Membrane Hyperpolarization Triggers Myogenin and Myocyte Enhancer Factor-2 Expression during Human Myoblast Differentiation.

S. König, V. Hinard, S. Arnaudeau, A. Béguet, C.R. Bader, and L. Bernheim.

Department of Basic Neurosciences, Centre Medical Universitaire, CH-1211 Geneva , Switzerland .  

Skeletal muscle formation arises through the differentiation and fusion of mononucleated myoblasts into multinucleated myotubes. The initiation and the progression through differentiation is a Ca2+?dependent multi-step process that involves the interplay of ionic channels, intracellular pathways, and myogenic transcription factors expressed at the onset of the differentiation process.We previously showed that human myoblasts must hyperpolarize to approximately -70 mV (i.e. their membrane potential must become more negative inside the cell) before they can proceed through the differentiation process. Hyperpolarization occurs through the expression of Kir2.1 K+ currents and generates an increase in intracellular Ca2+ that is necessary for myoblast differentiation and fusion to occur. The process of differentiation into myotubes is mainly regulated by two families of transcription factors, the myogenic bHLH family (MyoD, Myogenin, Myf5 and MRF4) and the MEF2 family (MEF2A-D). It is widely thought that myogenin is one of the earliest detectable markers of skeletal muscle differentiation.
In the present work, we show that functional Kir2.1 channels can be detected several hours before the expression of myogenin and MEF2 transcription factors. This result suggests that the hyperpolarization takes place in the very early stages of myoblast differentiation. Furthermore, drugs or antisense reducing Kir2.1 current (and thereby depolarizing myoblasts) inhibit expression/activity of myogenin and MEF2 as well as the rate of fusion of the myoblasts into myotubes. LY294002, an inhibitor of phosphatidylinositol 3-kinase which inhibits both myogenin/MEF2 expression did not affect Kir2.1 current indicating that Kir2.1 current acts upstream of myogenin and MEF2.Taken together, our results show that Kir2.1 channel activation is a required key early event that initiates myogenesis by turning on myogenin and MEF2 transcription factors via a hyperpolarization-activated Ca2+-dependent pathway. We propose that the Kir2.1-induced hyperpolarization might control regulatory pathways (p38 MAPK, CaMK and Calcineurin) implicated in the control of myogenic transcription factors.An ionic channel is thus an essential relay in the signaling pathway that leads to the expression of key myogenic transcription factors.