Transcriptional regulation of the rat type IIA phospholipase A2 gene by cAMP and interleukin-1beta in vascular smooth muscle cells: interplay of the CCAAT/enhancer binding protein (C/EBP), nuclear factor-kappaB and Ets transcription factors.
The abundant secretion of type IIA secreted phospholipase A(2) (sPLA(2)) is a major feature of the inflammatory process of atherosclerosis. sPLA(2) is crucial for the development of inflammation, as it catalyses the production of lipid mediators and induces the proliferation of smooth muscle cells. We have analysed the activation of sPLA(2) transcription by cAMP and interleukin-1beta (IL-1beta), and shown that the 500 bp region upstream of the transcription start site of the rat sPLA(2) gene is implicated in activation by synergistically acting cAMP and IL-1beta. We transiently transfected and stimulated rat smooth muscle cells in primary culture and measured the promoter activities of serial and site-directed deletion mutants of sPLA(2)-luciferase constructs. A distal region, between -488 and -157 bp, bearing a CAAT/enhancer binding protein (C/EBP)-responsive element (-242 to -223) was sufficient for cAMP/protein kinase A-mediated sPLA(2) promoter activation. We find evidence for the first time that activation of the sPLA(2) promoter by IL-1beta requires activation of an Ets-responsive element in the -184 to -180 region of the distal promoter via the Ras pathway and a nuclear factor-kappaB site at positions -141 to -131 of the proximal promoter. We also used electrophoretic mobility shift assays to identify five binding sites for the Sp1 factor; a specific inhibitor of Sp1, mithramycin A, showed that this factor is crucial for the basal activity of the sPLA(2) promoter.
UMR Physiologie et Physiopathologie, Université Pierre et Marie Curie, Case courrier 256, Bâtiment A, 5ème étage, 7 quai St Bernard, 75252 Paris cedex 05, France.
Valérie
V
Antonio
Arthur
A
Brouillet
Brigitte
B
Janvier
Claire
C
Monne
Gilbert
G
Bereziat
Marise
M
Andreani
Michel
M
Raymondjean
D000818:Animals; D001161:Arteriosclerosis / Q000235:genetics; D001483:Base Sequence; D001665:Binding Sites; D022762:CCAAT-Enhancer-Binding Proteins / Q000378:metabolism*; D016466:CHO Cells; D002478:Cells, Cultured; D006224:Cricetinae; D000242:Cyclic AMP / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D054501:Group II Phospholipases A2; D006801:Humans; D007375:Interleukin-1 / Q000378:metabolism* / Q000494:pharmacology; D008297:Male; D008969:Molecular Sequence Data; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D016328:NF-kappa B / Q000378:metabolism*; D010741:Phospholipases A / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D054467:Phospholipases A2; D011401:Promoter Regions, Genetic; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D050783:Proto-Oncogene Proteins c-ets; D051381:Rats; D017208:Rats, Wistar; D016329:Sp1 Transcription Factor / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic; D018631:ras Proteins / Q000235:genetics / Q000378:metabolism
D022762:CCAAT-Enhancer-Binding Proteins; D007375:Interleukin-1; D016328:NF-kappa B; D011518:Proto-Oncogene Proteins; D050783:Proto-Oncogene Proteins c-ets; D016329:Sp1 Transcription Factor; D014157:Transcription Factors; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D010741:Phospholipases A; D054501:Group II Phospholipases A2; D054467:Phospholipases A2; D018631:ras Proteins