Evidence for the involvement of cathepsin B in skeletal myoblast differentiation.
Our previous studies suggest that the cysteine protease cathepsin B (catB) is involved in skeletal myoblast differentiation (myogenesis). To test this hypothesis, we examined the effect of trapping one of the two catB alleles on the ability of C2C12 cells to differentiate. During differentiation, catB gene-trapped C2C12 mouse myoblasts (RT-27) demonstrated a similar pattern of intracellular catB activity and protein expression compared to that observed in control C2C12 myoblasts and myoblasts trapped in a gene other than catB. However, compared to control myoblast cell lines, levels of catB activity and protein at each stage of RT-27 differentiation were reduced. The reductions in levels of catB were associated with reductions in several myogenic phenotypes including reduced levels of creatine phosphokinase activity and myosin heavy chain protein, two late biochemical markers of myogenesis, and reduced myotube size and extent of myotube formation over time. Comparable reductions were not observed for myogenin protein, an early biochemical marker of myogenesis, or in myokinase activity and catB related cathepsin L-type activity, two non-specific proteins. Finally, both control and catB gene-trapped myoblasts secreted active catB at pH 7.0. However levels of active pericellular/secreted catB were 50% lower in catB gene-trapped myoblasts. Collectively, these results support a functional link between catB expression and skeletal myogenesis and suggest a role for active pericellular/secreted catB in myoblast fusion.
Jane
Derek T
DT
Department of Biological Sciences, University of Windsor, Windsor, Ontario N9B 3P4, Canada.
DaSilva
Luis
L
Koblinski
Jennifer
J
Horwitz
Marshall
M
Sloane
Bonnie F
BF
Dufresne
Michael J
MJ
D000483:Alleles; D000818:Animals; D015415:Biomarkers / Q000032:analysis; D015153:Blotting, Western; D002401:Cathepsin B / Q000235:genetics / Q000502:physiology*; D002454:Cell Differentiation; D002455:Cell Division; D002460:Cell Line; D003402:Creatine Kinase / Q000378:metabolism; D003470:Culture Media; D007700:Kinetics; D051379:Mice; D018482:Muscle, Skeletal / Q000166:cytology / Q000254:growth & development* / Q000378:metabolism; D018995:Myosin Heavy Chains / Q000378:metabolism; D013234:Stem Cells / Q000502:physiology
D015415:Biomarkers; D003470:Culture Media; D003402:Creatine Kinase; D002401:Cathepsin B; D018995:Myosin Heavy Chains