p63/73 homologues in surf clam: novel signaling motifs and implications for control of expression.

To understand the role of p53 gene family members during invertebrate embryonic development, we used polymerase chain reaction (PCR) to identify p63/73 homologues in the marine mollusc Spisula solidissima. Here, we report the sequences of two distinct p63/73-like homologues, both cloned from Spisula embryos. The first, Ssp63/73alpha is 2699 nucleotide (nt); the second, Spp63/73beta is 3920 nt. The nucleotide sequences of the two variants are nearly identical up to their stop codons but diverge in their 3'-untranslated regions (UTRs). The deduced amino acid sequence of both Ssp63/73 variants is 597 amino acids, coding for a protein with predicted molecular weight of approximately 68 kDa. We conclude that the two unique transcripts, containing 3' UTRs of variable lengths, represent tandem alternate polyadenylation sites for the Ssp63/73 gene. While alternative splicing has been well documented in the p63/73 gene family, this is the first report of alternate polyadenylation site choice as a control point for p63/73 gene expression in any species. In order to identify specific post-transcriptional as well as post-translational signals potentially involved in regulation of p63/73-like expression, we compared Ssp63/p73 nucleotide and Ssp63/73 deduced amino acid sequences to corresponding regions of other mammalian and nonmammalian p63 and p73 homologues. Within the Spisula 3' UTRs we identified multiple AU-rich elements (AREs) which may control translation activation. Within the deduced amino acid sequence, we identified potential sites for sumoylation, a post-translational process that has been identified in mammalian p63 and p73 proteins. Identification of these novel signaling sites provides information about potential mechanisms controlling expression of multiple p63/73 isoforms during development.