Blimp-1 was originally shown to regulate the expression of structural genes (J chain and Syndecan-1) during the differentiation of mature B lymphocytes into plasma cells (13
). Recently, it has been found that the expression of Blimp-1 in pre–B cells triggers an apoptotic response that appears to be mediated by the downregulation of c-myc expression (17
). Here we report that Blimp-1 can induce apoptosis across a broad range of immature and mature B cell lymphoma lines and can only promote differentiation in fully activated B cells. We also have identified the functional domain required for the induction of apoptosis and anticipate that Blimp-1 functions may depend upon the stage-specific expression of a cofactor.
The developmental stages represented by the 5B1b and 3B3 sublines appear to define a temporal window of B cell differentiation within which the function of Blimp-1 shifts. Partially activated 5B1b cells are impaired in their responses to IL-2/IL-5 stimulation and arrested at the centroblast stage. On the other hand, fully activated 3B3 cells respond to IL-2/IL-5 stimulation and differentiate into centroblast and, later, centrocyte-like cells (Fig. ). The transfection of 5B1b cells with the NZ dominant negative suppresses blasting and IgM secretion induced by IL-2/IL-5 treatment (Fig. ), thus indicating that Blimp-1 is an essential component in IL-2/IL-5 signaling during B cell maturation. In contrast to the antidifferentiation effect of NZ, overexpression of Blimp-1 in 5B1b but not in 3B3 cells results in marked apoptotic death that can be suppressed by the NZ dominant negative. This, in turn, indicates that the Blimp-1–mediated apoptotic machinery is still functional in the 5B1b cells. Similarly, the transfection of Blimp-1 in earlier stages of the B cell development, such as the pre–B (18-81), immature B (WEHI-231) and mature B (L10A) cell stages induces an apoptotic response. This effect is mainly dependent on a 69-aa region within the proline-rich domain (Fig. ). It is conceivable that the binding of an additional factor to this domain is required to elicit a response. For example, Blimp-1 may interact in vitro with YY1, a positive regulator of c-myc (17
), whose expression plays an important role in controlling the cell fate. Since the downregulation of c-myc was shown to be involved in the induction of apoptosis in both 18-81 (17
) and WEHI-231 (46
), its suppression by Blimp-1 is anticipated to play a role in the elimination of partially activated B cells. However, it appears that other genes that are involved in the regulation of the cell cycle may be targeted by Blimp-1.
The PR domain (aa 131-231) (16
) was suggested to modulate the repressor ability of RIZ (47
). Our results show that the deletion of 50% of this domain in the ΔN′ truncated protein had no significant effect on the ability of this protein to induce apoptosis in WEHI-231 cells. Therefore, the integrity of the PR domain may be unnecessary for the induction of apoptosis. In fact, the truncation of either the N′ or C′ acidic domains had only a minor effect on the apoptotic ability of Blimp-1 (Fig. B
Given that Blimp-1 is able to exert such different effects in closely related cells, the question arises as to how this is accomplished. One possible explanation is that different combinations of the zinc finger motifs may recognize distinct gene targets. A mechanism of this type was recently shown in the case of NeP1/CTCF, an 11–zinc finger protein that uses different combination of its zinc fingers to bind to either the c-myc gene or the lysozyme silencer, F1 (48
). The partial use of Blimp-1 zinc fingers for the recognition of potential targets was demonstrated in the work of Keller and Maniatis (32
). In that report the authors show that two out of the five PRDI-BF1 zinc fingers (and a stretch of 65 aa located N′ to these fingers) are sufficient for its binding to the PRDI element. Recently, we have found that the binding of NZ to the PRF1 element, but not to the PRD1 element, is sensitive to posttranslational modifications or conformational changes (Chi, J.-T., and E.J. Messika, unpublished results). Thus, Blimp-1 may be modulated to switch specificity to different targets due to interactions with stage-specific proteins, using different combinations of its zinc fingers, as the cell matures.
In summary, we propose that Blimp-1 may be part of a “gate keeper” mechanism in the differentiation process, as its upregulation causes apoptosis of immature or partially stimulated B cells. This may constitute an important selection mechanism to eliminate self-reactive B cells that have escaped earlier negative selection mechanisms or have been inappropriately (and partially) activated.