Cloning and characterization of cDNAs encoding trehalase from post-dormant embryos of the brine shrimp, Artemia franciscana

Abstract

To investigate post-dormant regulation of trehalose metabolism in the brine shrimp, we cloned and characterized two trehalase cDNAs from embryos of Artemia franciscana using a PCR probe corresponding to a highly conserved region among trehalases. The cDNAs consisted of 2496 and 2485 nucleotides, and had almost the same open reading frame encoding 703 amino acids which showed 46.6-42.6% similarities to trehalases of animals. The calculated molecular mass of the trehalase was 79,995 Da. The deduced sequence had a cleavable signal peptide, a cell adhesion motif, four potential N-glycosylation sites, trehalase signatures and a unique, long carboxyl terminal polypeptide containing a predicted transmembrane region and a potential cAMP-dependent phosphorylation site. Phylogenetic analyses showed a large divergence among trehalases of arthropods. Northern blots revealed the presence of two mRNAs. One of them, a 2.6 kb mRNA, was abundant in the dormant cysts and prenauplii. The other 5.0 kb mRNA was newly synthesized during post-dormant development. Possible mechanisms of trehalase regulation are presented on the basis of the results shown by the Northern blots and developmental changes of trehalase activity.