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An expression cloning approach was employed to identify the receptor for B-lymphocyte stimulator (BLyS) and identified the tumor necrosis factor receptor superfamily member TACI as a BLyS-binding protein. Expression of TACI in HEK293T cells confers on the cells the ability to bind BLyS with subnanomolar affinity. Furthermore, a TACI-Fc fusion protein recognizes both the cleaved, soluble form of BLyS as well as the membrane BLyS present on the cell surface of a recombinant cell line. TACI mRNA is found predominantly in B-cells and correlates with BLyS binding in a panel of B-cell lines. We also demonstrate that TACI interacts with nanomolar affinity with the BLyS-related tumor necrosis factor homologue APRIL for which no clear in vivo role has been described. BLyS and APRIL are capable of signaling through TACI to mediate NF-kappaB responses in HEK293 cells. We conclude that TACI is a receptor for BLyS and APRIL and discuss the implications for B-cell biology.

CIS is a cytokine-induced SH2-containing protein that was originally cloned as an interleukin (IL)-3-inducible gene. CIS is known to associate with the IL-3 receptor beta chain and erythropoietin receptor and to inhibit signaling mediated by IL-3 and erythropoietin. We now demonstrate that CIS also interacts with the IL-2 receptor beta chain (IL-2Rbeta). This interaction requires the A region of IL-2Rbeta (residues 313-382), which also mediates the association of IL-2Rbeta with Lck and Jak3. Correspondingly, CIS inhibits functions associated with both of these kinases: Lck-mediated phosphorylation of IL-2Rbeta and IL-2-mediated activation of Stat5. Thus, we demonstrate that CIS can negatively control at least two independent IL-2 signaling pathways. Although a functional SH2 binding domain of CIS was not required for its interaction with IL-2Rbeta in vitro, its phosphotyrosine binding capability was essential for the inhibitory action of CIS. On this basis, we have generated a mutant form of CIS protein with an altered SH2 domain that acts as a dominant negative and should prove useful in further understanding CIS action.

The interleukin-2 receptor alpha chain (IL-2Ralpha) is potently induced by antigens, mitogens, and certain cytokines that include IL-2 itself. This induction leads to the formation of high affinity IL-2 receptors when IL-2Ralpha is co-expressed with the beta (IL-2Rbeta) and gamma (gammac) chains of this receptor. We investigated the signaling pathways mediating IL-2-induced IL-2Ralpha mRNA expression using 32D myeloid progenitor cells stably transfected with either wild type IL-2Rbeta or mutants of IL-2Rbeta containing tyrosine to phenylalanine substitutions. Of the six cytoplasmic tyrosines in IL-2Rbeta, we have found that only the two tyrosines that mediate Stat5 activation (Tyr-392 and Tyr-510) contribute to IL-2-induced IL-2Ralpha gene expression and that either tyrosine alone is sufficient for this process. Interestingly, the IL-7 receptor contains a tyrosine (Tyr-429)-based sequence resembling the motifs encompassing Tyr-392 and Tyr-510 of IL-2Rbeta. Further paralleling the IL-2 system, IL-7 could activate Stat5 and drive expression of IL-2Ralpha mRNA in 32D cells transfected with the human IL-7R. However, IL-3 could not induce IL-2Ralpha mRNA in 32D cells, despite its ability to activate Stat5 via the endogenous IL-3 receptor. Moreover, the combination of IL-3 and IL-2 could not "rescue" IL-2Ralpha mRNA expression in cells containing an IL-2Rbeta mutant with phenylalanine substitutions at Tyr-392 and Tyr-510. These data suggest that Tyr-392 and Tyr-510 couple to an additional signaling pathway beyond STAT protein activation in IL-2-mediated induction of the IL-2Ralpha gene.