Essential role for the Menkes ATPase in activation of extracellular superoxide dismutase: implication for vascular oxidative stress.

Extracellular superoxide dismutase (SOD3), a secretory copper enzyme, plays an important role in atherosclerosis and hypertension by modulating the levels of extracellular superoxide anion (O2*-) in the vasculature. Little is known about the mechanisms by which SOD3 obtains its catalytic copper cofactor. Menkes ATPase (MNK) has been shown to transport cytosolic copper to the secretory pathway in nonvascular cells. We performed the present study to determine whether MNK is required for the activation of SOD3 in the vasculature. Here we show that MNK was highly expressed in the various vascular tissues and cells. Aortas and cultured fibroblasts from MNK mutant (MNK(mut)) mice showed a marked decrease in specific activity of SOD3, but not SOD1 (cytosolic form), which was partially restored by copper addition. Copper treatment in wild-type cells promoted the direct interaction and colocalization of SOD3 with MNK in the trans-Golgi network (TGN), suggesting that MNK transports copper to SOD3 in the TGN. Aortas of MNK(mut) mice revealed a decrease in activity of SOD3, but not SOD1, in association with a robust increase in O2*- levels. Finally, both MNK and SOD3 proteins were highly expressed in the intimal lesions of atherosclerotic vessels. In conclusion, vascular MNK plays an essential role in full activity of SOD3 through transporting copper to SOD3 in the TGN, thereby regulating O2*- levels in the vasculature. These studies provide a novel insight into vascular MNK as a critical modulator of "superoxide" stress, which may contribute to cardiovascular disease.