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 effusion lymphoma, and multicentric Castleman's disease; in all of these diseases, 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 activities mirror those of its mammalian counterparts. Like these proteins, vIL-6 mediates its effects through the gp130 signal transducer, but signaling is not dependent on the structurally related IL-6 receptor (IL-6R; gp80) subunit of the receptor-signal transducer complex. However, as we have shown previously, IL-6R can enhance vIL-6 signal transduction and can enable signaling through a gp130 variant (gp130.PM5) that is itself unable to support vIL-6 activity, indicating that IL-6R can form part of the signaling complex. Also, our analysis of a panel of vIL-6 mutants in transfection experiments in Hep3B cells (that express IL-6R and gp130) showed that most were able 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 gp130 and IL-6R and vIL-6-induced gp130-IL-6R complex formation, and we have extended our functional analyses of the vIL-6 variants to identify residues important 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 functional complex formation between vIL-6 and gp130 and that may be involved directly in binding to gp130 and IL-6R.
The Molecular Virology Laboratories, Department of Oncology, Johns Hopkins University, Baltimore, Maryland 21231, USA. m.havenga@crucell.com
H
H
Li
H
H
Wang
J
J
Nicholas
D015703:Antigens, CD / Q000378:metabolism*; D002478:Cells, Cultured; D050822:Cytokine Receptor gp130; D019288:Herpesvirus 8, Human / Q000378:metabolism*; D006801:Humans; D015850:Interleukin-6 / Q000737:chemistry / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000378:metabolism*; D019947:Receptors, Interleukin-6 / Q000502:physiology*; D015398:Signal Transduction; D013329:Structure-Activity Relationship; D014764:Viral Proteins / Q000737:chemistry / Q000378:metabolism*
D015703:Antigens, CD; C494199:IL6ST protein, human; D015850:Interleukin-6; D008562:Membrane Glycoproteins; D019947:Receptors, Interleukin-6; D014764:Viral Proteins; D050822:Cytokine Receptor gp130