Cooperation of multiple signaling pathways in CD40-regulated gene expression in B lymphocytes

Abstract

CD40͞CD40L interaction is essential for multiple biological events in T dependent humoral immune responses, including B cell survival and proliferation, germinal center and memory B cell formation, and antibody isotype switching and affinity maturation. By using high-density microarrays, we examined gene expression in primary mouse B lymphocytes after multiple time points of CD40L stimulation. In addition to genes involved in cell survival and growth, which are also induced by other mitogens such as lipopolysaccharide, CD40L specifically activated genes involved in germinal center formation and T cell costimulatory molecules that facilitate T dependent humoral immunity. Next, by examining the roles of individual CD40-activated signal transduction pathways, we dissected the overall CD40-mediated response into genes independently regulated by the individual pathways or collectively by all pathways. We also found that gene down-regulation is a significant part of the overall response and that the p38 pathway plays an important role in this process, whereas the NF-B pathway is important for the up-regulation of primary response genes. Our finding of overlapping independent control of gene expression modules by different pathways suggests, in principle, that distinct biological behaviors that depend on distinct gene expression subsets can be manipulated by targeting specific signaling pathways. C D40 is a member of the pleiotropic tumor necrosis factor receptor (TNFR) superfamily, which also includes such functionally diverse molecules as CD27, CD30, OX40, RANK, LT-␤R, the p75 low-affinity nerve growth factor receptor, Fas, and members of the death receptor family as well as TNFR1 and TNFR2 (1). CD40 stimulation activates B cells and promotes various aspects of a functional humoral immune response, including enhancement of survival and proliferation (2). Like many TNFR family members, CD40 activates the JNK͞SAPK and NF-B pathways (3, 4). Both of these pathways involve serine͞threonine kinases that regulate gene expression through activation of AP1 and Rel transcription factors, respectively. Another stress-responsive pathway that has also been reported to be activated by CD40 (5) is the p38 kinase pathway, which leads to the phosphorylation and activation of transcription factors such as ATF2 (6). Previous work has also linked CD40 to the activation of the extracellular signal-regulated kinase (ERK)͞mitogen-activated protein kinase (MAPK) pathway (7).In addition to serine͞threonine kinases, a link between CD40 signaling and tyrosine kinase activation has been suggested (8).One pathway shown to be activated by CD40 and to play an important role in CD40 signaling is that initiated by PI-3 kinase (PI-3K) (8-10). PI-3K phosphorylates membrane phospholipids (11), which can recruit and activate pleckstrin homology domaincontaining molecules such as the kinase protein kinase B. This kinase may then regulate the activity of forkhead-related transcription factors (12, 13).In each of the above pathways, one of the major final ...

Potential role of CARMA1 in CD40‐induced splenic B cell proliferation and marginal zone B cell maturation

“…Pharmacological inhibition of any of these pathways dramatically reduces the proliferation response induced by CD40 stimulation [37]. However, our data suggest that activation of these MAP kinase signaling pathways is not significantly affected in CARMA1 KO B cells and not likely a reason for the defective proliferation.…”

“…Proliferation of B cells upon CD40 stimulation requires NF-jB activation and NF-jBdependent induction genes such as cyclin-D2 [37,38]. However, our data indicated that activation of NF-jB is not defective in CD40-stimulated CARMA1 KO B cells (Fig.…”

“…CARMA1 KO B cells not only fail to activate NF-jB following BCR ligation but also fail to undergo activation and proliferation [20,23,24]. Moreover, biological response following CD40 stimulation in B cells requires both canonical (NF-jB1) B1) and non-canonical (NF-jB2) pathways [37,38]. Therefore, we asked whether activation of NF-jB is compromised in CARMA1 KO B cells upon CD40 stimulation.…”

“…Inhibition of any of these pathways has been shown to dramatically reduce CD40-induced proliferation [37]. Moreover, p42/p44 (ERK1/2) phosphorylation upon CD40 stimulation stabilizes c-Myc protein and favors cell-cycle progression [43].…”

Analysis of the Major Patterns of B Cell Gene Expression Changes in Response to Short-Term Stimulation with 33 Single Ligands

“…Whether Ltb expression is up-regulated in later time points beyond 4 h is unknown. CD40L was previously shown to induce strong and sustained up-regulation of Ltb in B cells following 24 h of stimulation (61). It is possible that costimulation with CD40L is required to up-regulate Ltb in B cells during T-dependent immune response.…”