Aminopeptidase-A. II. Genomic cloning and characterization of the rat promoter.
Aminopeptidase-A (APA) has a widespread tissue distribution consistent with a role in the metabolism of circulating or locally produced ANG II or CCK-8. APA is also highly expressed in pre-B lymphocytes, but its role in lymphoid cell development is unknown. To begin to understand the basis for cell-specific regulation of APA expression, we sought to clone and characterize the rat gene promoter. Screening of a rat genomic library with a partial rat APA cDNA resulted in isolation of a 12-kb clone found to contain the first exon and >3 kb of 5'-flanking sequence. Primer extension of rat kidney mRNA indicated that the major transcription start site was 312 bp upstream of the translation start codon and 22 bp downstream from a TATA box. Constructs containing portions of the 5'-flanking region placed upstream of a chloramphenicol acetyltransferase reporter gene indicated that expression was cell specific and that high activity could be obtained with constructs containing as little as 110 bp of 5'-flanking region sequence. We further identified an upstream regulatory element between -1063 and -348 that suppressed transcription in a cell-specific manner. This element (termed upstream suppressor of APA, or USA) also suppressed transcription of a heterologous promoter. These results indicate that the organization and regulation of the rat APA is not consistent with it being a housekeeping gene and further suggest that rat APA gene transcription might be regulated through the presence of a novel strong upstream suppressor element.
Jiang
Q
Q
Department of Pharmacology, Mount Sinai School of Medicine of the City University of New York, New York 10029, USA.
Troyanovskaya
M
M
Jayaraman
G
G
Healy
D P
DP
D000626:Aminopeptidases / Q000235:genetics*; D000818:Animals; D001483:Base Sequence / Q000235:genetics; D003001:Cloning, Molecular; D015870:Gene Expression; D017930:Genes, Reporter / Q000235:genetics; D016678:Genome; D043384:Glutamyl Aminopeptidase; D018374:LLC-PK1 Cells; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000235:genetics*; D051381:Rats; D013552:Swine; D014158:Transcription, Genetic
D000626:Aminopeptidases; D043384:Glutamyl Aminopeptidase