Persistent calcium elevation correlates with the induction of surface immunoglobulin-mediated B cell DNA synthesis.

Abstract

SummarySurface immunoglobulin (slg)-mediated stimulation of B lymphocytes induces a tyrosine kinase-dependent sequence of events leading to rapid and large elevations in intracellular ionized calcium ([Ca2+]i). These early biochemical events do not necessarily lead to proliferation of B cells, however, and conversely, the absence of or inhibition of these events does not necessarily prevent cellular proliferation. We now show by digital image analysis of single B cells that conditions which lead to B cell proliferation are associated with low-level but persistent sustained or cyclic elevations in [Ca 2 + ]i. In marked contrast, early and nonsustained elevations in [Ca 2 § ]i are induced in B cells by stimuli that lead to G1 transition but fail to progress to DNA synthesis. Thus, when B cells were stimulated with mitogenic and nonmitogenic anti-IgD antibodies, both of which induce entry of cells into G1 and early calcium transients of comparable magnitude, persistent low-level calcium elevations were only detected in cells stimulated with the mitogenic antibody. Furthermore, persistent calcium elevations were also seen when B cells were stimulated with a multivalent dextran-anti-Ig conjugate which induced very high levels of B cell proliferation in the absence of detectable phosphatidylinositol 4,5-biphosphate hydrolysis or elevations in [Ca2+]i as detected by flow cytometry. Finally, B cells from X-linked B cell-defective mice, which do not proliferate in response to anti-Ig antibody, show marked and early increases in [Ca2+]i, but do not show persistent calcium elevations. These data suggest that the rapid and large increases of [Ca 2 + ]i seen in lymphocytes within seconds after antigen receptor ligation may be associated with entry in G1, whereas low-level but persistent elevations may be the hallmark of a cell destined to synthesize DNA. kinase (8-10), all of which are intimately involved in cell growth of B cells as well as other cell types. Although it is clear that the summation of these events reflects an early response to sIg-mediated signal transduction in B cells which will lead to cell growth, the specific role of each event in regulating entry of the cell into G1 or S of the cell cycle is not known. Experiments both in human and in murine systems suggest that there is no correlation between the ability of an antigen or of anti-Ig antibody to induce phosphatidylinositol 4,5-biphosphate (PIP2) activation or tyrosine kinase activation and its ability to induce B cell DNA synthesis (11)(12)(13)(14). Thus, for example, inhibition of sIg-mediated PiP2 hydrolysis does not prevent B cell proliferation (15). Furthermore, stimulation of these early events by anti-Ig antibodies does not necessarily lead to B cell proliferation (13,16,17).

B cell TLRs and induction of immunoglobulin class-switch DNA recombination

“…Phosphorylation of CD79a Tyr182 by LPS, however, was slower than that following induction by anti–δ mAb/dex or soluble/monomeric anti–Igμ antibody. Also, like anti–δ mAb/dex, which induced a continuous increase of the free cytosolic Ca 2+ ion level, as a hallmark of persistent BCR signaling (200), LPS, but not lipid A or CD154, induced Ca 2+ elevation, which was detectable within minutes of stimulation and further increased after a longer period of stimulation (23). Furthermore, B cells deficient in known BCR signaling elements, such as CD19, Btk, Blnk, Lyn or Vav, displayed greatly reduced responses to LPS (201-205).…”

“…PI(3)K generates phosphatidylinositol (3,4,5) triphosphate on the inner leaflet of the plasma membrane, where signaling molecules such as phospholipase Cγ2 (PLCγ2) and AKT are recruited upon binding of phosphatidylinositol (3,4,5) triphosphate through their pleckstrin homology (PH) domain. Membrane-localized PLCγ2 cleaves phosphatidylinositol lipids to generate diacylglycerol (DAG), which activates protein kinase C-β (PKC-β) (224), and inositol triphosphate (IP3), which results in elevation of intracellular Ca 2+ levels (200, 225, 226). PKC-β activates the CARMA1 signalosome complex composed of CARMA1, BCL10 and MALT1, resulting in TRAF6 ubiquitination and subsequent TAK1-mediated IKK phosphorylation for canonical NF-κB activation and/or TAK1-triggered MAPK cascades for JNK, ERK and/or p38 activation (227).…”

“…In experiments showing BCR activation of the TAK1-IKK-canonical NF-κB pathway, BCR signaling was triggered by soluble/monomeric anti–μ antibody, which, however, does not extensively crosslink BCRs in Igδ hi Igμ lo mature B cells (200). BCR crosslinking by anti–δ mAb/dex did not induce immediate canonical NF-κB (p65) activation; rather, it increased the levels of the non-canonical NF-κB p52 subunit, which is processed from its precursor p100, within 24 – 48 h (the time frame of AID induction) in a fashion dependent on PI(3)K (23).…”

Activation-induced cell death in B lymphocytes

“…Indeed, for DNA synthesis to occur and for B cells to proliferate upon BCR crosslinking, there must be a small, but sustained increase in cytoplasmic free calcium [14]. Furthermore, B cell stimulation via BCR crosslinking can be successfully mimicked using calcium ionophores (such as ionomycin) to increase cytoplasmic free calcium, in combination with phorbol esters (such as PMA) to activate PKC [15].…”

Insulin-Like Growth Factor-1 Controls Type 2 T Cell-Independent B Cell Response

“…Previous studies have demonstrated that multivalent cross-linking of membrane IgD and specific costimulation with IL-4 and IL-5 serve as such a model for TI-2 responses (46,47). Therefore, we incubated purified splenic B cells from wt C57BL/6 mice with a dextran-coupled anti-IgD mAb (␣␦-dex), IL4, IL-5, and IGF-1, alone or in combination (Fig.…”