ABSTRACTHuman cDNAs encoding the precursor to pituitary adenylate cyclase activating polypeptide (PACAP) were cloned from human testis and cerebral cortex cDNA libraries. Nucleotide sequencing revealed that cDNA from the testis library encoded the entire precursor for PACAP, while cDNA from the brain library represented only the carboxy-terminal half of the precursor. The predicted human PACAP precursor consisted of 176 amino acid residues and was very similar to the ovine one (82). Both human and ovine precursors contained both PACAP and another peptide, PACAP-related peptide (PRP), having 29 amino acids. PACAP and PRP were preceded and followed by paired basic amino acids, recognized as important for post-translational processing. The PACAP precursor resembles the vasoactive intestinal peptide (VIP) precursor, which contains VIP and peptide histidine methionine/isoleucine amide (PHM/PHI). Structurally, PRP had some similarity to PHM/PHI, growth hormone-releasing hormone (GRH) and PACAP. Northern blot analysis indicated that a 3.0-kb transcript was expressed in the ovine hypothalamus. Tissue distribution of PACAP mRNA was also clarified in the rat. Southern blot analysis of human genomic DNA gives single bands with six restriction enzymes, indicating that a single copy of the PACAP gene is contained in a haploid genome. The cDNA for human PACAP precursor was expressed using COS-7 and Chinese hamster ovary (CHO) cells. Immunoreactive PACAP was secreted into the culture media of both transfected cell lines.
展开▼
