By differential screening of tumor necrosis factor α (TNF-α) and lipopolysaccharide (LPS)- activated endothelial cells (ECs), we have identified a cDNA clone that turned out to be a member of the inhibitor of apoptosis (iap) gene family. iap genes function to protect cells from undergoing apoptotic death in response to a variety of stimuli. These iap genes, hiap1, hiap2, and xiap were found to be strongly upregulated upon treatment of ECs with the inflammatory cytokines TNF-α, interleukin 1β, and LPS, reagents that lead to activation of the nuclear transcription factor κB (NF-κB). Indeed, overexpression of IκBα, an inhibitor of NF-κB, suppresses the induced expression of iap genes and sensitizes ECs to TNF-α–induced apoptosis. Ectopic expression of one member of the human iap genes, human X-chromosome–linked iap (xiap), using recombinant adenovirus overrules the IκBα effect and protects ECs from TNF-α– induced apoptosis. We conclude that xiap represents one of the NF-κB–regulated genes that counteracts the apoptotic signals caused by TNF-α and thereby prevents ECs from undergoing apoptosis during inflammation.

Exposure of endothelial and many other cell types to tumor necrosis factor ␣ generates both apoptotic and anti-apoptotic signals. The anti-apoptotic pathway leads to activation of the transcription factor NF-B that regulates the expression of genes such as A20 or members of the IAP gene family that protect cells from tumor necrosis factor ␣-mediated apoptosis. In turn, some anti-apoptotic genes have been shown to modulate NF-B activity. Here we demonstrate that XIAP, a NF-Bdependent member of the IAP gene family, is a strong stimulator of NF-B. Expression of XIAP leads to increased nuclear translocation of the p65 subunit of NF-B via a novel signaling pathway that involves the mitogen-activated protein kinase kinase kinase TAK1. We show that TAK1 physically interacts with NIK and with IKK2, and both XIAP or active TAK1 can stimulate IKK2 kinase activity. Thus, XIAP may be part of a system of regulatory loops that balance a cell's response to environmental stimuli.

The human invariant chain (I gamma) of class II histocompatibility antigens spans the membrane of the endoplasmic reticulum once. It exposes a small amino-terminal domain on the cytoplasmic side and a carboxy-terminal, glycosylated domain on the exoplasmic side of the membrane. When the exoplasmic domain of I gamma is replaced by the cytoplasmic protein chloramphenicol acetyltransferase (CAT), CAT becomes the exoplasmic, glycosylated domain of the resulting membrane protein I gamma CAT. Deletion of the hydrophilic cytoplasmic domain from I gamma CAT gives rise to a secreted protein from which an amino-terminal segment is cleaved, most likely by signal peptidase. We conclude that the membrane-spanning region of I gamma contains a signal sequence in its amino-terminal half and that hydrophilic residues at the amino-terminal end of a signal sequence can determine cleavage by signal peptidase.

Abstract. The intracellular transport and location of major histocompatibility complex (MHC) class II molecules and associated invariant chain (Ii) were investigated in a human melanoma cell line. In contrast to the class II molecules, which remain stable for >4 h after synthesis, the associated Ii is proteolytically processed within 2 h. During or shortly after synthesis the N112-terminal cytoplasmic and membrane-spanning segment is in some of the Ii molecules cleaved off ; during intracellular transport, class II associated and membrane integrated Ii is processed from its COOH terminus in distinct steps in endocytic compartments .Immunocytochemical studies at the light and electron microscopic level revealed the presence of class II molecules, but not of Ii on the cell surface . Intracel-

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