dual mechanism for the translation of subgenomic mrna from sindbis virus in infected and uninfected cells翻译的双重机制subgenomic信使rna从辛德毕斯病毒感染和未感染的细胞.pdfVIP

Dual Mechanism for the Translation of Subgenomic mRNA from Sindbis Virus in Infected and Uninfected Cells ´ ´ ´ ´ Miguel Angel Sanz, Alfredo Castello, Ivan Ventoso, Juan Jose Berlanga, Luis Carrasco* ´ ´ Centro de Biologıa Molecular ‘‘Severo Ochoa’’ (CSIC-UAM), Universidad Autonoma de Madrid, Cantoblanco, Madrid, Spain Abstract Infection of BHK cells by Sindbis virus (SV) gives rise to a profound inhibition of cellular protein synthesis, whereas translation of viral subgenomic mRNA that encodes viral structural proteins, continues for hours. To gain further knowledge on the mechanism by which this subgenomic mRNA is translated, the requirements for some initiation factors (eIFs) and for the presence of the initiator AUG were examined both in infected and in uninfected cells. To this end, BHK cells were transfected with different SV replicons or with in vitro made SV subgenomic mRNAs after inactivation of some eIFs. Specifically, eIF4G was cleaved by expression of the poliovirus 2A protease (2Apro) and the alpha subunit of eIF2 was inactivated by phosphorylation induced by arsenite treatment. Moreover, cellular location of these and other translation components was analyzed in BHK infected cells by confocal microscopy. Cleavage of eIF4G by poliovirus 2Apro does not hamper translation of subgenomic mRNA in SV infected cells, but bisection of this factor blocks subgenomic mRNA translation in uninfected cells or in cell-free systems. SV infection induces phosphorylation of eIF2a, a process that is increased by arsenite treatment. Under these conditions, translation of subgenomic mRNA occur