Molecular signals in B cell activation. II. IL-2-mediated signals are required in late G1 for transition to S phase after ionomycin and PMA treatment.
We report that sustained increase of intracellular calcium ion concentration and protein kinase C (PKC) activation maintained throughout the G1 phase of cell cycle do not provide sufficient signals to cause S-phase entry in rabbit B cells, and that additional signals transduced by IL-2 and IL-2 receptor interaction are essential for G1 to S transition. We have shown earlier that rabbit B cells can be activated to produce IL-2 and express functional IL-2 receptors after treatment with ionomycin and PMA. Herein we have compared the response of rabbit PBLs, which contain about 50% T cells, with those of purified B cells. After activation with ionomycin or PMA, comparable numbers of PBLs and B cells entered the cell cycle; but DNA synthesis by the PBL cultures was three to four times higher than that of cultures of purified B cells. Interestingly, IL-2 production by the PBL cultures was also three to four times higher than in B cell cultures, suggesting an involvement of IL-2 in inducing DNA synthesis in these cells. The hypothesis that IL-2, which is produced in early G1, acts in late G1 and is required for G1 to S transition in B cells was supported by the following observations: (i) IL-2 production by B cells was detected as early as 6 hr after activation and preceded DNA synthesis by at least 24 hr. (ii) B cell blasts in G1 (produced by treatment of resting B cells with ionomycin and PMA) showed DNA synthesis in response to IL-2, but showed very little DNA synthesis in response to restimulation with ionomycin and PMA. (iii) A polyclonal rabbit anti-human IL-2 antibody caused nearly complete inhibition of DNA synthesis by B cells activated by ionomycin and PMA. (iv) A PKC inhibitor, K252b, inhibited DNA synthesis in ionomycin and PMA-stimulated cells if added at the beginning of culture but was not inhibitory if added 16 hr later. We conclude that increased [Ca2+]i and PKC activation are not sufficient signals for G1 to S transition in B cells; entry into S is signaled by IL-2, and IL-2-mediated signal transduction probably does not involve increased [Ca2+]i or PKC activation.
Lagoo
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A
Department of Pathology and Laboratory Medicine, University of Texas, Medical School, Houston 77030.
Tseng
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CK
Sell
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D000818:Animals; D001402:B-Lymphocytes / Q000502:physiology*; D002118:Calcium / Q000502:physiology; D002453:Cell Cycle; D002478:Cells, Cultured; D004247:DNA / Q000096:biosynthesis; D066298:In Vitro Techniques; D007376:Interleukin-2 / Q000502:physiology*; D015759:Ionomycin / Q000008:administration & dosage*; D008213:Lymphocyte Activation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000502:physiology; D011817:Rabbits; D015375:Receptors, Interleukin-2 / Q000502:physiology; D015398:Signal Transduction; D013755:Tetradecanoylphorbol Acetate / Q000008:administration & dosage*; D013997:Time Factors
D007376:Interleukin-2; D015375:Receptors, Interleukin-2; D015759:Ionomycin; D004247:DNA; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium