Identification of P-Type ATPases in a Cell Line of AEDES Albolpicture C6/36

Abstract

Active cell membrane ions transport P-type-ATPases, which share a common acid-stable Asp residue that is phosphorilated during the pumping cycle, help cells maintain stable intracellular ion concentrations. The intracellular Na+ and K+ concentration is maintained by the operation of the Na+/K+ ATPase which transports 3 Na+ out- and 2 K+ in-wards balancing these ions leaks. The Na+/K+ ATPase is an ouabain sensitive P-type ATPase. The second Na+ pump is another ATPase which participates in Na+ transport across the membrane; it is ouabain insensitive but sensitive to bumetanide and furosemide [Pfluegers Arch: 316,1970;1; Biochim.Biophys.Acta: 901,1987;209]. The Na+/K+ ATPase is also present in Malpighian Tubules of Aedes [J.exp.Biol. 209,2006;4638]. P-type ATPases have been widely studied in high organisms. However, studies in invertebrates are needed. Na+/K+ ATPase has been identified in Drosophila melanogaster (EMBO J. 8,1989;193). However, Na+ ATPase reports are scarce. Our aim was to study P-Type ATPases in the C6/36 cell culture of Aedes alborpicture. ATPase activities were measured spectrophotometrically as liberated inorganic phosphorus. We found: (i) a Na+ stimulated ATPase in presence of ouabain without K+ which we associate with the second Na+ pump; (ii) a Na+ and K+ stimulated ATPase activity which is inhibited in the presence of ouabain which corresponds to the Na+/K+ ATPase; and (iii) that the consensus sequences of P-type ATPases were present on the genomic material of C6/36. We conclude that the differences of ATPase activities of C6/36 in the presence of Mg2+ plus Na+ plus ouabain (without K+) and of Mg2+ plus Na+ plus K+ (without ouabain) indicate the presence of Na+ and of Na+/K+ ATPases, respectively. Additional molecular biology experiments confirm the existence of P-type ATPases in this cellular culture. Correction et al.Biophysical JournalMay 04, 2011In Brief1334-Pos. Full-Text PDF Open Archive