Copper downregulates neprilysin activity through modulation of neprilysin degradation.
Copper plays a central role in conserved processes such as respiration, and in highly specialized processes, such as protein modification. The metalloprotease neprilysin (NEP) degrades a variety of bioactive peptides and is involved in many physiological processes. However, very little is known about the regulation of NEP activity. In the current study, we focused on the effect of Cu2+ on the enzymatic activity and protein stability of NEP. Using mouse neuroblastoma N2a cells, we found that the enzymatic activity of NEP was decreased by treatment with Cu2+ in a dose- and time-dependent manner. In our investigation of the mechanism by which Cu2+ downregulates NEP enzyme activity, we found that treatment with Cu2+ caused a decrease in the level of NEP as determined by Western blot analysis. Quantitative analysis of NEP mRNA with RT-PCR excluded the possibility that Cu2+ downregulates NEP protein at the gene transcription level. Moreover, specific proteasome inhibitors, MG132 and lactacystin, blocked the turnover of NEP, whereas inhibitors of lysosome had no significant effect, suggesting that Cu2+-induced degradation of NEP is via a proteasome pathway. Taken together, our data suggest that copper downregulates NEP activity through modulation of NEP protein degradation.
Li
Mi
M
Neuroscience Research Institute & Department of Neurobiology, Key Laboratory for Neuroscience Ministry of Education, Key Laboratory for Neuroscience Ministry of Public Health, Health Science Center, Peking University, Beijing, China.
Sun
Miao
M
Liu
Yi
Y
Yu
Jia
J
Yang
Huan
H
Fan
Dongsheng
D
Chui
Dehua
D
D000818:Animals; D045744:Cell Line, Tumor; D003300:Copper / Q000502:physiology*; D015536:Down-Regulation / Q000187:drug effects / Q000502:physiology*; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D051379:Mice; D015260:Neprilysin / Q000037:antagonists & inhibitors* / Q000378:metabolism* / Q000502:physiology; D046988:Proteasome Endopeptidase Complex / Q000502:physiology; D055550:Protein Stability; D015398:Signal Transduction / Q000502:physiology
D003300:Copper; D015260:Neprilysin; D046988:Proteasome Endopeptidase Complex