Loss of muscle innervation (i.e. denervation), as occur during aging, spinal cord injuries and motor-neuron diseases, leads to accelerated proteolysis and ultimately to muscle atrophy. The major fraction being degraded during atrophy is the myofibrils, whose loss accounts for the reduced strength, fatigue and disability. The mechanism for turnover of myofibrils during atrophy has long been uncertain, although the ubiquitin-proteasome system seems to play a major role. In this study we show that myofibril destruction in denervated muscle is preceded and accelerated by the depolymerization of the desmin cytoskeleton. Desmin depolymerization requires phosphorylation and ubiquitination by the ubiquitin ligase Trim32. This E3 is expressed in many cell types and mutations affecting this enzyme are associated with Limb Gridle Muscular Dystrophy 2H. Downregulation of Trim32 in denervated muscles by the electroporation of a shRNA prevented desmin disassembly and attenuated the loss of myofibrils, especially of thin filament. Interestingly, desmin phosphorylation was already evident 7 days after denervation, before there was any effect on myofibril content. Moreover, proteolytic products of desmin accumulated in muscles 10 days after denervation, before actin and myosin are lost. Finally, enhanced dissociation of desmin filaments by the expression of a dominant negative accelerated myofibril destruction 7 days after denervation. Thus, during atrophy, desmin depolymerization accelerates myofibril breakdown, and the complete loss of desmin, as seen with fasting, is not required for myofibril destruction.