Jun N-terminal kinase is essential for CD40-mediated IgE class switching in B cells.
BACKGROUND
CD40 ligation activates nuclear factor kappaB (NF-kappaB) and the mitogen-activated protein kinases p38 and C-Jun N-terminal kinase (JNK) and causes immunoglobulin class-switch recombination (CSR) in B cells. Both NF-kappaB and p38 are important for CD40-mediated CSR. The role of JNK activation in CD40-mediated isotype switching is unknown.


OBJECTIVE
We sought to determine the role of JNK activation in CD40-mediated isotype switching.


METHODS
Splenic B cells from BALB/c mice were stimulated with anti-CD40 mAb and IL-4 or with soluble CD40 ligand in the presence or absence of SP600125, an anthrapyrazolone inhibitor of JNK. The following events were examined: IgE production by means of ELISA; S(mu)-S(epsilon) deletional switch recombination by means of digestion circularization PCR; Cepsilon germline, mature epsilon, and activation-induced deaminase (AID) transcription by means of RT-PCR; and proliferation by tritiated thymidine incorporation and surface expression of CD23, CD54, and CD86 by means of FACS analysis.


RESULTS
SP600125 at 10 microM drastically inhibited JNK phosphorylation but had little effect on CD40-mediated p38 phosphorylation and expression of the NF-kappaB dependent genes c-Myc and bcl-xL. SP600125 inhibited IgE synthesis by approximately 88% but had no effect on B-cell proliferation and survival in response to anti-CD40 + IL-4 or on upregulation of CD23, CD54, and CD86 in response to CD40 ligation. Analysis of molecular events involved in IgE class switching revealed that SP600125 had no effect on the expression of C(epsilon) germline and AID transcripts. In contrast, SP600125 severely reduced S(mu)-S(epsilon) switch recombination and expression of mature epsilon transcripts.


CONCLUSIONS
These results demonstrate that JNK activation is essential for CD40-mediated CSR to IgE and suggest that JNK is important for AID activity in B cells.
Jabara
Haifa H
HH
Division of Immunology, Children's Hospital, KARP Building #10126, 1 Blackfan Circle, Boston, MA 02115, USA. Haifa.jubara@childrens.harvard.edu
Geha
Raif S
RS
D000818:Animals; D000873:Anthracenes / Q000494:pharmacology; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D001402:B-Lymphocytes / Q000187:drug effects / Q000276:immunology*; D019013:CD40 Antigens / Q000276:immunology*; D023201:CD40 Ligand / Q000276:immunology / Q000494:pharmacology; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D003564:Cytidine Deaminase; D043525:Cytosine Deaminase / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D017578:Immunoglobulin Class Switching / Q000187:drug effects / Q000276:immunology*; D007073:Immunoglobulin E / Q000187:drug effects / Q000276:immunology*; D015847:Interleukin-4 / Q000276:immunology / Q000494:pharmacology; D048031:JNK Mitogen-Activated Protein Kinases / Q000187:drug effects / Q000276:immunology* / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D010766:Phosphorylation / Q000187:drug effects; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000098830:AICDA (Activation-Induced Cytidine Deaminase)
D000873:Anthracenes; D000911:Antibodies, Monoclonal; D019013:CD40 Antigens; D023201:CD40 Ligand; C432165:pyrazolanthrone; D015847:Interleukin-4; D007073:Immunoglobulin E; D048031:JNK Mitogen-Activated Protein Kinases; D000098830:AICDA (Activation-Induced Cytidine Deaminase); D043525:Cytosine Deaminase; D003564:Cytidine Deaminase