Cloning of a novel phospholipase C-δ isoform from pacific purple sea urchin ( Strongylocentrotus purpuratus) gametes and its expression during early embryonic development

Abstract

Calcium (Ca 2+) is a ubiquitous intracellular messenger, controlling a diverse range of cellular processes, including fertilization and development of the embryo. One of the key mechanisms involved in triggering intracellular calcium release is the generation of the second messenger inositol-1,4,5-phosphate (IP 3) by the phospholipase C (PLC) class of enzymes. Although five distinct forms of PLC have been identified in mammals (β, γ, δ, ε, and ζ), only one, PLCγ, has thus far been detected in echinoderms. In the present study, we describe the isolation of a cDNA encoding a novel PLC isoform of the delta (δ) subclass, PLC-δsu, from the egg of the Pacific purple sea urchin Strongylocentrotus purpuratus. We also demonstrate the presence of this PLC within the sperm and in the early embryo. The PLC-δsu cDNA (2.44 kb) encodes a 742 amino acid polypeptide with an open reading frame of 84.6 kDa and a p I of 6.04. All of the characteristic domains found in mammalian PLCδ isoforms (PH domain, EF hands, an X–Y catalytic region, and a C2 domain) are present in PLC-δsu. A homology search revealed that PLC-δsu shares most sequence identity with bovine PLCδ2 (39%). We present evidence that PLC-δsu is expressed in unfertilized eggs, fertilized eggs, and in the early embryo. In addition to Northern and polymerase chain reaction (PCR) analyses, in situ hybridization experiments further demonstrated that the embryonic regions within which the PLC-δsu transcript can be detected during early embryonic development are associated with the highest levels of proliferative activity, suggesting a possible involvement with metabolism or cell cycle regulation.