Ca(2)+-ATPases are key regulators of Ca(2+) ion efflux in all eukaryotes. Animal cells have two distinct families of Ca(2+) pumps, with calmodulin-stimulated pumps (type IIB pumps) found exclusively at the plasma membrane. In plants, no equivalent type IIB pump located at the plasma membrane has been identified at the molecular level, although related isoforms have been identified in non-plasma membrane locations. Here, we identify a plant cDNA, designated SCA1 (for soybean Ca(2+)-ATPase 1), that encodes Ca(2+)-ATPase and is located at the plasma membrane. The plasma membrane localization was determined by sucrose gradient and aqueous two-phase membrane fractionations and was confirmed by the localization of SCA1p tagged with a green fluorescent protein. The Ca(2+)-ATPase activity of the SCA1p was increased approximately sixfold by calmodulin (K(1/2) approximately 10 nM). Two calmodulin binding sequences were identified in the N-terminal domain. An N-terminal truncation mutant that deletes sequence through the two calmodulin binding sites was able to complement a yeast mutant (K616) that was deficient in two endogenous Ca(2+) pumps. Our results indicate that SCA1p is structurally distinct from the plasma membrane-localized Ca(2+) pump in animal cells, belonging instead to a novel family of plant type IIB pumps found in multiple subcellular locations. In plant cells from soybean, expression of this plasma membrane pump was highly and rapidly induced by salt (NaCl) stress and a fungal elicitor but not by osmotic stress.

Ca 2 ؉ -ATPases are key regulators of Ca 2 ؉ ion efflux in all eukaryotes. Animal cells have two distinct families of Ca 2 ؉ pumps, with calmodulin-stimulated pumps (type IIB pumps) found exclusively at the plasma membrane. In plants, no equivalent type IIB pump located at the plasma membrane has been identified at the molecular level, although related isoforms have been identified in non-plasma membrane locations. Here, we identify a plant cDNA, designated SCA1 (for soybean Ca 2 ؉ -ATPase 1), that encodes Ca 2 ؉ -ATPase and is located at the plasma membrane. The plasma membrane localization was determined by sucrose gradient and aqueous two-phase membrane fractionations and was confirmed by the localization of SCA1p tagged with a green fluorescent protein. The Ca 2 ؉ -ATPase activity of the SCA1p was increased approximately sixfold by calmodulin ( K 1/2 ‫ف‬ 10 nM). Two calmodulin binding sequences were identified in the N-terminal domain. An N-terminal truncation mutant that deletes sequence through the two calmodulin binding sites was able to complement a yeast mutant (K616) that was deficient in two endogenous Ca 2 ؉ pumps. Our results indicate that SCA1p is structurally distinct from the plasma membrane-localized Ca 2 ؉ pump in animal cells, belonging instead to a novel family of plant type IIB pumps found in multiple subcellular locations. In plant cells from soybean, expression of this plasma membrane pump was highly and rapidly induced by salt (NaCl) stress and a fungal elicitor but not by osmotic stress. INTRODUCTIONCa 2 ϩ plays a central role as a second messenger in signal transduction of all eukaryotes (Bootman and Berrige, 1995;Clapham, 1995). The transient increase of cytosolic free Ca 2 ϩ concentration, [Ca 2 ϩ ] cyt , is correlated with a variety of external signals such as touch, temperature shift, abscisic acid, auxin, red light, fungal elicitors, salinity/drought, anoxia, gravity, gibberellic acid, cytokinin, oxidative stress, and hypoxia (Bush, 1995; Sanders et al., 1999). In turn, the increased [Ca 2 ϩ ] cyt triggers many signal transduction pathways, including the regulation of enzyme activity, ion channel activity, and gene expression, which results in diverse cellular responses (Bush, 1995). In addition to its role as a second messenger, Ca 2 ϩ also is important in regulating the processing of proteins in secretory pathways (Rudolph et al., 1989). Thus, cells require carefully regulated transport systems to control [Ca 2 ϩ ] in the cytoplasm and endomembrane compartments. Influx of Ca 2 ϩ to the cytosol occurs as a "downhill" transport through Ca 2 ϩ channels (Sanders et al., 1999). In contrast, the efflux of Ca 2 ϩ from the cytosol is mediated by two active Ca 2 ϩ transporters-Ca 2 ϩ pumps and Ca 2 ϩ /H ϩ antiporters-which are powered by ATP hydrolysis and proton motive force, respectively (Bush, 1995). In plants, Ca 2 ϩ -ATPases are believed to be a major Ca 2 ϩ transporter for the endoplasmic reticulum (ER), Golgi apparatus, vacuole, plastid inner membrane, and plasma membrane (Sander...

The infecting hyphae of Phytophthora capsici grew intercellularly in infected tissues of roots and stems of pepper (Capsicum annuum). The vascular tissues were not markedly disorganized even when heavily infected. Intercellularly growing hyphae penetrated the host cells by forming haustorium‐like bodies. The consistent features of ultrastructural changes in infected tissues of pepper roots and stems were degeneration of cell organelles and dissolution of host cell walls. The cytoplasm detached from the cell wall aggregated abundantly around some haustorium‐like bodies or the penetration sites of fungal hyphae. The host cell walls were palely stained, thinned and swollen, possibly being biochemically altered by the action of fungal macerating enzymes. Electron‐dense, wall‐like material was apposed on the outer wall of xylem vessel contacted by fungal hyphae. The infecting hyphae were also surrounded by granular, dark‐staining cytoplasm. Characteristics of host cell responses to the invading P. capsici were the deposition of papilla‐like material on host cell walls next to hyphae and the encasement of haustorium‐like bodies with wall appositions.