Enhanced T cell-independent antibody responses in c-Jun N-terminal kinase 2 (JNK2)-deficient B cells following stimulation with CpG-1826 and anti-IgM
Introduction
B cells undergo differentiation into antibody forming cells and/or class switching under the influence of T cells, their derived cytokines, and/or co-stimulators for B cells [1], [2], [3]. In addition to CD4+ T cell help, Toll-like receptor (TLR) ligand (TLR-L) also plays a crucial role in antibody production in B cells [4], [5]. For example, CpG-oligonucleotides (CpG ODN) stimulates B cells to cause T helper cell 1 (Th1)-like class switching through activation of TLR9 [6], [7], [8]. Combined stimulation with membrane immunoglobulin (mIg) engagement plus CpG ODN causes B cells to induce efficient class switching and/or B cell proliferation [9], [10]. A ligand for TLR4 lipopolysaccharide (LPS) induces class switching to IgG1 and IgE in combination with IL-4 and CD40-L [1], [11]. Class switch recombination requires activation-induced cytidine deaminase (AID) [12]. Accessibility of Ig S region to switch recombination machinery is promoted by germ line transcription of the Ig S region [13].
Signals through mIg and/or TLRs are mediated by several signal transduction pathway including NF-κB and mitogen-activated protein kinases (MAPKs) [14], [15], [16], [17]. MAPKs are composed of extracellular signal-regulated kinases (ERKs), p38MAPKs and c-Jun NH2-terminal kinases (JNKs) [18], [19], [20], [21]. JNKs comprise three JNK isoforms, namely, JNK1, JNK2, and JNK3. Both JNK1 and JNK2 are expressed relatively ubiquitously, whereas JNK3 is selectively expressed in brain and testis [19]. Gene-targeting experiments in mice have demonstrated that JNK1 and JNK2 have distinct and non-overlapping function in T cells in vitro [18], [22], [23], [24], [25]. T cells from JNK1-deficient (JNK1−/−) mice produce an increased amount of Th2 cytokine [22], whereas JNK2 is necessary for Th1 development [23]. However, the role for JNK2 in B cell activation and differentiation remains unresolved.
In the present study, we addressed whether JNK2 modulates antibody production in response to CpG ODN in the presence or absence of anti-IgM. Compared to the WT B cells, the JNK2−/− B cells showed enhanced IgG1 and IgG2a antibody production with comparable or slightly diminished level of B cell proliferation when treated with CpG ODN or CpG ODN plus anti-IgM. These findings are valuable for understanding the roles of JNKs in B cell activation and differentiation.
Section snippets
Mice
Genetically JNK2-deficient and WT mice were obtained from Jackson Laboratory (Bar Harber, ME, USA). Mice were backcrossed to C57BL/6J for six generations. Breeding and maintenance of mice were carried out under specific pathogen-free conditions at the Tokyo Medical University, and they were handled according to the guideline of the Ethical Committee of Animal Experiments of the Tokyo Medical University.
Isolation and activation of spleen B cells
For isolation of B lymphocytes, single-cell suspensions prepared from spleen were treated
JNK2 is phosphorylated in response to CpG ODN and anti-IgM
Because pathogen or pathogen-derived components contain CpG ODN as well as antigenic determinants [16], [30], we examined whether CpG ODN, a ligand for TLR-9, in the presence or absence of anti-IgM would activate JNK2, which plays a crucial role in the production of Th1 cytokine [23]. Incubation of B cells with either CpG ODN or anti-IgM for 30 min induced phosphorylation of JNK2 protein; the level of phosphorylation was much higher when the cells were stimulated for 30 min with both CpG ODN and
Discussion
Isotype switching in B cells is controlled by CD40-L, cytokines, and/or TLR signals. IL-4 directs IgG1 and IgE antibody production [1], [2], whereas IFN-γ and/or IL-27 favor(s) IgG2a antibody isotype switching [3], [32]. The IFN-γ production is induced by Th1 cells [33], development of which is regulated by JNK2 [23]. In the present study, using JNK2−/− B cells we examined whether JNK2 modulates antibody responses induced by CpG ODN and/or anti-IgM.
JNK2−/− B cells displayed an enhanced
Acknowledgement
This work was supported by a grant from the Intractable Immune System Disease Research Center of Tokyo Medical University, which is supported by the Ministry of Education, Culture, Sports, Science, and Technology of Japan.
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