Human Herpesvirus (HHV) 8 K1 Suppresses Fas-Mediated Apoptosis and Binds to the Fas Complex.

Kaposi’s sarcoma-associated herpesvirus (KSHV or HHV-8) is associated with multicentric Castleman’s disease, primary effusion lymphoma (PEL), and Kaposi’s sarcoma. Malignant tumors are frequently associated with resistance to Fas-mediated apoptosis, which is a common obstacle to the successful treatment of many different types of cancers. Oncoprotein K1 encoded by HHV-8 is a transmembrane protein with an immunoreceptor tyrosine-based activation motif (ITAM) sequence in its short cytoplasmic domain, which is constitutively phosphorylated (J Natl Cancer Inst Monogr. 28; 15–23, 2001). We have found that K1-expressing cells are resistant to apoptosis and K1 directly binds Fas to inhibit apoptosis. K1-transfected BJAB cells were resistant to Fas-induced apoptosis, but not to other stimuli including tumor necrosis-related apoptosis-inducing ligand or radiation. K1 transgenic mice showed lymph node hyperplasia and large spleens. Nine out of 12 K1 transgenic mice survived injection of 0.3 mg/gram activating anti-Fas (Jo2) antibody whereas 13 out of 22 control mice died with hepatocyte apoptosis (p=0.03). We prepared deletion and point mutants of K1 and Fas to map their respective binding sites.

Immunoprecipitation-immunoblotting analysis of the transfectant cell lysates showed that binding occurred through the extracellular domains of K1 and Fas. Also, we have found that preligand assembly domain-deleted Fas mutants (amino acids 1-66 deleted) still bind to K1. The deletion mutants encompassing cysteine-rich domains 1, 2, and 3 of the extracellular domain of Fas are currently being tested for mediating binding to K1. These results support the presence of an apoptosis regulatory system based on Fas-protein interactions. By characterizing this mechanism, we will lay the foundation of an apoptosis regulatory pathway that may explain apoptosis resistance of many cancers.