Increases in nuclear p65 activation in dystrophic skeletal muscle are secondary to increases in the cellular expression of p65 and are not solely produced by increases in IκB-α kinase activity

Abstract

Dystrophin-deficient muscle exhibits substantial increases in nuclear NF-κB activation. To examine potential mechanisms for this enhanced activation, the present study employs conventional Western blot techniques to provide the first determination of the relative expression of NF-κB signaling molecules in adult nondystrophic and dystrophic (mdx) skeletal muscle. The results indicate that dystrophic muscle is characterized by increases in the whole cell expression of IκB-α, p65, p50, RelB, p100, p52, IKK, and TRAF-3. The proportion of phosphorylated IκB-α, p65, NIK, and IKKβ, and the level of cytosolic IκB-α, were also increased in the mdx diaphragm. Proteasomal inhibition using MG-132 increased the proportion of phosphorylated IκB-α in nondystrophic diaphragm, but did not significantly increase this proportion in the mdx diaphragm. This result suggests that phosphorylated IκB-α accumulates in dystrophic cytosol because the rate of IκB-α degradation is lower than the effective rate of IκB-α synthesis and phosphorylation. Dystrophic increases in SUMO-1 (small ubiquitin modifier-1) protein and in Akt activation were also observed. The results indicate that increases in nuclear p65 activation in dystrophic muscle are not produced solely by increases in the activity of IκB-α kinase (IKK), but are due primarily to increases in the expression of p65 and other NF-κB signaling components.