Functional effects of overexpression of protein kinase C-alpha, -beta, -delta, -epsilon, and -eta in the mast cell line RBL-2H3.
The rat basophilic leukemic (RBL-2H3) cell line was stably transfected with the endogenously expressed Ca2+-dependent protein kinase C-alpha (PKC-alpha) and -betaI and the Ca2+-independent delta and epsilon isoforms to study their functional roles. In addition, the Ca2+-independent PKC-eta was expressed. All transfected PKC isoforms translocated to the membrane-containing fraction in response to aggregation of the IgE-sensitized high affinity receptor for IgE (Fc epsilonRI) with the Ag dinitrophenyl(25)-BSA. All PKC transfectants, except PKC-eta, showed increased proliferative responses, and aggregation of Fc epsilonRI further enhanced the rate of proliferation. The PKC transfectants also showed increased phosphoinositide hydrolysis in response to Ag aggregation of receptors. No marked differences in the Ca2+ responses of the transfectants to Ag or thapsigargin were observed. Overexpression of PKC-alpha or -epsilon specifically inhibited receptor-dependent cytosolic phospholipase A2 (cPLA2) activity, whereas this activity was enhanced in the PKC-betaI transfectant. Analysis of the secretory response revealed that overexpression of PKC-betaI and -eta significantly enhanced secretion. A broad spectrum of cytokine mRNAs was detected in all transfectants, and overexpression of PKC-betaI significantly enhanced the receptor-dependent production of IL-2 and IL-6 mRNA. These studies identify PKC-alpha and -epsilon as negative regulators of cPLA2 activity and demonstrate the importance of PKC-beta as a positive modulator of secretion, cPLA2 activity, and cytokine production in this mast cell line.
Chang
E Y
EY
Section on Chemical Immunology, National Institute of Arthritis and Musculoskeletal and Skin Diseases, National Institutes of Health, Bethesda, MD 20892, USA.
Szallasi
Z
Z
Acs
P
P
Raizada
V
V
Wolfe
P C
PC
Fewtrell
C
C
Blumberg
P M
PM
Rivera
J
J
D000818:Animals; D002118:Calcium / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D007527:Isoenzymes / Q000096:biosynthesis* / Q000235:genetics; D008407:Mast Cells / Q000201:enzymology*; D011493:Protein Kinase C / Q000096:biosynthesis* / Q000235:genetics; D064546:Protein Kinase C beta; D051571:Protein Kinase C-alpha; D051745:Protein Kinase C-delta; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D051381:Rats; D015398:Signal Transduction / Q000235:genetics*; D014407:Tumor Cells, Cultured
D007527:Isoenzymes; D012333:RNA, Messenger; C497260:Prkcd protein, rat; D011493:Protein Kinase C; D064546:Protein Kinase C beta; D051571:Protein Kinase C-alpha; D051745:Protein Kinase C-delta; D002118:Calcium