We have identified and characterized Diva, which is a novel regulator of apoptosis. Sequence analysis revealed that Diva is a member of the Bcl-2 family of proteins containing Bcl-2 homology domain 1, 2, 3, and 4 (BH1, BH2, BH3, and BH4) regions and a carboxyl-terminal hydrophobic domain. The expression of Diva mRNA was detected in multiple embryonic tissues but was restricted to the ovary and testis in adult mice. The expression of Diva promoted the death of 293T, Ramsey, and T47D cells as well as that of primary sensory neurons, indicating that Diva is a proapoptotic protein. Significantly, Diva lacks critical residues in the conserved BH3 region that mediate the interaction between BH3-containing proapoptotic Bcl-2 homologues and their prosurvival binding partners. Consistent with this, Diva did not bind to cellular Bcl-2 family members including Bcl-2, Bcl-X L , Bcl-w, Mcl-1, and A1/Bfl-1. Furthermore, mutants of Diva lacking the BH3 region fully retained their proapoptotic activity, confirming that Diva promotes apoptosis in a BH3-independent manner. Significantly, Diva interacted with a viral Bcl-2 homologue (vBcl-2) encoded by the Kaposi's sarcoma-associated herpesvirus. Consistent with these associations, apoptosis induced by Diva was inhibited by vBcl-2 but not by Bcl-X L . Importantly, Diva interacted with Apaf-1, an adapter molecule that activates caspase-9, a central death protease of the apoptotic pathway. The expression of Diva inhibited the binding of Bcl-X L to Apaf-1, as determined by immunoprecipitation assays. Thus, Diva represents a novel type of proapoptotic Bcl-2 homologue that promotes apoptosis independently of the BH3 region through direct binding to Apaf-1, thus preventing Bcl-X L from binding to the caspase-9 regulator Apaf-1.Apoptosis, a morphologically distinguished form of programmed cell death, is critical during development and tissue homeostasis and plays a role in the pathogenesis of a variety of diseases (1). Several regulatory components of the apoptotic pathway have been identified in various living organisms including man (1, 2). A family of proteins that includes Bcl-2, Bcl-X L , Bcl-w, Mcl-1, and A1/Bfl-1 shares conserved regions termed Bcl-2 homology domains 1, 2, 3, and 4 (BH1, BH2, BH3, and BH4) 1 and function by repressing apoptosis (3). The biochemical process by which Bcl-2 family members regulate cell death is poorly understood. Analyses of the nematode cell death regulators CED-3, CED-4, and CED-9 have provided important insight into the biochemical mechanism that regulates apoptosis. CED-9, the nematode homologue of Bcl-2 and Bcl-X L , binds to CED-4 and represses cell death by interacting and inhibiting the killing activity of CED-3 through CED-4 (4 -7). Recently, it was shown that Bcl-X L associates with Apaf-1, a mammalian homologue of CED-4, and inhibits the activation of procaspase-9 (8 -11).Whereas proteins like Bcl-2 and Bcl-X L inhibit cell death, structurally related proteins including Bax, Bak, Bad, Bik/ Nbk, Bid, Hrk, Bim, Bok/Mtd, and BNIP3 promote apo...

The intradermal inoculation in naive or in previously sensitized individuals of small amounts of Leishmania extract (Montenegro's skin test) induced or modulated, respectively, the immune response to Leishmania, as assessed by subsequent Montenegro's skin tests. These phenomena could hinder the interpretation of Montenegro's skin tests in a population already subjected to the test in the past and, in addition, could affect in an unknown way the development of mucosal lesions in people infected with L. braziliensis or L. amazonensis, since those lesions have been associated with hypersensitivity to Leishmania antigens. Anti-Leishmania antibody responses, assessed by enzyme-linked immunosorbent assay, were not induced in naive individuals by Montenegro's skin tests, but tended to become more intense following these tests in previously sensitized individuals.

SUMMARYSignalling through CD40 is essential for the development of immunoglobulin G (IgG) antibody responses, germinal centres and B-cell memory against T-dependent antigens. In addition, engagement of CD40 in B cells promotes cell survival by inducing the expression of anti-apoptotic members of the bcl-2 family of cell-death regulators. In the present study we analysed whether Tdependent immune responses can be developed in mice de®cient in CD40 if the anti-apoptotic activity mediated by the engagement of CD40 in B cells is compensated by the constitutive overexpression of anti-apoptotic genes of the bcl-2 family. We showed that the over-expression of either hbcl-2 or hbcl-x L transgenes in B cells is not suf®cient to restore IgG antibody responses and germinal centre formation in CD40-de®cient mice. These results indicate that CD40 functions, other than those mediated through survival, are required for the establishment of T-dependent B-cell responses.