Transitional B Cell Fate Is Associated with Developmental Stage-Specific Regulation of Diacylglycerol and Calcium Signaling upon B Cell Receptor Engagement

Abstract

Functional peripheral mature follicular B (FoB) lymphocytes are thought to develop from immature transitional cells in a BCR-dependent manner. We have previously shown that BCR cross-linking in vitro results in death of early transitional (T1) B cells, whereas late transitional (T2) B cells survive and display phenotypic characteristics of mature FoB cells. We now demonstrate that diacylglycerol (DAG), a lipid second messenger implicated in cell survival and differentiation, is produced preferentially in T2 compared with T1 B cells upon BCR cross-linking. Consistently, inositol 1,4,5-triphosphate is also produced preferentially in T2 compared with T1 B cells. Unexpectedly, the initial calcium peak appears similar in both T1 and T2 B cells, whereas sustained calcium levels are higher in T1 B cells. Pretreatment with 2-aminoethoxydiphenylborate, an inhibitor of inositol 1,4,5-triphosphate receptor-mediated calcium release, and verapamil, an inhibitor of L-type calcium channels, preferentially affects T1 B cells, suggesting that distinct mechanisms regulate calcium mobilization in each of the two transitional B cell subsets. Finally, BCR-mediated DAG production is dependent upon Bruton’s tyrosine kinase and phospholipase C-γ2, enzymes required for the development of FoB from T2 B cells. These results suggest that calcium signaling in the absence of DAG-mediated signals may lead to T1 B cell tolerance, whereas the combined action of DAG and calcium signaling is necessary for survival and differentiation of T2 into mature FoB lymphocytes.

B Cell Receptor and BAFF Receptor Signaling Regulation of B Cell Homeostasis

“…Thus, upon differentiation, T2 cells display enhanced expression of CD19, CD21, and BR3 (Ref. 15 and data not shown). This increases BCR signaling competence and BAFF-dependent survival potential of T2 relative to T1 B cells as was recently suggested (4).…”

Interferon‐γ–dependent inhibition of B cell activation by bone marrow–derived mesenchymal stem cells in a murine model of systemic lupus erythematosus

“…We isolated B cells from the spleens of (NZB ϫ NZW)F 1 mice by negative enrichment with the surface marker CD43 to avoid the inadvertent activation of B cells. CD43-negative cells were then sorted with antibodies specific for CD21, CD24, and CD23 to purify the follicular (CD23ϩCD24-CD21ϩ) and marginal zone (CD23-CD24ϩCD21ϩ) B cells (14).…”

Inositol 1,4,5-Trisphosphate 3-Kinase B Is a Negative Regulator of BCR Signaling That Controls B Cell Selection and Tolerance Induction

“…Of the many signals that occur following BCR activation, the elevation in cytosolic calcium is a particularly attractive candidate signal for linking BCR signal strength to fate determination. The degree of intracellular Ca 2ϩ elevation is proportional to the avidity of the BCR for self-ligands, and several studies have correlated alterations in intracellular Ca 2ϩ concentration ([Ca 2ϩ ] i ) 2 following BCR stimulation with changes in B cell fate (3)(4)(5)(6)(7)(8). Unfortunately, most mutations of the BCR or the downstream signaling proteins that alter B cell fate lead to a compendium of alterations in BCR signaling.…”