Signalling pathway interactions during differentiation of skeletal muscle satellite cells

Interactions between different signalling pathways are important for the control of complex cellular processes. In skeletal muscle, a terminally differentiated and mitotically inactive tissue, that is maintained by activation of adult stem cells, the Ras/MAP/Erk pathway is involved in control of proliferation, whereas the PI3-Kinase/Akt pathway is associated with differentiation. Interaction of these pathways in primary rabbit skeletal muscle satellite cells during proliferation, differentiation and maturation was examined by Luciferase reporter gene assays using fragments (~780bp) of the myosin-heavy-chain I (MHCI) or myosin-heavy-chain IIx (MHCIIx) promoter. Cells were cotransfected with expression vectors for constitutive active or dominant negative h-Ras and cultured for 2 days under proliferation or differentiation conditions. H-Ras showed no effect on the MHC promoter in proliferation medium, whereas in differentiation medium H-Ras abolished promoter activation caused by differentiation medium. In contrast, H-Ras expression caused a strong activation of the MHC promoters in differentiation medium after 6 days. Subsequent inhibitor studies with PI-3-kinase (LY294002) and MEK1/2 (PD98059) inhibitors revealed a differentiation-dependent interaction of Akt with Raf-1 that blocks the inhibitory effect of the MAP/Erk pathway on myogenic gene expression during differentiation.