Predominant contribution of DGKζ over DGKα in the control of PKC/PDK-1-regulated functions in T cells.
Diacylglycerol kinase (DGK)-mediated consumption of the diacylglycerol (DAG) generated in response to antigen recognition is an important mechanism to limit T-cell function. Targeting DGK activity presents new opportunities for therapeutic manipulation of the immune response, but assessment of individual DGK functions is complex. T cells express two DGK isoforms, DGKα and DGKζ, and there are no isoform-specific inhibitors. Here we used short interfering RNA-mediated gene silencing in human T cells and DGKα- and DGKζ-deficient mice to define DGK isoform-specific regulation of key signaling pathways during T-cell activation. Our results identify DGKζ as the predominant brake on basal/tonic conditions as well as on downstream T-cell receptor/co-stimulatory signals. DGKζ silencing triggers basal RasGTP activation and facilitates enhanced membrane stability of protein kinase C alpha as well as increased activity of AGC kinases. Downstream of T-cell receptor/co-stimulation, DGKζ silencing results in enhanced and maintained recruitment of PKC theta to the membrane, as well as phosphoinositide-dependent protein kinase-1 activation and scaffolding functions. Our studies identify a previously unrecognized DGKζ contribution as a negative regulator of the crosstalk between phospholipase C-gamma- and phosphoinositide 3-kinase-regulated pathways. This DGKζ input helps to explain previous observations in DGK-deficient mice and suggests that the development of isoform-specific DGK inhibitors is of great interest for the manipulation of distinct aspects of T-cell responses.
Department of Immunology and Oncology, Centro Nacional de Biotecnología (CNB-CSIC), Madrid, Spain.
Antonia
A
Ávila-Flores
Department of Immunology and Oncology, Centro Nacional de Biotecnología (CNB-CSIC), Madrid, Spain.
Javier
J
Arranz-Nicolás
Department of Immunology and Oncology, Centro Nacional de Biotecnología (CNB-CSIC), Madrid, Spain.
Elena
E
Andrada
Department of Immunology and Oncology, Centro Nacional de Biotecnología (CNB-CSIC), Madrid, Spain.
Denise
D
Soutar
Department of Immunology and Oncology, Centro Nacional de Biotecnología (CNB-CSIC), Madrid, Spain.
Isabel
I
Mérida
D000818:Animals; D018106:CD28 Antigens / Q000819:agonists / Q000378:metabolism; D002460:Cell Line; D002634:Chemotaxis, Leukocyte / Q000235:genetics / Q000276:immunology; D019852:Diacylglycerol Kinase / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation; D055786:Gene Knockout Techniques; D020868:Gene Silencing; D006801:Humans; D007376:Interleukin-2 / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D011493:Protein Kinase C / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D000081382:Pyruvate Dehydrogenase Acetyl-Transferring Kinase; D011948:Receptors, Antigen, T-Cell / Q000819:agonists / Q000378:metabolism; D015375:Receptors, Interleukin-2 / Q000378:metabolism; D015398:Signal Transduction; D013601:T-Lymphocytes / Q000276:immunology* / Q000378:metabolism*; D018808:Transcription Factor AP-1 / Q000378:metabolism; D014158:Transcription, Genetic; D018631:ras Proteins / Q000378:metabolism
D018106:CD28 Antigens; D007376:Interleukin-2; D000081382:Pyruvate Dehydrogenase Acetyl-Transferring Kinase; D011948:Receptors, Antigen, T-Cell; D015375:Receptors, Interleukin-2; D018808:Transcription Factor AP-1; D019852:Diacylglycerol Kinase; C512480:diacylglycerol kinase zeta, mouse; D017346:Protein Serine-Threonine Kinases; D011493:Protein Kinase C; D018631:ras Proteins