Expression of the Menkes gene homologue in mouse tissues lack of effect of copper on the mRNA levels

123

While molecular mechanisms for iron entry and storage within cells have been elucidated, no system to mediate iron efflux has been heretofore identified. We now describe an ATP requiring iron transporter in mammalian cells. 55 Fe is transported into microsomal vesicles in a Mg-ATP-dependent fashion. The transporter is specific for ferrous iron, is temperature-and time-dependent, and detected only with hydrolyzable nucleotides. It differs from all known ATPases and appears to be a P-type ATPase. The Fe-ATPase is localized together with heme oxygenase-1 to microsomal membranes with both proteins greatly enriched in the spleen. Iron treatment markedly induces ATP-dependent iron transport in RAW 264.7 macrophage cells with an initial phase that is resistant to cycloheximide and actinomycin D and a later phase that is inhibited by these agents. Iron release, elicited in intact rats by glycerol-induced rhabdomyolysis, induces ATP-dependent iron transport in the kidney. Mice with genomic deletion of heme oxygenase-1 have selective tissue iron accumulation and display augmented ATP-dependent iron transport in those tissues that accumulate iron.

Section: Discussionmentioning

confidence: 72%

A Mammalian Iron ATPase Induced by Iron

Barañano

Wolosker

Bae

et al. 2000

123

“…For Northern blot analysis, fractionation of ϳ10 g of RNA on a 1% (w/v) agarose gel containing 1ϫ MOPS buffer (pH 7) (20 mM MOPS, 2 mM NaOAc, 1 mM EDTA) and 0.66 M formaldehyde, transfer to a Hybond-Nϩ membrane (Amersham Biosciences), and hybridization of labeled probe DNA were carried out as previously described (41). Membranes were exposed to an imaging plate (Fujifilm) overnight at 4°C, and bands were visualized using Bio-imaging Analyzer Systems BAS-1800 (Raytest Isotopenmessgeraete).…”

Section: Methodsmentioning

confidence: 99%

A Single PDZ Domain Protein Interacts with the Menkes Copper ATPase, ATP7A

Stephenson

Dubach

Lim

et al. 2005

Self Cite

The homeostatic regulation of essential elements such as copper requires many proteins whose activities are often mediated and tightly coordinated through protein-protein interactions. This regulation ensures that cells receive enough copper without intracellular concentrations reaching toxic levels. To date, only a small number of proteins implicated in copper homeostasis have been identified, and little is known of the protein-protein interactions required for this process. To identify other proteins important for copper homeostasis, while also elucidating the protein-protein interactions that are integral to the process, we have utilized a known copper protein, the copper ATPase ATP7A, as a bait in a yeast two-hybrid screen of a human cDNA library to search for interacting partners. One of the ATP7A-interacting proteins identified is a novel protein with a single PDZ domain. This protein was recently identified to interact with the plasma membrane calcium ATPase b-splice variants. We propose a change in name for this protein from PISP (plasma membrane calcium ATPase-interacting single-PDZ protein) to AIPP1 (ATPase-interacting PDZ protein) and suggest that it represents the protein that interacts with the class I PDZ binding motif identified at the ATP7A C terminus. The interaction in mammalian cells was confirmed and an additional splice variant of AIPP1 was identified. This study represents an essential step forward in identifying the proteins and elucidating the network of protein-protein interactions involved in maintaining copper homeostasis and validates the use of the yeast two-hybrid approach for this purpose.

“…The Menkes (MNK) 1 protein (ATP7A) is a copper-translocating P-type ATPase expressed in most tissues except the liver (5)(6)(7)(8). Mutations in the Menkes gene that cause the loss of function of the MNK protein result in Menkes disease in humans, a potentially lethal X-linked disorder associated with severe systemic copper deficiency.…”

mentioning

confidence: 99%

“…Apart from eight highly conserved domains, the structure of human copper P-type ATPases differs considerably from other enzymes of that family (17). The most prominent feature of human Menkes protein and the related Wilson (WND) protein, a copper P-type ATPase expressed in the liver, is six repeats of the putative metalbinding motifs (GMXCXXC) at the N-terminal domain (5)(6)(7)(8).…”

mentioning

confidence: 99%

The Regulation of Catalytic Activity of the Menkes Copper-translocating P-type ATPase

Voskoboinik

Mar

Strausak

et al. 2001

112

113

The Menkes protein is a transmembrane copper translocating P-type ATPase. Mutations in the Menkes gene that affect the function of the Menkes protein may cause Menkes disease in humans, which is associated with severe systemic copper deficiency. The catalytic mechanism of the Menkes protein, including the formation of transient acylphosphate, is poorly understood. We transfected and overexpressed wild-type and targeted mutant Menkes protein in yeast and investigated its transient acyl phosphorylation. We demonstrated that the Menkes protein is transiently phosphorylated by ATP in a copper-specific and copper-dependent manner and appears to undergo conformational changes in accordance with the classical P-type ATPase model. Our data suggest that the catalytic cycle of the Menkes protein begins with the binding of copper to high affinity binding sites in the transmembrane channel, followed by ATP binding and transient phosphorylation. We propose that putative copper-binding sites at the N-terminal domain of the Menkes protein are important as sensors of low concentrations of copper but are not essential for the overall catalytic activity.Copper is an essential trace element: its ability to redox cycle between Cu(I) and Cu(II) states is utilized by cuproenzymes participating in redox reactions. However, these same properties make excess copper toxic to biological systems (1). Finely tuned complex mechanisms of copper homeostasis have evolved to allow the regulated uptake of copper, its delivery to target proteins, and detoxification by chelation and/or efflux from the cell (2-4). Copper-translocating P-type ATPases found in a variety of organisms are implicated in the delivery of copper to some cuproenzymes and in the efflux of copper from the cell (2-4).The Menkes (MNK) 1 protein (ATP7A) is a copper-translocating P-type ATPase expressed in most tissues except the liver (5-8). Mutations in the Menkes gene that cause the loss of function of the MNK protein result in Menkes disease in humans, a potentially lethal X-linked disorder associated with severe systemic copper deficiency. Menkes patients suffer from neurological and connective tissue abnormalities as a result of copper deficiency, which reduces the activity of copper-dependent enzymes (9). Through clinical and laboratory studies on Menkes disease patients, the role of the MNK protein in the absorption of dietary copper from gut epithelium, delivery of copper to cuproenzymes, and efflux from the cell were established (9, 10).P-type ATPases are multispanning membrane proteins that translocate ions (e.g. H ϩ , Na, and Cd 2ϩ ) across biological membranes against an electrochemical and concentration gradient using ATP as an energy source (11, 12). The catalytic cycle of P-type ATPases is characterized by the coupled reactions of cation translocation and ATP hydrolysis with a transient aspartyl phosphate formed as a part of the reaction cycle. The phosphorylation results in the enzyme changing its conformation from the high affinity cation and nucleotide binding s...