Identification of the promoter for the gene encoding the bifunctional enzyme, peptidylglycine alpha-amidating monooxygenase

DNA Cell Biol. 1996 Dec;15(12):1093-104. doi: 10.1089/dna.1996.15.1093.

Abstract

The gene encoding rat peptidylglycine alpha-amidating monooxygenase (PAM) contains 26 protein-coding exons. We identified two non-overlapping genomic clones encoding the 5' untranslated region (UTR) of the PAM gene. Exon 1 has 69 nucleotides flanked by perfect splice acceptor and donor sites, with a TATA motif 25 nucleotides upstream. Exon 0 lacks TATA or CAAT motifs and is embedded in a G + C-rich 800-nucleotide CpG island. The major products identified by RNase protection initiated in exon 0; only a minority of mRNAs initiated in exon 1. 5'-rapid amplification of cDNA ends (RACE) identified the same major transcriptional start sites in exon 0 in the atrium and neurointermediate pituitary. The 2.0-kb fragment upstream of exon 0 and the 1.3-kb fragment upstream of exon 1 were placed upstream of a luciferase-based reporter gene in both sense and antisense orientations. Expression of luciferase was observed in neuroendocrine and nonneuroendocrine cells with both sense constructs. A 0.2-kb fragment of the exon 0 PAM promoter containing multiple GC box elements supported expression of luciferase activity in all cell types. Expression of reporter genes in cells that do not normally express PAM suggests a need for more upstream or intronic information, a role for methylation, or a need for chromatin scaffolding for tissue-specific expression of the endogenous gene.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Base Sequence
  • Cell Line
  • Cloning, Molecular
  • DNA, Complementary / genetics
  • DNA, Recombinant
  • Exons / genetics
  • Gene Expression Regulation, Enzymologic / physiology
  • Genes
  • Male
  • Mixed Function Oxygenases / genetics*
  • Molecular Sequence Data
  • Multienzyme Complexes*
  • Promoter Regions, Genetic / genetics*
  • RNA, Messenger / genetics
  • Rats
  • Rats, Sprague-Dawley
  • Sequence Analysis, DNA
  • Testis
  • Transcription, Genetic / genetics

Substances

  • DNA, Complementary
  • DNA, Recombinant
  • Multienzyme Complexes
  • RNA, Messenger
  • Mixed Function Oxygenases
  • peptidylglycine monooxygenase