Vaccinia Virus Multiplication Relies On Rac-1/Jnk/Jun Pathway
Salgado, A.P.C.1,2*; Andrade, L.G. 1,2; Ferreira, P.C.P. 2; Kroon, E.G. 2 & Bonjardim, C.A. 1,2
1-Depto Microbiologia/Grupo de Transdução de Sinal, 2-Lab. de Vírus, Universidade Federal de Minas Gerais, CEP 31 270-901 -Belo Horizonte, MG. Telefone: 0XX31-3499 2752. E-mail: apsalgado@hotmail.com *
Vaccinia virus (VV) is a member of the Poxviridae family, a group of large, double-stranded DNA viruses that replicate in the cytoplasm of the host cells. Poxvirus infection induces extensive changes to both cell metabolism and structure. Its propagation is dependent on both virus-encoded and cellular protein kinases. We have shown that VV-stimulated pathway MEK/ERK1,2/ RSK2 leads to both phosphorylation of the ternary complex factor Elk1 and expression of the early growth response (egr-1) gene, which kinetically paralleled MAPK activation. The recruitment of this pathway is biologically relevant (ANDRADE et al., Biochem. J. v 381, 437, 2004). Our group has recently demonstrated that another pathway activated after VV infection, the c-jun N-terminal protein kinases (JNK1/2) is also required for successful VV replication (manuscript in preparation). In the attempt to connect the JNK/c-Jun cascade with upstream signals generated at cytoskeleton by tyrosine kinase after VV infection, we are now studying Rac-1, a small Rho family GTPase-protein associated with a variety of biological responses in reacting to extracellular signals. In this study, A31-derived cell clones stably-expressing dominant-negative Rac-1 (DN Rac-1) were selected and then infected with VV (MOI of 10) for 4h. Cell lysates were then prepared to western blot and immunoblotted with anti-phospho JNK .A significant reduction in both VV-induced- JNK and c-JUN- phosphorylation (downstream targets of Rac-1) was found. To confirm the role played by Rac-1 on VV biology, two DNs Rac-1 clones were infected with VV at MOI of 10 for times varying from 3, 6, 12, 24, 36 to 48 h. Virus was then collected and titrated by plaque assay. Our results are consistent with the requirement of Rac-1 for VV multiplication, since more than one log decrease in virus yield was detected in both DNs.
Financial support: CNPq, CAPES, FAPEMIG
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