A C-terminal, calmodulin-like regulatory domain from the plasma membrane Ca2+-pumping ATPase.

Abstract

A cDNA that encodes what appears to be the inhibitory domain of the plasma membrane calcium-pumping ATPase (Ca2+-ATPase) has been isolated by screening a Agtll bovine brain cDNA library with antibodies prepared against the human erythrocyte membrane Ca2 + -ATPase. This screening resulted in isolation of a bacteriophage containing a 1.5-kilobase cDNA insert encoding a 71-residue polypeptide, the remainder being a large 3' terminal noncoding region. A portion of this deduced peptide sequence was identical to that of a peptide isolated from a V8 protease digest of the human erythrocyte Ca2+-ATPase except for 1 residue. Antibodies purified by immunoabsorption to the fusion protein containing this cDNA-encoded polypeptide reacted only with those fragments of a limited trypsin digest of the human erythrocyte Ca2+-ATPase that contain the inhibitory domain. Moreover, these antibodies were able to partially stimulate basal enzyme activity and block further activation by calmodulin. The encoded polypeptide bears homology to the glutamic acid-rich regions N-terminal to the Ca2+-binding loops of calmodulin and to a lesser extent with the loops themselves. This encoded polypeptide also represents the C terminus of the Ca2+-ATPase. Portions of the isolated cDNA were homologous to the 3' noncoding region of the sarcoplasmic reticulum Ca2+-ATPase cDNA, indicating a possible mechanism for the evolution of these distinct membrane Ca2+ pumps.

Recent advances in the molecular characterization of plasma membrane Ca2+ pumps

“…Screening of various rat, bovine and human cDNA libraries has so far led to the identification of at least five different mammalian PMCA isoforms that are encoded by separate genes (Brandt et al, 1988;Shull & Greeb, 1988;Greeb & Shull, 1989;Strehler et al, 1989aStrehler et al, , 1990. The genes for two human isoforms, hPMCA1 and hPMCA4, have recently been mapped in the genome and were shown to be localized on chromosomes 12 and 1, respectively (S. Olson, M.G.…”

Thapsigargin, a tumor promoter, discharges intracellular Ca2+ stores by specific inhibition of the endoplasmic reticulum Ca2(+)-ATPase.

“…Increasingly, thapsigargin is used as a probe for intracellular Ca2+-storage and -release processes ( ple, the plasma membrane Ca2+ pump has been shown to be sensitive to cGMP-dependent protein kinase (32), Ca2+/ calmodulin (33), and the hormone glucagon (34); the cardiac SR Ca2+-ATPase is regulated by phospholamban, which is a substrate for multiple signal-transduction kinases (35); and skeletal muscle SR Ca2+-ATPase has been proposed to be directly responsible for Ca2+-dependent Ca2+ efflux from SR (36), a situation directly analogous to the ER Ca2+-ATPase acting as an induced Ca2+ leak channel.…”

Regulation of Platelet Plasma Membrane Ca2+-ATPase by cAMP-dependent and Tyrosine Phosphorylation

“…Polyclonal antibodies against human erythrocyte PMCA were raised in rabbits as described previously (10), and monoclonal anti-PMCA antibody 5F10 (11) was obtained from Affinity BioReagents (Neshanic Station, NJ). Isoform-specific anti-PMCA antibodies (9) were kindly provided by Dr. Ernesto Carafoli (Swiss Federal Institute of Technology, Zurich, Switzerland).…”