Molecular cloning of an intracellular P-type ATPase from Dictyostelium that is up-regulated in calcium-adapted cells.
Results from a number of laboratories suggest that intracellular Ca2+ is involved in the regulation of Dictyostelium discoideum growth and development. To learn more about the regulation and function of intracellular Ca2+ in this organism, we have cloned and sequenced cDNAs that encode a putative P-type Ca2+ ATPase designated patA. The deduced protein product of this gene (PAT1) has a calculated molecular mass of 120,718 daltons. It exhibits about 46% amino acid identity with Ca2+ ATPases of the plasma membrane Ca2+ ATPase family and lower identity with sarco(endo)plasmic reticulum Ca2+ ATPase family members and monovalent cation pumps. However, PAT1 lacks the highly conserved calmodulin-binding domain present in the C-terminal region of most plasma membrane Ca2+ ATPase-type enzymes. When Dictyostelium amoebae are adapted to grow in the presence of 80 mM CaCl2, both the patA message and protein product are up-regulated substantially. These cells also exhibit an increase in the rate and magnitude of intracellular P-type Ca2+ uptake activity. Immunofluorescence analysis indicates that PAT1 colocalizes with bound calmodulin to intracellular membranes, probably components of the contractile vacuole complex. The presence of PAT1 on the contractile vacuole suggests that in Dictyostelium this organelle might function in Ca2+ homeostasis as well as in water regulation.
D000064:Acclimatization; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D015153:Blotting, Western; D002118:Calcium / Q000494:pharmacology*; D000252:Calcium-Transporting ATPases / Q000032:analysis / Q000096:biosynthesis* / Q000737:chemistry; D002147:Calmodulin / Q000378:metabolism; D003001:Cloning, Molecular; D017931:DNA Primers; D018076:DNA, Complementary; D004023:Dictyostelium / Q000187:drug effects / Q000201:enzymology* / Q000502:physiology; D019084:Fluorescent Antibody Technique, Indirect; D015971:Gene Expression Regulation, Enzymologic; D005800:Genes, Fungal; D007700:Kinetics; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D015183:Restriction Mapping; D017386:Sequence Homology, Amino Acid
D002147:Calmodulin; D017931:DNA Primers; D018076:DNA, Complementary; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D000252:Calcium-Transporting ATPases; D002118:Calcium