Investigation of the Met-267 Arg exchange in isoform 1 of the human plasma membrane calcium pump in patients with essential hypertension by the amplification-created restriction site technique

Abstract

 Alterations in Ca²⁺ homeostasis have been proposed to be a primary factor in the pathogenesis of essential hypertension. In this disease increased intracellular Ca²⁺ levels have repeatedly been reported in various cell types. Because of its prominent role in cellular calcium homeostasis in vascular smooth muscle cells, modifications of the plasma membrane Ca²⁺-ATPase (PMCA) pump have been suggested to contribute to an increased contractile tone of small blood vessels. This pump is a calmodulin-dependent Ca²⁺-ATPase that ejects Ca²⁺ from the cytosol into the extracellular space. Recently a mutational thymidine (T)→guanosine (G) transversion in isoform 1 of the PMCA has been identified resulting in the substitution of a methionine (Met) by an arginine (Arg) at amino acid position 267 in a highly conserved domain of the pump molecule. The aim of our study was to determine the prevalence of this polymorphism in the normal population and to investigate whether the Met-267 Arg occurs more frequently in patients with essential hypertension than in normotensives. To detect the mutational change we modified a method based on the technique of amplification-created restriction sites (ACRS) using three base exchanges in the diagnostic primer. Samples from 100 hypertensive and 60 normotensive subjects revealed a thymidine at nucleotide position 981. These data suggest that ACRS is feasible in spite of extensive primer modifications (e.g., three mismatched bases) in contrast to the previously used one or two and may therefore be conceptually suitable to detect almost any base changes in the genome. The described T→G transversion is a rare polymorphism and is presumably not related to common forms of essential hypertension.