Structure of cDNA clones and genomic DNA FMRFamide-related peptides (FaRPs) in Helix

A complementary DNA (cDNA) library was prepared from poly(A)(+) RNA isolated from the central ganglia of Helix aspersa from which two classes of FaRP-encoding cDNA clones were identified by hybridization with the Aplysia FMRF-1 clone and oligonucleotides based on known Helix peptides. One type of cDNA (exemplified by HF-1) encodes only the tetrapeptides (FMRFamide and FLRFamide) and is very similar to the tetrapeptide-encoding precursors of other molluscan species. The other type of cDNA (represented by HF-4) encodes no tetrapeptides, but only N-terminally extended peptides, including all of the heptapeptides previously detected in the nervous system as well as some novel predicted peptides, which may be processed into free bio-active peptides. The overall structure of the precursor polypeptide encoded by HF-4 is markedly different from that encoded by HF-1 and more closely resembles the Drosophila FaRP precursor. Restriction digestion and hybridization analysis of genomic DNA indicates that each class of cDNA comes from a single genomic locus and that the two genomic loci span about 14 kbp. Parts of the genomic DNA sequence homologous to HF-1 were determined by PCR of Helix pomatia DNA. All of the coding sequence contained in HF-1 appears to be on one exon since it is contiguous in the genomic PCR products. In the coding region, the sequences from H. aspersa and H. pomatia are about 95% identical, but they are only about 80% identical in the noncoding region.