A natural hepatocyte growth factor/scatter factor autocrine loop in myoblast cells and the effect of the constitutive Met kinase activation on myogenic differentiation.
As a rule, hepatocyte growth factor/scatter factor (HGF/SF) is produced by mesenchymal cells, while its receptor, the tyrosine kinase encoded by the met proto-oncogene, is expressed by the neighboring epithelial cells in a canonical paracrine fashion. In the present work we show that both HGF/SF and met are coexpressed by undifferentiated C2 mouse myoblasts. In growing cells, the autocrine loop is active as the receptor exhibits a constitutive phosphorylation on tyrosine that can be abrogated by exogenously added anti-HGF/SF neutralizing antibodies. The transcription of HGF/SF and met genes is downregulated when myoblasts stop proliferating and differentiate. The coexpression of HGF/SF and met genes is not exclusive to C2 cells since it has been assessed also in other myogenic cell lines and in mouse primary satellite cells, suggesting that HGF/SF could play a role in muscle development through an autocrine way. To analyze the biological effects of HGF/SF receptor activation, we stably expressed the constitutively activated receptor catalytic domain (p65(tpr-met)) in C2 cells. This active kinase determined profound changes in cell shape and inhibited myogenesis at both morphological and biochemical levels. Notably, a complete absence of muscle regulatory markers such as MyoD and myogenin was observed in p65(tpr-met) highly expressing C2 clones. We also studied the effects of the ectopic expression of human isoforms of met receptor (h-met) and of HGF/SF (h-HGF/SF) in stable transfected C2 cells. Single constitutive expression of h-met or h-HGF/SF does not alter substantially the growth and differentiation properties of the myoblast cells, probably because of a species-specific ligand-receptor interaction. A C2 clone expressing simultaneously both h-met and h-HGF/SF is able to grow in soft agar and shows a decrease in myogenic potential comparable to that promoted by p65(tpr-met) kinase. These data indicate that a met kinase signal released from differentiation-dependent control provides a negative stimulus for the onset of myogenic differentiation.
Anastasi
S
S
Istituto Pasteur-Fondazione Cenci Bolognetti, Dipartimento di Biotecnologie Cellulari ed Ematologia, Sezione di Genetica Molecolare, Università di Roma La Sapienza, 00161 Roma, Italy.
Giordano
S
S
Sthandier
O
O
Gambarotta
G
G
Maione
R
R
Comoglio
P
P
Amati
P
P
D000818:Animals; D002454:Cell Differentiation / Q000502:physiology; D004285:Dogs; D015536:Down-Regulation / Q000502:physiology; D004789:Enzyme Activation; D015870:Gene Expression / Q000502:physiology; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000235:genetics / Q000378:metabolism; D017228:Hepatocyte Growth Factor / Q000494:pharmacology*; D006801:Humans; D007686:Kidney Tubules, Distal / Q000166:cytology; D008099:Liver / Q000166:cytology; D051379:Mice; D008809:Mice, Inbred C3H; D009132:Muscles / Q000737:chemistry / Q000166:cytology* / Q000201:enzymology; D017570:MyoD Protein / Q000235:genetics / Q000378:metabolism; D018008:Myogenin / Q000235:genetics / Q000378:metabolism; D010770:Phosphotransferases / Q000378:metabolism*; D000090063:Proto-Oncogene Mas; D019859:Proto-Oncogene Proteins c-met; D012333:RNA, Messenger / Q000378:metabolism; D020794:Receptor Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013045:Species Specificity; D018243:Teratocarcinoma; D014162:Transfection; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000201:enzymology
C000715470:MAS1 protein, human; C503799:MYOG protein, human; D017570:MyoD Protein; C503801:Myog protein, mouse; D018008:Myogenin; D000090063:Proto-Oncogene Mas; D012333:RNA, Messenger; D011994:Recombinant Proteins; D017228:Hepatocyte Growth Factor; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D010770:Phosphotransferases; D019859:Proto-Oncogene Proteins c-met; D020794:Receptor Protein-Tyrosine Kinases