Detection of direct binding of human herpesvirus 8-encoded interleukin-6 (vIL-6) to both gp130 and IL-6 receptor (IL-6R) and identification of amino acid residues of vIL-6 important for IL-6R-dependent and -independent signaling

Human herpesvirus 8 (HHV-8) is associated with Kaposi's sarcoma, primary ef fusion lymphoma, and multicentric Castleman's disease; in all of these dise ases, interleukin-6 (IL-6) has been implicated as a likely mitogenic and/or angiogenic factor, HHV-8 encodes a homologue of IL-6 (viral IL-6 [vIL-6]) that has been shown to be biologically active in several assays and whose a ctivities mirror those of its mammalian counterparts. Like these proteins, vIL-6 mediates its effects through the gp130 signal transducer, but signali ng is not dependent on the structurally related IL-6 receptor (IL-6R; gp80) subunit of the receptor-signal transducer complex. However, as we have sho wn previously, IL-6R can enhance vIL-6 signal transduction and can enable s ignaling through a gp130 variant (gp130.PM5) that is itself unable to suppo rt vIL-6 activity, indicating that IL-6R can form part of the signaling com plex. Also, our analysis of a panel of vIL-6 mutants in transfection experi ments in Hep3B cells (that express IL-6R and gp130) showed that most were a ble to function normally in this system. Here, we have used in vitro vIL-6- receptor binding assays to demonstrate direct binding of vIL-6 to both gp13 0 and IL-6R and vIL-6-induced gp130-IL-6R complex formation, and we have ex tended our functional analyses of the vIL-6 variants to identify residues i mportant for IL-6R-independent and IL-6R-dependent signaling through native gp130 and gp130.PM5, respectively. These studies have identified residues in vIL-6 that are important for IL-6R-independent and IL-6R-mediated functi onal complex formation between vIL-6 and gp130 and that may be involved dir ectly in binding to gp130 and IL-6R.