Phospholipase A2s are families of enzymes that catalyze the hydrolysis of the sn-2 ester bond of phospholipids. Cytosolic Phospholipase A2 is a high molecular weight PLA2 which preferentially hydrolyzes phospholipids containing arachidonic acid in the sn-2 position. In some cells, cPLA2 activity and protein levels may be regulated by cytokines and growth factors. Therefore, it was of interest to further investigate the regulation of gene expression of this enzyme by characterizing the 5' promoter region of the cPLA2 gene. A probe consisting of the 5' portion of the known cDNA sequence of cPLA2 was used to screen a genomic library in Lambda Phage. A clone was isolated from the library which contained a 20 kB insert. After Eco RI digestion of the insert, a 5.7 kB fragment which bound the probe was subcloned into a Bluescript phage and sequenced. The transcription initiation site was identified by primer extension and by rapid amplification of 5' cDNA ends. DNA sequence analysis of the 595 base pairs 5' to the transcription initiation site revealed a 48 base pair purine-pyrimidine dinucleotide repeat, five interferon gamma response elements and one interferon gamma activated sequence. The promoter does not have a TATA box. The 595 basepair sequence has promoter activity in a luciferase reporter gene system. Current work involves the construction of reporter genes with approximately l, 2, and 3kB inserts in order to further study the properties of the 5' region of the cPLA2 gene. An understanding of the mechanisms of induction of gene expression of this enzyme may allow for novel approaches to the control of eicosanoid production in inflammatory diseases of the airways.

Agency
National Institute of Health (NIH)
Institute
Clinical Center (CLC)
Type
Intramural Research (Z01)
Project #
1Z01CL000149-02
Application #
5201104
Study Section
Project Start
Project End
Budget Start
Budget End
Support Year
2
Fiscal Year
1995
Total Cost
Indirect Cost
Name
Clinical Center
Department
Type
DUNS #
City
State
Country
United States
Zip Code