COORDINATE ACTIVATION OF LYSOSOMAL, CA2-ACTIVATED AND ATP-UBIQUITIN-DEPENDENT PROTEINASES IN THE UNWEIGHTED RAT SOLEUS MUSCLE()

Nine days of hindlimb suspension resulted in atrophy (55%) and loss of protein (53%) in rat soleus muscle due to a marked elevation in prote in breakdown (66%, P < 0.005). To define which proteolytic system(s) c ontributed to this increase, soleus muscles from unweighted rats were incubated in the presence of proteolytic inhibitors. An increase in ly sosomal and Ca2+-activated proteolysis (254%, P < 0.05) occurred in th e atrophying incubated muscles. In agreement with the measurements in vitro, cathepsin B, cathepsins B+L and m-calpain enzyme activities inc reased by 111%, 92% and 180% (P < 0.005) respectively in the atrophyin g muscles. Enhanced mRNA levels for these proteinases (P < 0.05 to P < 0.001) paralleled the increased enzyme activities, suggesting a trans criptional regulation of these enzymes. However, the lysosomal and Ca2 +-dependent proteolytic pathways accounted for a minor part of total p roteolysis in both control (9%) and unweighted rats (18%). Furthermore the inhibition of these pathways failed to suppress increased protein breakdown in unweighted muscle. Thus a non-lysosomal Ca2+-independent proteolytic process essentially accounted for the increased proteolys is and subsequent muscle wasting. Increased mRNA levels for ubiquitin, the 14 kDa ubiquitin-conjugating enzyme E2 (involved in the ubiquityl ation of protein substrates) and the C2 and C9 subunits of the 20 S pr oteasome (i.e. the proteolytic core of the 26 S proteasome that degrad es ubiquitin conjugates) were observed in the atrophying muscles (P < 0.02 to P < 0.001). Analysis of C9 mRNA in polyribosomes showed equal distribution into both translationally active and inactive mRNA pools, in either unweighted or control rats. These results suggest that incr eased ATP-ubiquitin-dependent proteolysis is most probably responsible for muscle wasting in the unweighted soleus muscle.