Involvement of MAPKs, NF-kappaB and p300 co-activator in IL-1beta-induced cytosolic phospholipase A2 expression in canine tracheal smooth muscle cells.
Cytosolic phospholipase A2 (cPLA2) plays a pivotal role in mediating agonist-induced arachidonic acid release for prostaglandin (PG) synthesis during stimulation with interleukin-1beta (IL-1beta). However, the mechanisms underlying IL-1beta-induced cPLA2 expression and PGE2 synthesis by canine tracheal smooth muscle cells (CTSMCs) have not been defined. IL-1beta induced cPLA2 protein and mRNA expression, PGE2 production, and phosphorylation of p42/p44 MAPK, p38 MAPK (ATF2), and JNK (c-Jun) in a time- and concentration-dependent manner, determined by Western blotting, RT-PCR, and ELISA, which was attenuated by the inhibitors of MEK1/2 (U0126), p38 MAPK (SB202190), and JNK (SP600125), or transfection with dominant negative mutants of MEK1/2, p38, and JNK, respectively. Furthermore, IL-1beta-induced cPLA2 expression and PGE2 synthesis was inhibited by a selective NF-kappaB inhibitor (helenalin) or transfection with dominant negative mutants of NF-kappaB inducing kinase (NIK), IkappaB kinase (IKK)-alpha, and IKK-beta. Consistently, IL-1beta stimulated both IkappaB-alpha degradation and NF-kappaB translocation into nucleus in these cells. NF-kappaB translocation was blocked by helenalin, but not by U0126, SB202190, and SP600125. MAPKs together with NF-kappaB-activated p300 recruited to cPLA2 promoter thus facilitating the binding of NF-kappaB to cPLA2 promoter region and expression of cPLA2 mRNA. IL-1beta-induced cPLA2 expression and PGE2 production was inhibited by actinomycin D and cycloheximide, indicating the involvement of transcriptional and translational events in these responses. These results suggest that in CTSMCs, IL-1beta-induced cPLA2 expression and PGE2 synthesis was independently mediated through activation of MAPKs and NF-kappaB pathways and was connected to p300 recruitment and activation.
Luo
Shue-Fen
SF
Department of Internal Medicine, Chang Gung University, Kwei-San, Tao-Yuan, Taiwan.
Lin
Chih-Chung
CC
Chen
Hsin-Chieh
HC
Lin
Wei-Ning
WN
Lee
I-Ta
IT
Lee
Chiang-Wen
CW
Hsiao
Li-Der
LD
Yang
Chuen-Mao
CM
D000818:Animals; D003600:Cytosol / Q000201:enzymology*; D015232:Dinoprostone / Q000096:biosynthesis; D004285:Dogs; D048049:Extracellular Signal-Regulated MAP Kinases / Q000502:physiology; D005260:Female; D053583:Interleukin-1beta / Q000494:pharmacology*; D048031:JNK Mitogen-Activated Protein Kinases; D008297:Male; D020928:Mitogen-Activated Protein Kinases / Q000502:physiology*; D032389:Myocytes, Smooth Muscle / Q000201:enzymology*; D016328:NF-kappa B / Q000502:physiology*; D054467:Phospholipases A2 / Q000235:genetics*; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000032:analysis; D014132:Trachea / Q000201:enzymology*; D050880:p300-CBP Transcription Factors / Q000502:physiology*; D048051:p38 Mitogen-Activated Protein Kinases / Q000502:physiology
D053583:Interleukin-1beta; D016328:NF-kappa B; D012333:RNA, Messenger; D050880:p300-CBP Transcription Factors; D048049:Extracellular Signal-Regulated MAP Kinases; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D054467:Phospholipases A2; D015232:Dinoprostone