Kaposi's sarcoma-associated herpesvirus glycoproteins B and K8.1 regulate virion egress and synthesis of vascular endothelial growth factor and viral interleukin-6 in BCBL-1 cells.
Kaposi's sarcoma-associated herpesvirus (KSHV) viral glycoproteins play important roles in the infectious life cycle and have been implicated in KSHV-associated endothelial cell transformation, angiogenesis, and KS-induced malignancies. KSHV-associated primary effusion lymphomas (PELs) secrete high levels of vascular endothelial growth factor (VEGF) and viral interleukin-6 (vIL-6) in vitro and VEGF, vIL-6, and basic-fibroblast growth factor (b-FGF) in mouse xenografts. KSHV-encoded glycoproteins B (gB) and K8.1 stimulate VEGF secretion, most likely mediated by direct or indirect binding to cell surface receptors, including the gB-specific alphaVbeta3 and alpha3beta1 integrins. In this study, the short interfering RNA (siRNA)-mediated inhibition of either gB or K8.1 transcription by anti-gB or -K8.1 siRNAs caused a substantial reduction in virion egress and a decrease in both vIL-6 and VEGF production. Similarly, the treatment of BCBL-1 cells with anti-gB or anti-K8.1 antibodies caused a substantial reduction in vIL-6 and VEGF production. Codon-optimized versions of either wild-type gB, mutant gB having the RGD amino acid motif changed to RAA, or K8.1 efficiently rescued virion egress and VEGF and vIL-6 production. These results suggest that the binding of gB via its RGD motif to integrin receptors was not responsible for the observed gB-associated regulation of VEGF and vIL-6 transcription. Conditioned medium collected from BCBL-1 cells transfected with anti-gB and anti-K8.1 siRNAs or treated with anti-gB and anti-K8.1 antibodies exhibited a significantly reduced ability to induce the formation of the capillary network of endothelial cells compared to the ability of medium from mock-infected BCBl-1 cells. Furthermore, medium obtained from BCBL-1 cells expressing smaller amounts of gB and K8.1 produced a substantial reduction in endothelial cell migration in a vertical migration assay compared to that of control medium containing wild-type levels of gB and K8.1. These results suggest a functional linkage between gB/K8.1 synthesis and VEGF/vIL-6 transcriptional regulation via paracrine and/or autocrine signaling pathways.
Subramanian
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R
Department of Pathobiological Sciences, School of Veterinary Medicine, Louisiana State University, Baton Rouge, LA, USA.
Sehgal
I
I
D'Auvergne
O
O
Kousoulas
K G
KG
D019943:Amino Acid Substitution; D000818:Animals; D001483:Base Sequence; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D017931:DNA Primers / Q000235:genetics; D042783:Endothelial Cells / Q000821:virology; D005814:Genes, Viral; D006023:Glycoproteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D019288:Herpesvirus 8, Human / Q000235:genetics / Q000472:pathogenicity* / Q000502:physiology*; D006801:Humans; D015850:Interleukin-6 / Q000096:biosynthesis* / Q000235:genetics; D051379:Mice; D016297:Mutagenesis, Site-Directed; D009389:Neovascularization, Pathologic; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis* / Q000235:genetics; D014759:Viral Envelope Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D014764:Viral Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*
D017931:DNA Primers; D006023:Glycoproteins; D015850:Interleukin-6; C113454:K8.1 protein, Human herpesvirus 8; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D011994:Recombinant Proteins; D042461:Vascular Endothelial Growth Factor A; D014759:Viral Envelope Proteins; D014764:Viral Proteins; C116207:glycoprotein B, human herpesvirus 8