N-terminally extended FMRFamide-related peptides of Helix aspersa: processing of the precursor protein and distribution of the released peptides.
Sequencing of cDNA clones reveals a precursor protein that can be processed into 10 different hepta-FaRPs. Two of the peptides are previously undescribed and are N-terminally extended forms of-YMRFamide, making them the only methionine-containing peptides in the precursor. They are separated from the main cluster of hepta-FaRPs by a recognition site (RQKR) for the Golgi-resident proteolytic enzyme furin. Antisera raised against the synthetic peptide KQDPFLRFGK specifically stain the clusters of neurons in the parietal ganglia that have been shown to contain hepta-FaRP mRNA. These antisera recognize two major protein bands of 35 and 23 kDa on immunoblots. Evidence is presented to identify the larger band as the precursor protein and the smaller band as the fragment containing the main cluster of hepta-FaRPs produced after furin cleavage. A series of immunostaining bands of 22-13 kDa suggests sequential and non-preferential N- or C-terminal cleavage at the mainly monobasic (K and R) sites that link all of the peptide sequences throughout the 23-kDa fragment, to yield the preamidated hepta-FaRPs. Immunostaining of sections shows punctate staining in the perikarya of the parietal cluster neurons commensurate with label within the endoplasmic reticulum and Golgi apparatus. Staining is followed through the axons to many fibers in the nerve trunks and is picked up as fine processes within the skin. These observations indicate that the antiserum used here recognizes one or more of the processed hepta-FaRPs, a view confirmed by radioimmunoassay. The abundance of immunoreactive fibers within the skin suggests a major role for the peptides in this tissue.
Cottrell
G A
GA
School of Biological and Medical Sciences, University of St. Andrews, Fife, Scotland.
Lutz
E M
EM
Price
D A
DA
Sommerville
J
J
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D018076:DNA, Complementary / Q000235:genetics; D019835:FMRFamide; D006372:Helix, Snails / Q000378:metabolism*; D007447:Invertebrate Hormones / Q000378:metabolism; D015335:Molecular Probes / Q000235:genetics; D008969:Molecular Sequence Data; D009479:Neuropeptides / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism*; D011498:Protein Precursors / Q000235:genetics / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D012333:RNA, Messenger / Q000235:genetics; D011863:Radioimmunoassay; D014018:Tissue Distribution
D018076:DNA, Complementary; D007447:Invertebrate Hormones; D015335:Molecular Probes; D009479:Neuropeptides; D010446:Peptide Fragments; D011498:Protein Precursors; D012333:RNA, Messenger; D019835:FMRFamide