Reovirus-induced modification of cap-dependent translation in infected L cells.

Abstract

The translational apparatus in cell-free extracts prepared from L cells infected with reovirus undergoes a time-dependent transition from cap dependence to cap independence. Extracts from uninfected L cells translate capped reovirus mRNA at high efficiency and synthesize the expected three size classes of reovirus polypeptides, and the translation is sensitive to m7G(5')ppp. This same extract translates uncapped mRNA at a much lower efficiency. In contrast, extracts from infected L cells translate uncapped reovirus mRNA at high efficiency and synthesize the correct three size classes of polypeptides, and the translation is not sensitive to inhibition by m7G(5')ppp. Infected cell extracts translate capped mRNA at reduced efficiency (z25%), the translation is not sensitive to inhibition by m7G(5')ppp, and the correct three size classes of viral polypeptides are not synthesized. These observations may explain how reovirus takes over the host translational apparatus.the viral mRNA in this process. The recent development of a method for making efficient cell-free protein-synthesizing systems of extremely low background activity from L cells (14) has enabled us to compare the in vitro translation of exogenously supplied reovirus mRNA in extracts that have been prepared from infected and uninfected L cells. Using extracts from infected cells, we found that the translation of uncapped reovirus mRNA is highly efficient and that the ability of these extracts to translate this form of reovirus mRNA increases with time of infection. Our observations lead us to conclude that the host cell translational machinery becomes modified as a result of viral infection. A model is proposed for the reovirus-induced takeover of the host cell translation machinery, in which uncapped viral mRNAs are preferentially translated at late times during infection.

The Regulation of Translation in Reovirus-infected Cells

“…Contrary to Skup and co-workers (Skup & Millward, 1980;Skup et al, 1981) and in agreement with Detjen et al (1982), the results obtained led us to consider that reovirus-infected HeLa cells are able to translate capped viral mRNAs late during virus infection. However, in reovirus-infected HeLa cells, no inhibition of cellular protein synthesis is apparent at the time when most viral proteins are synthesized, which suggests that there is an excess of translation components and that no competition between cellular and viral mRNAs is established in intact infected cells.…”

“…Contrary to this view, Skup & Millward (1980) have suggested that the inhibition of translation after reovirus infection of L cells occurs by a mechanism similar to that already suggested for poliovirus. The model proposes that the reovirus mRNA synthesized early in infection is capped, and the infected cells translate this kind of mRNA efficiently, but not uncapped mRNAs.…”

“…Both are thought to block protein synthesis by different mechanisms (Jen et al, 1980;Detjen et al, 1981). According to the model suggested for reovirus-infected L cells, the translation of reovirus uncapped m R N A s is blocked early during infection, and thus is only possible in the late phase (Skup & Millward, 1980;Skup et al, 1981). Fig.…”

“…The model proposes that the reovirus mRNA synthesized early in infection is capped, and the infected cells translate this kind of mRNA efficiently, but not uncapped mRNAs. Late in infection, the viral mRNA produced is uncapped and it is efficiently translated because the protein-synthesizing machinery has been modified in such a way that only uncapped mRNAs are recognized (Skup & Millward, 1980;Skup et al, 1981). However, the validity of the reovirus and poliovirus models has been questioned recently (Detjen et al, 1982;Alonso & Carrasco, 1982b).…”

“…According to the model proposed by Skup and co-workers (Skup & Millward, 1980;Skup et al, 1981) for regulation of translation in reovirus-infected cells, a modification in the proteinsynthesizing apparatus late in reovirus infection takes place, thus preventing the cell from translating capped mRNAs. To test this, reovirus-infected HeLa cells were superinfected late in infection with VSV, a virus that possesses typical capped m R N A , after which the synthesis of VSV proteins was analysed.…”