Role of PKCθ in T-cell signalling
1
Vol.21 No.5 203 MAY 2000 UPDATE IMMUNOLOGY TODAY Current literature T-cell signalling and subsequent cytokine gene expression is dependent on the activa- tion of members of the serine/threonine- specific protein kinase C (PKC) family. Conventional PKCs are activated by both diacylglycerol (DAG) and Ca 21 . PKCu, along with the d, e and h enzymes, is a member of the novel subclass of PKCs that are activated by DAG only. PKCu is expressed only in T cells and is rapidly recruited to the contact point between the T cell and the APC. In this study, Sun et al. generated a PKCu-knockout mouse and ex- amined the role of PKCu in TCR signalling. FACS analysis studies indicated that cell numbers in the thymus, spleen and lymph nodes of PKCu 2/2 mice were normal, and the processes of negative and positive selection in the thymus did not require PKCu. Peripheral T-cell signalling was found to be dependent on PKCu. Purified T cells were stimulated to proliferate using plate-bound anti-CD3 antibody, with or without anti-CD28 stimulation. In comparison with wild-type mice, proliferation of T cells from PKC 2/2 mice was significantly reduced. This was also true of T cells stimulated to proliferate in mixed lymphocyte reactions. FACS analy- sis studies showed that the reduced prolif- erative responses of PKCu T cells was due to decreased expression of both interleukin 2 (IL-2) and the high-affinity IL-2 receptor. The induction of cytokine gene expression following T-cell signalling is dependent on several transcription factors, including NF-AT, AP-1 and NF-kB. The absence of PKCu had no effect on the translocation of NF-AT1 and 2 to the nucleus after activation of wild-type and PKCu 2/2 T cells. Mature PKCu T cells showed a significant defect in the activation of NF-kB activation following stimulation with anti-CD3, or anti-CD3 plus anti-CD28. By contrast, NF-kB activation was unaffected in activated PKCu 2/2 thymocytes, suggesting that an alternative PKC enzyme is impor- tant for T-cell signalling in thymocytes. This study indicates that PKCu is impor- tant in linking T-cell signalling and NF-kB activation in a unique signalling pathway in mature T cells. Elaine Bell Very early B-lineage commitment further dissected Tudor, K.R.S. et al. (2000) Functional assess- ment of precursors from murine bone mar- row suggests a sequence of early B lineage differentiation events. Immunity 12, 335–345 Defining the earliest steps in B-cell differen- tiation has become something of a Holy Grail, and has been greatly assisted by the development of reagents that are able to probe the surface and intracellular molecules that define B-lineage activities. Here, Tudor and colleagues have distinguished two sub- sets of mouse common lymphoid progenitor (CLP) cells that, although phenotypically part of the CLP pool, appear to be defining a very early B-lineage predisposition. The earliest indication of lymphoid com- mitment to either T- or B-cell lineage is the activity of terminal deoxynucleotide trans- ferase (TdT), an enzyme essential for antigen receptor diversification. At what stage can these cells be clearly suggested to be compo- nents of either the T- or B-lineage pathway? Apparently uncommitted, lineage marker- negative (Lin 2 ) TdT 1 cells from murine mar- row display c-kit at low density, as well as interleukin 7 receptor (IL-7R) and Flk-2/Flt-3 receptors. However, single cells with these characteristics generated CD45RA 1 CD19 2 as well as CD19 1 lymphocytes at high clonal frequency in culture. In contrast to other studies on the earliest B-lineage committed cells, an important result here indicates that these early pro-B cells have detectable CD24 expression but no CD11b. These Lin 2 TdT 1 c-kit lo IL-7Ra 1 Flk-2 1 Flt-3 1 CD24 lo/1 cells can be further subdivided on the basis of expression of two CLP markers, DX5 and Ly6C, both of which are absent from cells with the most significant B-lineage clonal precursor frequency. It has only recently been possible to study the differentiation potential of CLP cells iso- lated on the basis of early TdT expression. This report proposes that there are two Lin 2 TdT 1 c-kit lo IL-7Ra 1 Flk-2 1 Flt-3 1 CD24 lo/1 pro-B-cell populations, and that they may have an early and intermediate precur- sor—product relationship as they move from Ly6c 1 DX5 1 to Ly6c 2 DX5 2 status. Rob Brines Role of PKCu in T-cell signalling Sun, Z. et al. (2000) PKCu is required for TCR-induced NF-kB activation in mature but not immature T lymphocytes. Nature 404, 402–407 Autoantibodies are associated with a wide variety of autoimmune diseases. Although the structure and function of antigens tar- geted by autoantibodies have been well characterized, there is little evidence to support the role of such antibodies in the pathogenesis of autoimmune diseases. Rheumatic diseases are particularly asso- ciated with autoantibodies to nuclear anti- gens. In systemic lupus erythematosus (SLE) and autoimmune bowel diseases a high fre- quency of autoantibodies against poly (ADP-ribose) polymerase (PARP) has been detected. PARP is a nuclear enzyme that de- tects and binds DNA strand breaks. It con- tains an N-terminal DNA binding domain and two zinc finger motifs (F1 and F2) that are involved in detecting DNA breaks dur- ing DNA repair. IgG autoantibodies against the zinc finger domains of PARP are preva- lent in SLE and autoimmune bowel diseases. Previously, Decker et al. examined the possible pathogenic role of these antibodies in SLE. Autoantibodies to the F2 motif of PARP purified from patients with SLE were shown not to inhibit the catalytic activity of PARP. In this study, the role of PARP au- toantibodies in preventing PARP cleavage by caspase-3 was examined. This process is crucial in the execution phase of apoptosis, to ensure that PARP is inactivated and does not carry out futile DNA repair. Autoanti- bodies to PARP F2 efficiently inhibited cas- pase-3-mediated PARP cleavage in vitro and prolonged the survival of cells undergoing apoptosis. These findings indicate that au- toantibodies to PARP might contribute to the pathogenesis of SLE by prolonging the survival of cells important to the auto- immune process. Elaine Bell Pathogenic properties of PARP autoantibodies in autoimmune disease Decker, P. et al. (2000) Inhibition of caspase-3-mediated poly (ADP-ribose) polymerase (PARP) apoptotic cleavage by human PARP autoantibodies and effect on cells undergoing apoptosis. J. Biol. Chem. 275, 9043–9046
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Transcript of Role of PKCθ in T-cell signalling
V o l . 2 1 N o . 5 2 0 3
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U P D A T EI M M U N O L O G Y TO D AY
Current literature
T-cell signalling and subsequent cytokinegene expression is dependent on the activa-tion of members of the serine/threonine-specific protein kinase C (PKC) family. Conventional PKCs are activated by bothdiacylglycerol (DAG) and Ca21. PKCu,along with the d, e and h enzymes, is amember of the ÔnovelÕ subclass of PKCs thatare activated by DAG only. PKCu is expressed only in T cells and is rapidly recruited to the contact point between the T cell and the APC. In this study, Sun et al.generated a PKCu-knockout mouse and ex-amined the role of PKCu in TCR signalling.
FACS analysis studies indicated that cellnumbers in the thymus, spleen and lymphnodes of PKCu2/2 mice were normal, and theprocesses of negative and positive selectionin the thymus did not require PKCu.
Peripheral T-cell signalling was found tobe dependent on PKCu. Purified T cells werestimulated to proliferate using plate-boundanti-CD3 antibody, with or without anti-CD28stimulation. In comparison with wild-typemice, proliferation of T cells from PKC2/2
mice was significantly reduced. This was
also true of T cells stimulated to proliferatein mixed lymphocyte reactions. FACS analy-sis studies showed that the reduced prolif-erative responses of PKCu T cells was due todecreased expression of both interleukin 2(IL-2) and the high-affinity IL-2 receptor.
The induction of cytokine gene expressionfollowing T-cell signalling is dependent onseveral transcription factors, including NF-AT,AP-1 and NF-kB. The absence of PKCu hadno effect on the translocation of NF-AT1 and2 to the nucleus after activation of wild-typeand PKCu2/2 T cells. Mature PKCu T cellsshowed a significant defect in the activationof NF-kB activation following stimulationwith anti-CD3, or anti-CD3 plus anti-CD28.By contrast, NF-kB activation was unaffectedin activated PKCu2/2 thymocytes, suggestingthat an alternative PKC enzyme is impor-tant for T-cell signalling in thymocytes.
This study indicates that PKCu is impor-tant in linking T-cell signalling and NF-kBactivation in a unique signalling pathway inmature T cells.
Elaine Bell
Very early B-lineage commitmentfurther dissectedTudor, K.R.S. et al. (2000) Functional assess-ment of precursors from murine bone mar-row suggests a sequence of early B lineage differentiation events. Immunity 12, 335–345
Defining the earliest steps in B-cell differen-tiation has become something of a Holy Grail, and has been greatly assisted by thedevelopment of reagents that are able toprobe the surface and intracellular moleculesthat define B-lineage activities. Here, Tudorand colleagues have distinguished two sub-sets of mouse common lymphoid progenitor(CLP) cells that, although phenotypicallypart of the CLP pool, appear to be defining avery early B-lineage predisposition.
The earliest indication of lymphoid com-mitment to either T- or B-cell lineage is theactivity of terminal deoxynucleotide trans-ferase (TdT), an enzyme essential for antigenreceptor diversification. At what stage canthese cells be clearly suggested to be compo-nents of either the T- or B-lineage pathway?Apparently uncommitted, lineage marker-negative (Lin2) TdT1 cells from murine mar-row display c-kit at low density, as well asinterleukin 7 receptor (IL-7R) and Flk-2/Flt-3receptors. However, single cells with thesecharacteristics generated CD45RA1CD192
as well as CD191 lymphocytes at high clonalfrequency in culture. In contrast to otherstudies on the earliest B-lineage committedcells, an important result here indicates thatthese Ôearly pro-BÕ cells have detectableCD24 expression but no CD11b. TheseLin2TdT1c-kitloIL-7Ra1Flk-21Flt-31CD24lo/1
cells can be further subdivided on the basisof expression of two CLP markers, DX5 andLy6C, both of which are absent from cellswith the most significant B-lineage clonalprecursor frequency.
It has only recently been possible to studythe differentiation potential of CLP cells iso-lated on the basis of early TdT expression.This report proposes that there are twoLin2TdT1c-kitloIL-7Ra1Flk-21Flt-31CD24lo/1
pro-B-cell populations, and that they mayhave an ÔearlyÕ and ÔintermediateÕ precur-sorÐproduct relationship as they move fromLy6c1DX51 to Ly6c2DX52 status.
Rob Brines
Role of PKCu in T-cell signallingSun, Z. et al. (2000) PKCu is required for TCR-induced NF-kB activation in mature but not immature T lymphocytes. Nature 404, 402–407
Autoantibodies are associated with a widevariety of autoimmune diseases. Althoughthe structure and function of antigens tar-geted by autoantibodies have been wellcharacterized, there is little evidence to support the role of such antibodies in thepathogenesis of autoimmune diseases.
Rheumatic diseases are particularly asso-ciated with autoantibodies to nuclear anti-gens. In systemic lupus erythematosus (SLE)and autoimmune bowel diseases a high fre-quency of autoantibodies against poly(ADP-ribose) polymerase (PARP) has beendetected. PARP is a nuclear enzyme that de-tects and binds DNA strand breaks. It con-tains an N-terminal DNA binding domainand two zinc finger motifs (F1 and F2) thatare involved in detecting DNA breaks dur-ing DNA repair. IgG autoantibodies againstthe zinc finger domains of PARP are preva-lent in SLE and autoimmune bowel diseases.
Previously, Decker et al. examined thepossible pathogenic role of these antibodiesin SLE. Autoantibodies to the F2 motif ofPARP purified from patients with SLE wereshown not to inhibit the catalytic activity ofPARP. In this study, the role of PARP au-toantibodies in preventing PARP cleavageby caspase-3 was examined. This process iscrucial in the execution phase of apoptosis,to ensure that PARP is inactivated and doesnot carry out futile DNA repair. Autoanti-bodies to PARP F2 efficiently inhibited cas-pase-3-mediated PARP cleavage in vitro andprolonged the survival of cells undergoingapoptosis. These findings indicate that au-toantibodies to PARP might contribute tothe pathogenesis of SLE by prolonging thesurvival of cells important to the auto-immune process.
Elaine Bell
Pathogenic properties of PARP autoantibodies in autoimmune diseaseDecker, P. et al. (2000) Inhibition of caspase-3-mediated poly (ADP-ribose) polymerase (PARP)apoptotic cleavage by human PARP autoantibodies and effect on cells undergoing apoptosis. J. Biol. Chem. 275, 9043–9046
