Identification of a calmodulin-regulated soybean Ca2+-ATPase (SCA1) that is located in the plasma membrane

Ca2+-ATPases are key regulators of Ca2+ ion efflux in all eukaryotes. Anima l cells have two distinct families of Ca2+ pumps, with calmodulin-stimulate d pumps (type IIB pumps) found exclusively at the plasma membrane. In plant s, no equivalent type IIB pump located at the plasma membrane has been iden tified at the molecular level, although related isoforms have been identifi ed in non-plasma membrane locations. Here, we identify a plant cDNA, design ated SCA1 (for (s) under bar oybean (C) under bar a(2+)-(A) under bar TPase (1) under bar), that encodes Ca2+-ATPase and is located at the plasma memb rane. The plasma membrane localization was determined by sucrose gradient a nd aqueous two-phase membrane fractionations and was confirmed by the local ization of SCA1p tagged with a green fluorescent protein. The Ca2+-ATPase a ctivity of the SCA1p was increased approximately sixfold by calmodulin (K(1 /2)similar to N10 nM). Two calmodulin binding sequences were identified in the N-terminal domain. An N-terminal truncation mutant that deletes sequenc e through the two calmodulin binding sites was able to complement a yeast m utant (K616) that was deficient in two endogenous Ca2+ pumps. Our results i ndicate that sCA1p is structurally distinct from the plasma membrane-locali zed Ca2+ pump in animal cells, belonging instead to a novel family of plant type IIB pumps found in multiple subcellular locations. In plant cells fro m soybean, expression of this plasma membrane pump was highly and rapidly i nduced by salt (NaCl) stress and a fungal elicitor but not by osmotic stres s.