Kaposi's sarcoma-associated herpesvirus induces sustained NF-kappaB activation during de novo infection of primary human dermal microvascular endothelial cells that is essential for viral gene expression.
In vitro Kaposi's sarcoma-associated herpesvirus (KSHV) infection of primary human dermal microvascular endothelial (HMVEC-d) cells and human foreskin fibroblast (HFF) cells is characterized by the induction of preexisting host signal cascades, sustained expression of latency-associated genes, transient expression of a limited number of lytic genes, and induction of several cytokines, growth factors, and angiogenic factors. Since NF-kappaB is a key molecule involved in the regulation of several of these factors, here, we examined NF-kappaB induction during de novo infection of HMVEC-d and HFF cells. Activation of NF-kappaB was observed as early as 5 to 15 min postinfection by KSHV, and translocation of p65-NF-kappaB into nuclei was detected by immunofluorescence assay, electrophoretic mobility shift assay, and p65 enzyme-linked immunosorbent assay. IkappaB phosphorylation inhibitor (Bay11-7082) reduced this activation significantly. A sustained moderate level of NF-kappaB induction was seen during the observed 72 h of in vitro KSHV latency. In contrast, high levels of ERK1/2 activation at earlier time points and a moderate level of activation at later times were observed. p38 mitogen-activated protein kinase was activated only at later time points, and AKT was activated in a cyclic manner. Studies with UV-inactivated KSHV suggested a role for virus entry stages in NF-kappaB induction and a requirement for KSHV viral gene expression in sustained induction. Inhibition of NF-kappaB did not affect target cell entry by KSHV but significantly reduced the expression of viral latent open reading frame 73 and lytic genes. KSHV infection induced the activation of several host transcription factors, including AP-1 family members, as well as several cytokines, growth factors, and angiogenic factors, which were significantly affected by NF-kappaB inhibition. These results suggest that during de novo infection, KSHV induces sustained levels of NF-kappaB to regulate viral and host cell genes and thus possibly regulates the establishment of latent infection.
Sadagopan
Sathish
S
Department of Microbiology and Immunology, H. M. Bligh Cancer Research Laboratories, Chicago Medical School, Rosalind Franklin University of Medicine and Science, 3333 Green Bay Road, North Chicago, IL 60064, USA.
Sharma-Walia
Neelam
N
Veettil
Mohanan Valiya
MV
Raghu
Hari
H
Sivakumar
Ramu
R
Bottero
Virginie
V
Chandran
Bala
B
D021581:Active Transport, Cell Nucleus; D043925:Angiogenesis Inducing Agents / Q000378:metabolism; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D016207:Cytokines / Q000096:biosynthesis; D042783:Endothelial Cells / Q000378:metabolism / Q000821:virology*; D005347:Fibroblasts / Q000821:virology; D015870:Gene Expression; D005786:Gene Expression Regulation; D005814:Genes, Viral; D019288:Herpesvirus 8, Human / Q000502:physiology*; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000096:biosynthesis; D008856:Microscopy, Fluorescence; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D008954:Models, Biological; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009570:Nitriles / Q000494:pharmacology; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D013450:Sulfones / Q000494:pharmacology; D014157:Transcription Factors / Q000096:biosynthesis; D053586:Virus Internalization; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
C434003:3-(4-methylphenylsulfonyl)-2-propenenitrile; D043925:Angiogenesis Inducing Agents; D016207:Cytokines; D036341:Intercellular Signaling Peptides and Proteins; D016328:NF-kappa B; D009570:Nitriles; D013450:Sulfones; D014157:Transcription Factors; D051057:Proto-Oncogene Proteins c-akt; D048052:Mitogen-Activated Protein Kinase 3; D048051:p38 Mitogen-Activated Protein Kinases