title: Identification of a 27 bp 5'-flanking region element responsible for the low level constitutive expression of the human cytosolic phospholipase A2 gene.
abstract: The cytosolic phospholipase A2 (cPLA2) gene codes for an enzyme that liberates arachidonic acid from membrane phospholipids, and thus plays a pivotal role in the production of the prostaglandin and leukotriene mediators of inflammation, as well as in a variety of cell signalling pathways. After preliminary studies demonstrated the cPLA2 gene is expressed in a variety of human tissues and was localized to the q arm of chromosome 1 between markers F13B and D1S74, we cloned and characterized the 5'-flanking region of this gene in order to identify the elements controlling its low level constitutive expression. The 5'-flanking region has features typical of a housekeeping gene with no TATA box or CAAT box, although atypical in that it is not GC rich, has no SP1 sites, and has a long run of CA repeats. Analysis of fragments of the 5'-flanking region demonstrated that 541 bp 5' to exon 1 supported reporter gene activity at a level 30% of the SV40 promoter. Interestingly, similar activity was observed by deleting most of the 5'-flanking region down to a 27 bp region containing a sequence with homology to the initiator sequence in the terminal deoxynucleotidyl transferase gene and a polypyrimidine tract similar to the initiator element of the mouse ribosomal protein gene. Within this 27 bp region, a 10 bp fragment (-17 to -8 bp) within the polypyrimidine tract is critical for the baseline expression of the human cPLA2 gene. While the 5'-flanking region contains a putative composite AP-1 and glucocorticoid response element, this region does not respond to tumor necrosis factor-alpha (TNF) and/or glucocorticoids in a cell line (HEp-2) that exhibits upregulation of cPLA2 mRNA transcript levels by TNF. The observations that the expression of the cPLA2 gene is tightly controlled at a relatively low level is consistent with the evolving concept that modulation of expression of this critical enzyme is primarily at the post-translational level.
authors.affiliation: Division of Pulmonary and Critical Care Medicine, New York Hospital-Cornell Medical Center, New York 10021.
authors.forename: A
authors.initials: A
authors.lastname: Miyashita
authors.forename: R G
authors.initials: RG
authors.lastname: Crystal
authors.forename: J G
authors.initials: JG
authors.lastname: Hay
mesh_terms: D001483:Base Sequence; D002874:Chromosome Mapping; D002878:Chromosomes, Human, Pair 1; D003600:Cytosol / Q000201:enzymology*; D003907:Dexamethasone / Q000494:pharmacology; D015870:Gene Expression; D017930:Genes, Reporter; D006801:Humans; D008969:Molecular Sequence Data; D009928:Organ Specificity; D010741:Phospholipases A / Q000235:genetics*; D054467:Phospholipases A2; D012150:Polymorphism, Restriction Fragment Length; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000032:analysis; D011993:Recombinant Fusion Proteins; D012091:Repetitive Sequences, Nucleic Acid; D016385:TATA Box; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
chemical_list: D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D014409:Tumor Necrosis Factor-alpha; D003907:Dexamethasone; D010741:Phospholipases A; D054467:Phospholipases A2