Copper efflux from murine microvascular cells requires expression of the menkes disease Cu-ATPase.

Abstract

Previously, we showed that the transport of Cu by PC12 pheochromocytoma cells and C6 glioma cells correlated with the expression of a Cu-transporting ATPase (Atp7a) that has been linked to Menkes disease. Here, we show that cerebrovascular endothelial (CVE) cells that comprise the blood-brain barrier (BBB) also express the gene for the Cu-ATPase. By using reverse transcription-polymerase chain reaction (RT-PCR) and primers designed from mouse Atp7a cDNA, we amplified a 925-bp and a 760-bp cDNA fragment from two extreme regions of Atp7a mRNA from murine CVE cells; 777 bp of the 925-bp fragment and 677 bp of the 760-bp fragment had a 99.7 and 100% sequence homology, respectively, with mouse Atp7a cDNA. The 777-bp sequences covered the heavy metal binding (Hmb) domain and the 677-bp fragment coded for residues at the -COOH terminus of Atp7a. A functional analysis showed that Cu efflux was blocked by the sulfhydryl reagent p-chloromercuribenzoate (p-CMB), a potential inhibitor of Atp7a function. This study provides strong evidence that a Cu-ATPase in the BBB controls the penetration of Cu into the brain and that lesions to the Cu-ATPase in CVE cells are a primary cause of low brain Cu levels in Menkes disease.

• Authors: Y Qian, E Tiffany-Castiglioni, J Welsh, E D Harris
• Journal: The Journal of nutrition (J Nutr) Vol. 128 Issue 8 Pg. 1276-82 (Aug 1998) ISSN: 0022-3166 [Print] United States
• PMID: 9687544 (Publication Type: Comparative Study, Journal Article, Research Support, U.S. Gov't, Non-P.H.S., Research Support, U.S. Gov't, P.H.S.)

Chemical References

• Atp7a protein, mouse
• Carrier Proteins
• Cation Transport Proteins
• DNA, Complementary
• Recombinant Fusion Proteins
• Copper
• RNA-Directed DNA Polymerase
• Adenosine Triphosphatases
• Copper-Transporting ATPases

Topics

• Adenosine Triphosphatases (chemistry, genetics, metabolism)
• Amino Acid Sequence
• Animals
• Base Sequence
• Biological Transport
• Blood-Brain Barrier
• Brain (blood supply)
• Carrier Proteins (chemistry, genetics, metabolism)
• Cation Transport Proteins
• Copper (metabolism)
• Copper-Transporting ATPases
• DNA, Complementary (chemistry)
• Endothelium, Vascular (enzymology)
• Gene Expression
• Menkes Kinky Hair Syndrome (enzymology)
• Mice
• Microcirculation (enzymology)
• Molecular Sequence Data
• Polymerase Chain Reaction
• RNA-Directed DNA Polymerase
• Recombinant Fusion Proteins
• Sequence Homology