Molecular cloning, cDNA sequence, and localization of a prohormone convertase (PC2) from the Aplysia atrial gland.
Neuropeptides and peptide hormones are synthesized as part of larger precursor proteins that are processed post-translationally by subtilisin-related calcium-dependent prohormone convertases (PCs), frequently at multiple basic sites, to generate biologically active peptides. The atrial gland of Aplysia californica produces large quantities of egg-laying hormone (ELH)-related peptides, providing a unique opportunity to study prohormone processing. We have screened an Aplysia atrial gland cDNA library using a Lymnaea stagnalis PC2 probe and have isolated an Aplysia PC2-related 4.6-kb cDNA partial clone that was truncated on the 5' end. The remaining 5' atrial gland PC2 nucleotide sequence was obtained by reverse transcription/polymerase chain reaction (RT-PCR). The composite cDNA structure (5.6 kb) was deduced from sequence analysis of the RT-PCR product combined with the sequence obtained from the cDNA clone. The deduced cDNA of Aplysia atrial gland PC2 encoded a putative preproendoprotease of 653 amino acids that was evolutionarily related to other eukaryotic PC2s, and showed the strongest sequence identity with recently reported Aplysia nervous tissue PC2 sequences. In situ hybridization demonstrated extensive expression of PC2 in atrial gland secretory cells. The cDNA clone contained a relatively long 3'untranslated region (3'-UTR) of 3,632 nucleotides. Strikingly, the 3'-UTR also contained several major nucleotide repeat sequences including the microsatellite repeats, (CA)n and (TG)n, and a TA-rich region comprised largely of the triplet repeat (TTA)n. The characterized Aplysia PC2 is a candidate endoprotease that may play an important role in the processing of ELH-related precursors in the atrial gland and represents the first example of PC2 expression in exocrine tissue.
Nagle
G T
GT
Department of Anatomy and Neurosciences, University of Texas Medical Branch, Galveston 77555.
Garcia
A T
AT
Knock
S L
SL
Gorham
E L
EL
Van Heumen
W R
WR
Kurosky
A
A
D000595:Amino Acid Sequence; D000818:Animals; D001048:Aplysia / Q000201:enzymology / Q000235:genetics*; D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D005088:Exocrine Glands / Q000201:enzymology; D006325:Heart Atria; D008969:Molecular Sequence Data; D045706:Proprotein Convertase 2; D011498:Protein Precursors / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis; D012091:Repetitive Sequences, Nucleic Acid / Q000235:genetics; D015183:Restriction Mapping; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013381:Subtilisins / Q000737:chemistry / Q000235:genetics*
D018076:DNA, Complementary; D011498:Protein Precursors; D012333:RNA, Messenger; D013381:Subtilisins; D045706:Proprotein Convertase 2