta; D011993:Recombinant Fusion Proteins; C508961:Tgfb1 protein, mouse; C508962:Tgfb1 protein, rat; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D018995:Myosin Heavy Chains