title: A novel calmodulin-regulated Ca2+-ATPase (ACA2) from Arabidopsis with an N-terminal autoinhibitory domain.
abstract: To study transporters involved in regulating intracellular Ca2+, we isolated a full-length cDNA encoding a Ca2+-ATPase from a model plant, Arabidopsis, and named it ACA2 (Arabidopsis Ca2+-ATPase, isoform 2). ACA2p is most similar to a "plasma membrane-type" Ca2+-ATPase, but is smaller (110 kDa), contains a unique N-terminal domain, and is missing a long C-terminal calmodulin-binding regulatory domain. In addition, ACA2p is localized to an endomembrane system and not the plasma membrane, as shown by aqueous-two phase fractionation of microsomal membranes. ACA2p was expressed in yeast as both a full-length protein (ACA2-1p) and an N-terminal truncation mutant (ACA2-2p; Delta residues 2-80). Only the truncation mutant restored the growth on Ca2+-depleted medium of a yeast mutant defective in both endogenous Ca2+ pumps, PMR1 and PMC1. Although basal Ca2+-ATPase activity of the full-length protein was low, it was stimulated 5-fold by calmodulin (50% activation around 30 nM). In contrast, the truncated pump was fully active and insensitive to calmodulin. A calmodulin-binding sequence was identified within the first 36 residues of the N-terminal domain, as shown by calmodulin gel overlays on fusion proteins. Thus, ACA2 encodes a novel calmodulin-regulated Ca2+-ATPase distinguished by a unique N-terminal regulatory domain and a non-plasma membrane localization.
authors.affiliation: Department of Cell Biology, The Scripps Research Institute, La Jolla, California 92037, USA. Harper@Scripps.edu
authors.forename: J F
authors.initials: JF
authors.lastname: Harper
authors.forename: B
authors.initials: B
authors.lastname: Hong
authors.forename: I
authors.initials: I
authors.lastname: Hwang
authors.forename: H Q
authors.initials: HQ
authors.lastname: Guo
authors.forename: R
authors.initials: R
authors.lastname: Stoddard
authors.forename: J F
authors.initials: JF
authors.lastname: Huang
authors.forename: M G
authors.initials: MG
authors.lastname: Palmgren
authors.forename: H
authors.initials: H
authors.lastname: Sze
mesh_terms: D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000201:enzymology*; D001483:Base Sequence; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D002147:Calmodulin / Q000378:metabolism*; D018076:DNA, Complementary; D005816:Genetic Complementation Test; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D011485:Protein Binding; D012441:Saccharomyces cerevisiae / Q000235:genetics; D017386:Sequence Homology, Amino Acid
chemical_list: D002147:Calmodulin; D018076:DNA, Complementary; D010940:Plant Proteins; D000252:Calcium-Transporting ATPases