Characterization of the promoter region of the rat neprilysin gene

Chingwen Li, Louis B. Hersh

Research output: Contribution to journalArticlepeer-review

15 Scopus citations


The neprilysin gene is composed of three distinct 5' noncoding exons which can be joined to the first coding exon to generate multiple mRNA species, all encoding the same protein. Genomic fragments containing upstream sequences of each of these three noncoding exons from the rat neprilysin gene were subcloned in the promoterless vector pXp1, which contains the luciferase reporter gene. Expression was compared between a neprilysin positive human spinal cord cell line, HSC-2, and neprilysin negative lines MCF-7, a human breast adenocarcinoma cell line and Hep G2, a human liver carcinoma cell line. The first and second promoter regions showed high activity in the positive cell line, but low activity in the negative cell lines. An analysis of the exon 1 promoter region showed that the proximal 85 nucleotides exhibited basal promoter activity. An enhancer-like sequence was found to be located within a 22-bp fragment located at -136 to -115. Scanning mutagenesis of a 29-bp fragment containing the enhancer-like sequence showed that changes in each 5- or 6-bp segment throughout this fragment decreased activity; however, mutations of the segment encompassing positions 19 to 24 eliminated >98% of the promoter activity. Binding of nuclear proteins from HSC-2 cells to this 29-bp fragment was observed by gel shift analysis. The ability of mutations within the 29-bp fragment to affect enhancer activity correlated with the ability of these mutant oligonucleotides to compete for the wild- type sequence in gel shift assays.

Original languageEnglish
Pages (from-to)189-195
Number of pages7
JournalArchives of Biochemistry and Biophysics
Issue number1
StatePublished - Oct 1 1998

Bibliographical note

Funding Information:
This work is supported in part by a grant from the NIH: NIDA DA 02243.


  • Cell-specific expression
  • Enhancer activity
  • Gene regulation

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology


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