Title: AN APYRASE GENE ENCODING A CALCIUM-DEPENDENT, STRUCTURAL HOMOLOGUE OF THE SALIVARY APYRASES FROM BLOOD FEEDING ARTHROPODS HAS BEEN IDENTIFIED IN OSTERTAGIA THIRD AND FOURTH STAGE LARVAE.
Submitted to: American Association of Veterinary Parasitologists
Publication Type: Abstract Only
Publication Acceptance Date: May 20, 2006
Publication Date: July 18, 2006
Citation: Zarlenga, D.S., Gasbarre, L.C. 2006. An apyrase gene encoding a calcium-dependent, structural homologue of the salivary apyrases from blood feeding arthropods has been identified in ostertagia third and fourth stage larvae [abstract]. American Association of Veterinary Parasitologists.
Apyrases (ATP diphosphohydrolase) comprise a ubiquitous class of glycosylated enzymes involved in hydrolyzing extracellular nucleoside di- and triphosphates to mononucleotides and orthophosphate in the presence of Mg 2+ and/or Ca 2+. To date, most apyrases can be structurally linked to the yeast GTPase/CD39 which is part of the heat shock70/sugar kinase/actin superfamily. This group of proteins has been associated with motility, adhesion, secretion, regulation of hemostasis, non-synaptic information transfer and platelet formation. A second class of soluble apyrases which counteract the blood-clotting mechanisms of the host has been identified in hematophagous arthropods. Herein, we identified an apyrase gene that structurally conforms to the soluble, calcium-activated apyrases found in arthropods, by immunologically screening an Ostertagia ostertagi L4 cDNA expression library with hyperimmune bovine serum. Host antibodies to the O. ostertagi apyrase (OoAP) were detected as soon as 14 dpi and increased through 30 dpi. Mouse antibodies generated against the recombinant protein (rOoAP) were used for immunohistochemical and Western blot analyses. Studies demonstrated that OoAP is produced or collects within the muscle layer below the hypodermis, and is predominantly found in L3 and minimally expressed in L4. Also, mouse anti-rOoAP recognized conserved amino acid epitopes in L3 antigen from Haemonchus, Cooperia, and Oesophagostomum. With respect to activity, rOoAP was found to be calcium-dependent with a pH maximum between 6.5 and 7.5 where the greatest functional activity was observed on ATP, ADP, UTP and UDP. Little or no activity was observed when either CTP or AMP were used as substrates, respectively. Striking correlations will be raised between the ability of extracellular ATP and orthophosphate to regulate muscle development and movement, and the role this class of apyrases may play in the ontogeny of O. ostertagi.