SIGNAL TRANSDUCTION IN T LYMPHOCYTE ACTIVATION
Principal Investigator: Amnon Altman
Affiliation: La Jolla Institute for Allergy and Immunology
Abstract: The importance of PKC? in TCR signals controlling T cell activation and differentiation is well established. The selective requirement of PKC? in allergic and autoimmune diseases, but not in antiviral responses, highlights it as a promising drug target. Realization of this potential requires a clear understanding of the mechanism(s) that regulate the unique, non-redundant functions and highly selective immunological synapse (IS) localization of PKC? in T cells, which remain unknown. Our work during the last funding period revealed that within the large PKC family, PKC? represents a unique paradigm with respect to the mechanisms that regulate these events and, ultimately, lead to the activation of critical transcription factors, which determine T cell fate. We propose to apply a multi-disciplinary approach in order to address the following unresolved questions: Aim 1) Positive regulation of PKC? activation and localization. Using biochemical, genetic, imaging and biophysical approaches, we will address the hypothesis that the localization and activation of PKC? in T cells are positively regulated by specialized mechanisms and plasma membrane interactions owing to the unique properties of its regulatory C1 and V3 domains. We will conduct a detailed structure- function analysis of the C1A, C1B and V3 domains of PKC?, their phosphorylation, and their interaction with binding partners. Aim 2) Autoinhibition by the PKC? C2 domain. Our studies indicate that the PKC? N- terminal C2 domain negatively regulates its function, and that autoinhibition is relieved by TCR-induced phosphorylation of Tyr-90, and, furthermore, that C2 itself is a pTyr- and PIP2-binding domain. We will investigate the mechanisms through which autoinhibition is effected, and characterize C2-binding ligands and their functional significance. We will pursue Aims 1 and 2 as a collaborative consortium with Dr. Wonhwa Cho (U. Illinois), who will provide his expertise in biophysical and two-photon imaging approaches to further elucidate the specialized mechanisms that regulate PKC?. Aim 3) Regulation of Ca2+/NFAT signaling by PKC?. PKC?-/- T cells display a Ca2+/NFAT signaling defect, but the mechanism linking PKC? to Ca2+ signaling is unknown. Based on our recent findings, and given the important role of Ca2+ signaling in determining the balance between productive T cell activation and anergy/apoptosis, we will characterize this pathway. Specifically, we will address the hypothesis that PKC? mediates a positive feedback loop that promotes PLC31 activation and, hence, Ca2+/NFAT signaling, via its functional and/or physical association with Tec family tyrosine kinase(s). Our studies will elucidate novel mechanisms that control the unique membrane localization and function of PKC? in T cells. The new information will likely provide a rational basis for using PKC? as a drug target in autoimmune and allergic diseases, and in T cell malignancies. PUBLIC HEALTH RELEVANCE: Protein kinase C-8 (PKC?), an enzyme expressed in T lymphocytes of the immune system, is important for the activation and survival of these cells and their ability to mount many beneficial as well as pathological immune responses, e.g., protection against certain infections, and autoimmune and allergic diseases. However, the mechanisms, which control the unique functions and intracellular localization of PKC? in T lymphocytes (which are not shared by other related members of the PKC family), are largely unknown. Here we propose to use a complementary set of biochemical, genetic, imaging and biophysical approaches in order to elucidate these mechanisms and the regulatory interactions of PKC? with other proteins and membrane lipids. Understanding of these pathways is likely to establish a more rational basis for the use of PKC? as a drug target in autoimmune and allergic diseases, and in T cell leukemias/lymphomas
Funding Period: -------------------- - --------------------
more information: NIH RePORT
- Impaired activation and localization of LAT in anergic T cells as a consequence of a selective palmitoylation defectMatthias Hundt
Division of Cell Biology, La Jolla Institute for Allergy and Immunology, San Diego, California 92121, USA
Immunity 24:513-22. 2006....
- TNFR-associated factor 6 regulates TCR signaling via interaction with and modification of LAT adapterJi Ji Xie
State Key Laboratory of Biocontrol, Key Laboratory of Gene Engineering of Ministry of Education, School of Life Sciences, Sun Yat Sen University, Guangzhou 510006, China
J Immunol 190:4027-36. 2013....
- The yin and yang of protein kinase C-theta (PKCθ): a novel drug target for selective immunosuppressionElizabeth Yan Zhang
Division of Cell Biology, La Jolla Institute for Allergy and Immunology, La Jolla, CA, USA
Adv Pharmacol 66:267-312. 2013..Early progress in developing such drugs is being made, but additional studies on the role of PKCθ in the human immune system are urgently needed...
- In and out of the bull's eye: protein kinase Cs in the immunological synapseKok Fai Kong
Division of Cell Biology, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA
Trends Immunol 34:234-42. 2013..Further work is needed to identify mechanisms underlying PKC recruitment or exclusion at the IS, potential redundancy among IS-localized PKCs, and the relevance of PKC localization for IS dynamics and lymphocyte activation...
- Protein kinase Cθ C2 domain is a phosphotyrosine binding module that plays a key role in its activationRobert V Stahelin
Department of Chemistry, University of Illinois, Chicago, IL 60607, USA
J Biol Chem 287:30518-28. 2012..Collectively, these studies establish the C2 domain of PKCθ as a Tyr(P)-binding domain and suggest that the domain may play a major role in PKCθ activation via its Tyr(P) binding...
- Protein kinase C-η controls CTLA-4-mediated regulatory T cell functionKok Fai Kong
1 Division of Cell Biology, La Jolla Institute for Allergy and Immunology, La Jolla, California, USA 2
Nat Immunol 15:465-72. 2014..These results reveal a CTLA-4-PKC-η signaling axis required for contact-dependent suppression and implicate this pathway as a potential cancer immunotherapy target. ..
- PKCdelta acts upstream of SPAK in the activation of NKCC1 by hyperosmotic stress in human airway epithelial cellsLaura Smith
Department of Pediatrics, Case Western Reserve University, Cleveland, Ohio 44106, USA
J Biol Chem 283:22147-56. 2008..Inhibition of PKCdelta activity with rottlerin blocked the increase in SPAK kinase activity. The results indicate that PKCdelta acts upstream of SPAK to increase activity of NKCC1 during hyperosmotic stress...
- Mechanism of diacylglycerol-induced membrane targeting and activation of protein kinase CthetaHeather R Melowic
Department of Chemistry, University of Illinois, Chicago, Illinois 60607, USA
J Biol Chem 282:21467-76. 2007..Collectively, these results show that PKC has a unique membrane binding and activation mechanism that may account for its subcellular targeting properties...
- Protein kinase C theta (PKCtheta): a key player in T cell life and deathKeitaro Hayashi
Division of Cell Biology, La Jolla Institute for Allergy and Immunology, 9420 Athena Circle, La Jolla, CA 92037, USA
Pharmacol Res 55:537-44. 2007..Therefore, PKCtheta is a critical enzyme that regulates T cell function at multiple stages, and it represents an attractive drug target for allergic and autoimmune diseases...
- Filamin A is required for T cell activation mediated by protein kinase C-thetaKeitaro Hayashi
Division of Cell Biology, La Jolla Institute for Allergy and Immunology, San Diego, CA 92121, USA
J Immunol 177:1721-8. 2006....
- Antigen-independent signalosome of CARMA1, PKCθ, and TNF receptor-associated factor 2 (TRAF2) determines NF-κB signaling in T cellsTakanori So
Divisions of Molecular Immunology and Cell Biology, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA
Proc Natl Acad Sci U S A 108:2903-8. 2011..Thus, by recruiting TCR-related intracellular molecules into the TRAF2 complex, OX40 provides the T cell with a high level of NF-κB activity needed for longevity...
- A motif in the V3 domain of the kinase PKC-θ determines its localization in the immunological synapse and functions in T cells via association with CD28Kok Fai Kong
Division of Cell Biology, La Jolla Institute for Allergy and Immunology, La Jolla, California, USA
Nat Immunol 12:1105-12. 2011....