Stimulation of myofibrillar protein degradation and expression of mRNA encoding the ubiquitin-proteasome system in C2C12 myotubes by dexamethasone: Effect of the proteasome inhibitor MG-132

Addition of the synthetic glucocorticoid, dexamethasone (Dex) to serum-depr ived C2C12 myotubes elicited time- and concentration-dependent changes in N -tau-methylhistidine (3-MH), a marker of myofibrillar protein degradation. Within 24 h, 100 nM Dex significantly decreased the cell content of 3-MH an d increased release into the medium. Both of these responses had increased in magnitude by 48 h and then declined toward basal values by 72 h. The inc rease in the release of 3-MH closely paralleled its loss from the cell prot ein. Furthermore, Dex also decreased the 3-MH:total cell protein ratio, sug gesting that myofibrillar proteins were being preferentially degraded. Incu bation of myotubes with the peptide aldehyde, MC-132, an inhibitor of prote olysis by the (ATP)-ubiquitin (Ub)-dependent proteasome, prevented both the basal release of 3-MH (> 95%) and the increased release of 3-MH into the m edium in response to Dex (> 95%). Northern hybridization studies demonstrat ed that Dex also elicited similar time- and concentration-dependent increas es in the expression of mRNA encoding two components (14 kDa E-2 Ub-conjuga ting enzyme and Ub) of the ATP-Ub-dependent pathway. The data demonstrate t hat Dex stimulates preferential hydrolysis of myofibrillar proteins in C2C1 2 myotubes and suggests that the ATP-Ub-dependent pathway is involved in th is response. (C) 1999 Wiley-Liss, Inc.