DIAPHRAGM MUSCLE PROTEIN UBIQUITINATION FOLLOWING UNILATERAL DENERVATION

Abstract

Muscle protein content reflects a balance between protein synthesis and degradation, which are likely regulated by neuromuscular activity and/or neurotrophin influence. In previous studies, we found that unilateral denervation (DNV) of the rat diaphragm muscle (DIAm) decreases myosin heavy chain (MHC) protein content after approximately 7 days. However, we did not determine whether the decrease in DIAm MHC protein content was the result of a decrease in protein synthesis, an increase in protein degradation, or both. We hypothesize that protein degradation plays a significant role in the decrease in DIAm protein content following DNV. In this study, we analyzed the ubiquitination state of cellular proteins as a marker of protein degradation. Conjugation of ubiquitin to cellular proteins targets proteins for degradation by proteosomes. We analyzed DIAm from adult male Sprague-Dawley rats 3, 7 and 14 days after DNV. The DIAm was homogenized and protein ubiquitination was analyzed by Western analysis. No difference in ubiquitination was detected between control (CTL), sham surgery-treated (sham), and 3-day DNV animals. However, following 7 and 14 days of DNV, there was an approximate five-fold increase in protein ubiquitination in DIAm compared to CTL and sham groups. These results demonstrate that DIAm protein ubiquitination increases between 3 and 7 days following DNV, and support the hypothesis that protein degradation contributes to the DNV-induced decrease in DIAm MHC protein content. Supported by NIH grants AR51173 and HL37680.