Cloning and characterization of a muscle isoform of a Na,K-ATPase alpha subunit (SNaK1) from Schistosoma mansoni.
A cDNA encoding a Na,K-ATPase alpha subunit homologue, designated SNaK1, was isolated from an adult cDNA library of Schistosoma mansoni. The 3.8 kb DNA contained a 3021 bp open reading frame potentially encoding a 1,007 amino acid protein that had an Mr of 111,817 and a pl of 5.48. Homology searches for SNaK1 revealed approximately 70% sequence identity with a variety of Na, K-ATPases from evolutionarily diverse organisms. SNaK1 is predicted to contain 10 transmembrane regions typical of this protein family as well as other conserved domains, such as the phosphorylation site and ATP binding domain. Antibodies raised against an amino terminal peptide detected the protein in membrane preparations of eggs, cercariae and adult males and females, suggesting a general role for SNaK1. The mobility of the protein differed in various life-stages suggestive of post-transcriptional or post-translational modification. Immunolocalization of SNaK1 in sections of adult worms using epifluorescence and electron microscopy, revealed antibody labelling in the subtegumental and peripheral layers. Strong staining was discernible in the peripheral muscle band indicating that SNaK1 plays a central role in muscle contraction in adult parasites and may be the primary target of ouabain action. Staining was also detected in the secretory bodies in sections of ducts in this region and over the RER of the presumed gastrodermis. Immunogold labelling was also localized over neuronal vesicles in axons associated with the peripheral muscle layer.
Skelly
P J
PJ
Department of Immunology and Infectious Diseases, Harvard School of Public Health, Boston, Massachusetts 02115, USA. psk@hsph.harvard.edu
Dougan
P M
PM
Maule
A
A
Day
T A
TA
Shoemaker
C B
CB
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D003001:Cloning, Molecular; D018509:DNA, Helminth / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D005260:Female; D006801:Humans; D007150:Immunohistochemistry; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics; D008297:Male; D016253:Microscopy, Immunoelectron; D008969:Molecular Sequence Data; D009132:Muscles / Q000201:enzymology*; D016133:Polymerase Chain Reaction; D012550:Schistosoma mansoni / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D012689:Sequence Homology, Nucleic Acid; D000254:Sodium-Potassium-Exchanging ATPase / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics*
D018509:DNA, Helminth; D007527:Isoenzymes; D000254:Sodium-Potassium-Exchanging ATPase