Abstract

Kaposi's sarcoma-associated herpesvirus (KSHV), etiologically associated with Kaposi's sarcoma, uses integrins (α3β1, αVβ3, and αVβ5) and associated signaling to enter human dermal microvascular endothelial cells (HMVEC-d), an in vivo target of infection. KSHV infection activated c-Cbl, which induced the selective translocation of KSHV into lipid rafts (LRs) along with the α3β1, αVβ3, and xCT receptors, but not αVβ5. LR-translocated receptors were monoubiquitinated, leading to productive macropinocytic entry, whereas non-LR-associated αVβ5 was polyubiquitinated, leading to clathrin-mediated entry that was targeted to lysosomes. Because the molecule(s) that integrate signal pathways and productive KSHV macropinocytosis were unknown, we immunoprecipitated KSHV-infected LR fractions with anti-α3β1 antibodies and analyzed them by mass spectrometry. The tyrosine kinase EphrinA2 (EphA2), implicated in many cancers, was identified in this analysis. EphA2 was activated by KSHV. EphA2 was also associated with KSHV and integrins (α3β1 and αVβ3) in LRs early during infection. Preincubation of virus with soluble EphA2, knockdown of EphA2 by shRNAs, or pretreatment of cells with anti-EphA2 monoclonal antibodies or tyrosine kinase inhibitor dasatinib significantly reduced KSHV entry and gene expression. EphA2 associates with c-Cbl-myosin IIA and augmented KSHV-induced Src and PI3-K signals in LRs, leading to bleb formation and macropinocytosis of KSHV. EphA2 shRNA ablated macropinocytosis-associated signaling events, virus internalization, and productive nuclear trafficking of KSHV DNA. Taken together, these studies demonstrate that the EphA2 receptor acts as a master assembly regulator of KSHV-induced signal molecules and KSHV entry in endothelial cells and suggest that the EphA2 receptor is an attractive target for controlling KSHV infection.

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Authors

Chakraborty S, Veettil MV, Bottero V, Chandran B

Institution

H. M. Bligh Cancer Research Laboratories, Department of Microbiology and Immunology, Chicago Medical School, Rosalind Franklin University of Medicine and Science, North Chicago, IL 60064, USA.

Source

Proceedings of the National Academy of Sciences of the United States of America 109:19 2012 May 8 pg E1163-72

MeSH

Blotting, Western
Cells, Cultured
Endothelial Cells
Gene Expression Regulation, Viral
HEK293 Cells
Herpesvirus 8, Human
Host-Pathogen Interactions
Humans
Integrin alpha3beta1
Membrane Microdomains
Microscopy, Fluorescence
Pinocytosis
Protein Binding
RNA Interference
Receptor, EphA2
Reverse Transcriptase Polymerase Chain Reaction
Signal Transduction
Virus Internalization

Pub Type(s)

Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

22509030