Poliovirus-directed protein synthesis

Further studies on VP4 isolated from poliovirus particles without SDS and with VP4 bound to viral RNA are expected to yield additional support for specifying a biological function of VP4. It will also be interesting to investigate the effect of VP4 on the initiation of poliovirus-directed protein synthesis in vitro. 

An alternative type of explanation for the specific discrimination against host cell protein synthesis in poliovirus-infected cells stemmed from the observation that initiation of protein synthesis could be selectively inhibited in HeLa cells and in poliovirus-infected HeLa cells by increasing the osmolarity of the growth medium (Sa-borio et al., 1974). The inhibition was independent of the solute used to increase the osmolarity. However, virus-directed protein synthesis was observed to be relatively more resistant to inhibition by hypertonic medium than was cellular protein synthesis, a fact which was interpreted as indicating that initiation of viral RNA translation was intrinsically more efficient than that of cellular mRNA (Nuss et al., 1975). These workers, therefore, proposed that the virus-specific or virus-induced factor involved in suppression of host protein synthesis could function by indiscriminantly lowering the rate of peptide chain initiation. Under such conditions, translation of viral mRNA, when it was synthesized, could occur due to its inherently strong affinity... 

One way of searching for the presence of inhibitors of polypeptide initiation in infected cells was to add cytoplasmic fractions from virus infected cells to a cell-free system from rabbit reticulocytes. This system initiates the synthesis of new polypeptide chains at a very high rate. Cytoplasm from poliovirus infected HeLa cells, but not from uninfected cells, inhibited protein synthesis in the reticulocyte lysate (59) The inhibitor was isolated and identified as double-stranded (ds) RNA (60). To study the effect of ds RNA on host and viral protein synthesis, a cell-free system from HeLa cells was developed which initiated translation on endogenous cellular or viral mRNA. When added to this system, the ds RNA was found to inhibit the translation of both cellular and viral mRNAs (61). Furthermore, measurement of the amount of ds RNA present in cells early in infection (61, 62) revealed that an insufficient quantity was present to act as a direct agent of protein synthesis inhibition. 

In this review I have outlined several theories that have been proposed to explain the mechanism by which picornaviruses inhibit cellular protein synthesis. Some theories seem less likely than others. Inhibition by ds ENA, for example, is no longer thought to be a likely possibility. In cell-free extracts ds ENA inhibits both cellular and viral mRNA translation (61). The inhibitor of cellular protein synthesis would be expected to be selective in its inhibitory activity. It is also apparent that picornavirus infection does not result in the degradation or alteration of cellular mENA (9> 27, 29 51). So,.too, experiments demonstrating that protein synthesis inhibition takes place in the absence of significant viral ENA synthesis (I4) tend to weaken the argument that protein synthesis inhibition results from direct competition of viral mENA with cellular ENA for initiation factor eIE-4D (47) As mentioned earlier, superinfection with poliovirus of cells infected with VSV prevents VSY mENA translation (J2, 56). In lysates from uninfected HeLa cells, however, 7SY mENA translation is favored over poliovirus mENA translation when both mENA species are present in equimolar saturating concentrations (55) If competition were a major cause of cellular protein synthesis inhibition, one would have expected poliovirus mENA to out-compete VSV mENA in cell-free translation, not the contrary. 

The suggestion that viral mRNA is an inherently efficient mRNA and can effectively compete with cellular mRNAs for some limiting component of the protein-synthesizing machinery has acquired experimental support for picornaviruses other than polioviruses. A direct competition model for shut-off of host cell translation has been proposed, specifically, to describe the shut-off induced by the car-dioviruses, EMC, and mengo (see Section 7, below). However, these models do not apply to poliovirus-induced inhibition because, as noted above, cessation of protein synthesis in poliovirus-infected cells occurs before detectable viral RNA is synthesized and shut-off does occur after infection in the presence of guanidine or with a mutant virus temperature-sensitive for RNA synthesis at restrictive temperature. 

A block at the level of initiation of protein synthesis has also been suggested as the mechanism of shut-off by vesicular stomatitis virus (VSV Stanners et al., 1977 Jaye et al., 1982 Gillies and Stollar, 1982). As in poliovirus-induced shut-off, degradation of host mRNAs does not seem to play a role in VSV-induced shut-off since host mRNA can be extracted from infected cells and translated in vitro (Ehrenfeld and Lund, 1977). However, recent work has demonstrated that the synthesis of VSV leader RNA is directly related to inhibition of host RNA synthesis (Grinnell and Wagner, 1983). Unlike poliovirus mRNAs, VSV mRNAs are capped and require cap-binding protein for translation (Banerjee, 1980 Rose et al., 1978). The mechanism of VSV-induced shut-off is presently under active investigation to determine if competition between mRNAs (Lodish and Porter, 1980,... 

To initiate a virus growth cycle, poliovirus RNA has to act first as mRNA. Since double-stranded RNA is not able to initiate protein synthesis directly (Miura and Muto, 1965), RF-RNA has either to be melted or to be used as a template for the synthesis of new RNA which in turn can serve as mRNA. Therefore the first steps of a virus growth cycle may differ according to whether infection is by viral RNA or RF-RNA. Whereas the initiation of a virus... 

---