A Conditional Mutation Affecting Localization of the Menkes Disease Copper ATPase

Abstract

Copper is an essential co-factor for several key metabolic processes. This requirement in humans is underscored by Menkes disease, an X-linked copper deficiency disorder caused by mutations in the copper transporting P-type ATPase, MNK. MNK is located in the transGolgi network where it transports copper to secreted cuproenzymes. Increases in copper concentration stimulate the trafficking of MNK to the plasma membrane where it effluxes copper. In this study, a Menkes disease mutation, G1019D, located in the large cytoplasmic loop of MNK, was characterized in transfected cultured cells. In copper-limiting conditions the G1019D mutant protein was retained in the endoplasmic reticulum. However, this mislocalization was corrected by the addition of copper to cells via a process that was dependent upon the copper binding sites at the N-terminal region of MNK. Reduced growth temperature and the chemical chaperone, glycerol, were found to correct the mislocalization of the G1019D mutant, suggesting this mutation interferes with protein folding in the secretory pathway. These findings identify G1019D as the first conditional mutation associated with Menkes disease and demonstrate correction of the mislocalized protein by copper supplementation. Our findings provide a molecular framework for understanding how mutations that affect the proper folding of the MNK transporter in Menkes patients may be responsive to parenteral copper therapy.

Mutations associated with Charcot–Marie–Tooth disease cause SIMPLE protein mislocalization and degradation by the proteasome and aggresome–autophagy pathways

“…First, mutation-induced mislocalization away from the early endosomal membrane would remove SIMPLE protein from its site of action, thereby contributing to CMT pathogenesis by a lossof-function mechanism (Kim et al, 2002;Lupo et al, 2009;Roberts et al, 2010). Although the cellular function of SIMPLE is unknown, it has been proposed that SIMPLE might participate in protein sorting at the early endosome (Shirk et al, 2005).…”

In and Out of the ER: Protein Folding, Quality Control, Degradation, and Related Human Diseases

“…One indication that the folding defect of the mutant protein may be reverted by the use of chemical chaperones is the increase of the polypeptide folding efficiency upon expression at lower temperature (72,195). Another is the increase in the export of the select functional polypeptide from the ER upon inhibition of ERAD (392) as the folding and degradation processes are for many proteins in kinetic competition (268a).…”

“…In any case, studies with chemical chaperones offered an important proof of principle and paved the way for characterization of several, substrate-specific pharmacological chaperones that proved efficient in cultured cells or that have already been used in animal and/or clinical trials (22,58,232,271). Here are a few selected examples: V2 receptor antagonists rescued mutant vasopressin receptor from proteasomal degradation (272); gonadotropin-releasing hormone (GnRH) peptidomimetics rescued secretion of several GnRH mutants (177); the cromophore retinal rescued maturation of mutant opsin (285); specific inhibitors facilitated maturation and transport of mutant voltage-gated potassium channels (93), beta-glucocerebrosidase (334), and ␣-D-galactosidase (90); copper supplementation alleviated maturation defects of the mutant copper transporting ATPase (195); and 4-phenylbutyrate corrected folding defects of several cystic fibrosis channel mutations (434).…”

Dynamics of endogenous ATP7A (Menkes protein) in intestinal epithelial cells: copper-dependent redistribution between two intracellular sites

“…30,48,59,60). Many mutations causing Menkes and Wilson diseases have been identified and studies conducted to understand a specific mutation's effect on protein expression, stability, or function (10,18,19,40,61). In addition, studies of the cellular basis of the normal proteins' dual functions have revealed that both ATP7A and ATP7B proteins change their intracellular locations in response to changes in copper levels (30).…”