Interleukin-2 (IL-2) withdrawal is a physiologic process inducing cell death in activated T lymphocytes. Glucocorticoidinduced leucine zipper (GILZ) has recently been identified as a protein modulating T-cell receptor activation by repressing various signaling pathways. We report here that IL-2 deprivation leads to expression of GILZ in T lymphocytes. We then characterized the human gilz promoter and showed that FoxO3 (Forkhead box class O3) binding to the Forkhead responsive elements identified in the promoter is necessary for induction of gilz expression upon IL-2 withdrawal. To assess the functional consequences of this induction, we used 2 strategies, GILZ overexpression and GILZ silencing in murine IL-2-dependent CTLL-2 cells. GILZ overexpression protects CTLL-2 cells from IL-2 withdrawal-induced apoptosis, whereas cell death is accelerated in cells unable to express GILZ. Concomitantly, the expression of Bim is inhibited in GILZ-overexpressing cells and enhanced when GILZ expression is impaired. Furthermore, GILZ inhibits FoxO3 transcriptional activity that leads to inhibition of Bim expression but also to down-regulation of GILZ itself. Therefore, GILZ is a transiently expressed protein induced upon IL-2 withdrawal that protects T cells from the onset of apoptosis. ( IntroductionA complex network of cytokines controls the development, maturation, homeostasis, and responses of the immune system. Immune responses typically involve clonal expansion of activated T cells, which must be eliminated at the end of the response to preserve homeostasis. 1 Interleukin-2 (IL-2) is secreted by activated T cells and plays a role in their survival and proliferation. IL-2 is also involved in the regulation of homeostasis of the lymphoid tissue, as IL-2 induces apoptosis of activated T cells in an "active" way involving regulation of Fas and Fas ligand (FasL) expression and in a "passive" way due to deprivation of this survival factor. 2 Binding of IL-2 to its receptor triggers a signaling cascade that induces, among others, the phosphoinositide 3-kinase (PI3K) pathway involved in T-cell proliferation and survival. 3,4 One of the downstream effectors of PI3K signaling is protein kinase B (PKB)/Akt. PKB is a serine/threonine kinase promoting cell survival by targeting proapoptotic proteins. 5 PKB exerts one of its inhibitory effects by phosphorylating members of the Forkhead family of transcription factors (Forkhead box class O1 [FoxO1], FoxO3, and FoxO4), which contain 3 RXRXXS/T consensus phosphorylation sites for PKB. 6 Interruption of the PI3K/PKB pathway following IL-2 deprivation results in dephosphorylation of FoxO proteins leading to their translocation to the nucleus and transcription of target genes.Brunet et al 7 were the first to ascribe a role to FoxO proteins in programmed cell death, reporting that overexpression of a constitutive active form of FoxO3 induces apoptosis of Jurkat T cells in a Fas-dependent manner. Fas-independent mechanisms of apoptosis induced by FoxO proteins have also been observed in othe...

T cell–dependent immune responses develop soon after birth, whereas it takes 2 yr for humans to develop T cell–independent responses. We used this dissociation to analyze the repertoire diversification of IgM+IgD+CD27+ B cells (also known as “IgM memory” B cells), comparing these cells with switched B cells in children <2 yr of age, with the aim of determining whether these two subsets are developmentally related. We show that the repertoire of IgM+IgD+CD27+ B cells in the spleen and blood displays no sign of antigen-driven activation and expansion on H-CDR3 spectratyping, despite the many antigenic challenges provided by childhood vaccinations. This repertoire differed markedly from those of switched B cells and splenic germinal center B cells, even at the early stage of differentiation associated with μ heavy chain expression. These data provide evidence for the developmental diversification of IgM+IgD+CD27+ B cells, at least in very young children, outside of T cell–dependent and –independent immune responses.

Klhl6 belongs to the KLHL gene family, which is composed of an N-terminal BTB-POZ domain and four to six Kelch motifs in tandem. Several of these proteins function as adaptors of the Cullin3 E3 ubiquitin ligase complex. In this article, we report that Klhl6 deficiency induces, as previously described, a 2-fold reduction in mature B cells. However, we find that this deficit is centered on the inability of transitional type 1 B cells to survive and to progress toward the transitional type 2 B cell stage, whereas cells that have passed this step generate normal germinal centers (GCs) upon a T-dependent immune challenge. Klhl6-deficient type 1 B cells showed a 2-fold overexpression of genes linked with cell proliferation, including most targets of the anaphase-promoting complex/cyclosome complex, a set of genes whose expression is precisely downmodulated upon culture of splenic transitional B cells in the presence of BAFF. These results thus suggest a delay in the differentiation process of Klhl6-deficient B cells between the immature and transitional stage. We further show, in the BL2 Burkitt's lymphoma cell line, that KLHL6 interacts with Cullin3, but also that it binds to HBXIP/Lamtor5, a protein involved in cell-cycle regulation and cytokinesis. Finally, we report that KLHL6, which is recurrently mutated in B cell lymphomas, is an off-target of the normal somatic hypermutation process taking place in GC B cells in both mice and humans, thus leaving open whether, despite the lack of impact of Klhl6 deficiency on GC B cell expansion, mutants could contribute to the oncogenic process.