The ubiquitin-protein ligase Nedd4 targets Notch1 in skeletal muscle and distinguishes the subset of atrophies caused by reduced muscle tension.
Ubiquitination-dependent proteolysis is a fundamental process underlying skeletal muscle atrophy. Thus, the role of ubiquitin ligases is of great interest. There are no focused studies in muscle on the ubiquitin ligase Nedd4. We first confirmed increased mRNA expression in rat soleus muscles due to 1-14 days of hind limb unloading. Nedd4 protein localized to the sarcolemmal region of muscle fibers. Hind limb unloading, sciatic nerve denervation, starvation, and diabetes led to atrophy of soleus, plantaris, and gastrocnemius muscles, but only unloaded and denervated muscles showed a marked increase in Nedd4 protein expression. This increase was strongly correlated with decreased Notch1 expression, a known target of Nedd4 in other cell types. Overexpression of dominant negative Nedd4 in soleus muscles completely reversed the unloading-induced decrease of Notch1 expression, indicating that Nedd4 is required for Notch1 inactivation. Overexpression of wild-type Nedd4 in soleus muscles of weight bearing rats caused a decrease in Notch1 protein, indicating that Nedd4 is sufficient for Notch1 down-regulation. To further show that Notch1 is a Nedd4 substrate in muscle, conditional overexpression of Nedd4 in C2C12 myotubes induced ubiquitination of Notch1. This is the first finding of a Nedd4 substrate in muscle and of an ubiquitin ligase, the activity of which distinguishes disuse from cachexia atrophy.
Koncarevic
Alan
A
Department of Health Sciences, Boston University, Boston, Massachusetts, USA.
Jackman
Robert W
RW
Kandarian
Susan C
SC
D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D056827:Endosomal Sorting Complexes Required for Transport; D005260:Female; D015870:Gene Expression; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D019417:Hindlimb Suspension / Q000502:physiology; D007150:Immunohistochemistry; D009121:Muscle Denervation; D018485:Muscle Fibers, Skeletal / Q000166:cytology / Q000378:metabolism; D009129:Muscle Tonus / Q000502:physiology; D018482:Muscle, Skeletal / Q000378:metabolism* / Q000473:pathology / Q000503:physiopathology; D009133:Muscular Atrophy / Q000235:genetics / Q000378:metabolism* / Q000503:physiopathology; D032446:Myoblasts / Q000166:cytology / Q000378:metabolism; D000075702:Nedd4 Ubiquitin Protein Ligases; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D051881:Receptor, Notch1 / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012508:Sarcolemma / Q000378:metabolism; D014162:Transfection; D044767:Ubiquitin-Protein Ligases / Q000235:genetics / Q000378:metabolism*; D014452:Ubiquitins / Q000378:metabolism
D056827:Endosomal Sorting Complexes Required for Transport; D012333:RNA, Messenger; D051881:Receptor, Notch1; D011993:Recombinant Fusion Proteins; D014452:Ubiquitins; D049452:Green Fluorescent Proteins; C000618479:NEDD4L protein, rat; D000075702:Nedd4 Ubiquitin Protein Ligases; C000618484:Nedd4 protein, rat; D044767:Ubiquitin-Protein Ligases