RhoA-GTPase facilitates entry of Kaposi's sarcoma-associated herpesvirus into adherent target cells in a Src-dependent manner.
Kaposi's sarcoma-associated herpesvirus (KSHV) (human herpesvirus 8) binds to adherent target cell surface heparan sulfate molecules via its envelope glycoproteins gB and gpK8.1A, to integrins via gB, to the transporter CD98/xCT complex, and possibly to another molecule(s). This is followed by virus entry overlapping with the induction of preexisting host cell signal pathways, such as focal adhesion kinase, Src, phosphatidylinositol 3-kinase (PI3-K), Rho-GTPases, protein kinase C-zeta, and extracellular signal-regulated kinase 1/2. Here, using hemagglutinin-tagged plasmids expressing wild-type, dominant-positive, and dominant-negative forms of RhoA in HEK (human embryonic kidney) 293 cells, we investigated the role of RhoA-GTPase in virus entry. The dominant-negative form of RhoA GTPase and treatment of target cells with Clostridium difficile toxin B (CdTxB), a specific inactivator of Rho-GTPases, significantly blocked KSHV entry. KSHV infection induced closely similar levels of FAK and PI3-K in all three cell types. In contrast, very strong Src activation was observed in KSHV-infected dominant-positive RhoA cells compared to wild-type cells, and only moderate Src activation was seen in dominant-negative cells. Inhibition of Src activation by CdTxB and reduction of RhoA activation by Src inhibitors suggest that KSHV-induced Src is involved in RhoA activation, which in turn is involved in a feedback-sustained activation of Src. Since the decreased entry in RhoA dominant-negative cells may be due to inefficient signaling downstream of RhoA, we examined the induction of RhoA-activated Dia-2, which is also known to induce Src. Dia-2 coimmunoprecipitated with activated Src, which was inhibited by Src inhibitors, in the infected cells. Together with the reduced virus entry in RhoA dominant-negative cells, these results suggest that activated RhoA-dependent Dia-2 probably functions as a link between RhoA and Src in KSHV-infected cells, mediating the sustained Src activation, and that KSHV-induced Src and RhoA play roles in facilitating entry into adherent target cells.
Veettil
Mohanan Valiya
MV
Department of Microbiology and Immunology, Chicago Medical School, Rosalind Franklin University of Medicine and Science, 3333 Green Bay Road, North Chicago, IL 60064, USA.
Sharma-Walia
Neelam
N
Sadagopan
Sathish
S
Raghu
Hari
H
Sivakumar
Ramu
R
Naranatt
Pramod P
PP
Chandran
Bala
B
D002448:Cell Adhesion / Q000502:physiology*; D002460:Cell Line; D015967:Gene Expression Regulation, Viral; D019288:Herpesvirus 8, Human / Q000235:genetics / Q000502:physiology*; D006801:Humans; D015398:Signal Transduction; D020741:rho GTP-Binding Proteins / Q000502:physiology*; D019061:src-Family Kinases / Q000378:metabolism*
D019061:src-Family Kinases; D020741:rho GTP-Binding Proteins