Cytokines in the tumor necrosis factor (TNF) family regulate development and function of the immune system. We have isolated a new member of this family, designated Apo-2 ligand (Apo-2L), via an expressed sequence tag. Apo-2L is a 281-amino acid protein, related most closely to Fas/Apo-1 ligand. Transfected Apo-2L is expressed at the cell surface with its C terminus exposed, indicating a type II transmembrane protein topology. Like Fas/Apo-1 ligand and TNF, the C-terminal extracellular region of Apo-2L (amino acids 114 -281) exhibits a homotrimeric subunit structure. Soluble Apo-2L induces extensive apoptosis in lymphoid as well as non-lymphoid tumor cell lines. The effect of Apo-2L is not inhibited by soluble Fas/Apo-1 and TNF receptors; moreover, expression of human Fas/Apo-1 in mouse fibroblasts, which confers sensitivity to induction of apoptosis by agonistic anti-Fas/Apo-1 antibody, does not confer sensitivity to Apo-2L. Hence, Apo-2L acts via a receptor which is distinct from Fas/Apo-1 and TNF receptors. These results suggest that, along with other family members such as Fas/Apo-1 ligand and TNF, Apo-2L may serve as an extracellular signal that triggers programmed cell death.
TNF1 family cytokines modulate host defense mechanisms via a corresponding family of receptors (1-3). Most TNF family cytokines are expressed as type II transmembrane proteins, whose C-terminal extracellular domain is processed proteolytically to form a soluble homotrimeric molecule. In contrast, the members of the TNF receptor (TNFR) family are type I transmembrane proteins. In both families, sequence homology is found mainly in the extracellular regions, which mediate ligand-receptor binding.Members of the TNF family have diverse biological actions, including T cell co-stimulation, induction of B cell proliferation and differentiation, and macrophage activation (1-3). In addition, certain TNF family members regulate the elimination of unwanted immune cells by inducing programmed cell death (apoptosis). For example, Fas/Apo-1 ligand (Fas/Apo-1L) plays a key role in peripheral deletion of self-reactive lymphocytes, as suggested by the autoimmune phenotype of the mouse mutations lpr and gld, which occur, respectively, in the Fas/Apo-1 receptor or ligand genes (4, 5). In addition, Fas/Apo-1L is involved in apoptosis of B cells and CD4 ϩ T cells subsequent to stimulation by antigen, while TNF mediates a similar function in CD8ϩ T cells (4 -7). In this article, we describe a new member of the TNF cytokine family. We have designated this protein Apo-2L, because it resembles the Fas/Apo-1L in its amino acid sequence, as well as in its ability to induce apoptosis. Apo-2L appears to act via a receptor which is distinct from Fas/Apo-1 and TNF receptors, suggesting that its biological role is unique.
EXPERIMENTAL PROCEDURESCloning of Apo-2L cDNA-To isolate a full-length cDNA that contains the sequence of expressed sequenced tag (EST) HHEA47M, a gt11 bacteriophage library of human placental cDNA (ϳ1 ϫ 10 6 clones) (HL1075b, Clontech) was screene...