Ubiquitin-conjugating enzyme E214k/HR6B is dispensable for increased protein catabolism in muscle of fasted mice.
Activated skeletal muscle proteolysis in catabolic states has been linked to an upregulation of the ATP-ubiquitin-dependent proteolytic system. Previous studies suggested that the N-end rule pathway is primarily responsible for the bulk of skeletal muscle proteolysis. The activity of this pathway is dependent on the 14-kDa ubiquitin-conjugating enzyme E2(14k) (HR6B) and the ubiquitin protein ligase Ubr1. To address the requirement of E2(14k) in muscle proteolysis, we examined muscle protein metabolism in wild-type (WT) mice and mice lacking the E2(14k) gene (KO) in fed and fasted (48 h) states. Baseline body weight, muscle mass, and protein content were similar, and these parameters decreased similarly upon fasting in the two genotypes. There were also no effects of genotype on the rate of proteolysis in soleus muscle. The fasting-induced increase in the amount of ubiquitinated proteins was the same in WT and KO mice. The absence of any significant effect of loss of E2(14k) function was not due to a compensatory induction of the closely related isoform HR6A. Total intracellular concentration of E2(14k) and HR6A in the WT mice was 290 +/- 40 nM, but the level in the KO mice (reflecting the level of HR6A) was 110 +/- 9 nM. This value is about threefold the apparent Michaelis-Menten constant (K(m)) of E2(14k) (approximately 40 nM) for stimulating conjugation in muscle extracts. Because the HR6A isoform has a K(m) of 16 nM for stimulating conjugation, the HR6A levels in the muscles of KO mice appear sufficient for supporting conjugation mediated by this pathway during fasting.
Adegoke
Olasunkanmi A J
OA
Polypeptide Laboratory, Department of Medicine, McGill University, Montreal, Quebec H3A 2B2, Canada.
Bédard
Nathalie
N
Roest
Henk P
HP
Wing
Simon S
SS
D000818:Animals; D001835:Body Weight / Q000502:physiology; D005215:Fasting / Q000502:physiology; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D008025:Ligases / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D009124:Muscle Proteins / Q000378:metabolism*; D018482:Muscle, Skeletal / Q000033:anatomy & histology / Q000378:metabolism*; D009929:Organ Size / Q000502:physiology; D010447:Peptide Hydrolases / Q000378:metabolism; D025801:Ubiquitin / Q000378:metabolism; D044763:Ubiquitin-Conjugating Enzymes
D007527:Isoenzymes; D009124:Muscle Proteins; D025801:Ubiquitin; C512786:Ube2a protein, mouse; C465394:Ube2b protein, mouse; D044763:Ubiquitin-Conjugating Enzymes; D010447:Peptide Hydrolases; D008025:Ligases