Protein synthesis Shut Off

The data available suggests that poliovirus induces the synthesis of some inhibitor which causes the slow inactivation of initiation factor eIE-4B. While it appears that there may be a break-through in the problem of protein synthesis shut-off, many questions still remain unanswered. For example, the nature of the eIF-4B inhibitor is unknown. Is it a viral product, or is it a cellular factor induced by the virus Is the eIF-4B inactivated by proteolytic digestion or is it inactivated by modification ... 

ABREU, S. and LUCAS-LENAEU, J. Cellulax protein synthesis shut-off by mengovirus Translation of nonviral and viral mRNAs in extracts from uninfected and infected Ehrlich ascites tumor cells. J. Virol. (1976), 18, 184-192. 

Vaccinia virus infection causes a rapid and drastic inhibition of host cell protein synthesis (shut-off) (Kit and Dubbs, 1962 Shatkin, 1963 Salzman and Sebring, 1967 Holowczak and Joklik, 1967 Moss and Salzman, 1968 Metz and Esteban, 1972). This rapid inhibition is followed by the degradation of host polyribosomes (Joklik and Mer-igan, 1966) and the formation of viral polyribosomes (Metz et al., 1975). Shut-off can also occur under some restrictive conditions (for review, see Bablanian, 1975). 

About 1.5-2 hours after infection the first viral structural proteins can be detected in cells efficiently infected by SFV (see Kaariainen and S6-derlund, 1978). By 3-4 hours the synthesis of host cell proteins is shut off. The mechanism of host protein shutoff is not yet known, but studies by van Steeg (1982) suggest that it is the viral capsid protein itself which is responsible for the selective inhibition of host protein synthesis. The capsid protein seems to reduce the activity of the initiation factors elF-4B and CAP binding protein below levels necessary for the formation of the 80 S initiation complex from host mRNA. Translation of the viral 26 S RNA is, however, unaffected. More studies are clearly needed to substantiate this interesting possibility. 

Involvement of a viral structural protein in shut-off. A structural protein may be involved in the shut-off of cellular protein synthesis. This conclusion was derived from experiments by Steiner-Pryor and Cooper (14)> who observed that certain temperature sensitive mutants of poliovirus defective in the structmal proteins of the virus particle were mable to inhibit cellular protein synthesis at the non-permissive temperature. 

Figure 14-2. Regulation of cyclic AMP-dependent protein kinase A (PKA) by cyclic AMP. Activation of adenylate cyclase by binding of G( -GTP amplifies the signal by synthesis of many molecules of cyclic AMP. Cyclic AMP binding to PKA causes dissociation of the regulatory subunits from the catalytic subunits, which carry on the signal. Phosphodiesterase regulates the concentration of cyclic AMP by catalyzing its hydrolysis to AMP, which shuts off the signal.

The fate of HSV-1 infection can be either lytic or latent. During the lytic replication cycle, which occurs in mucosal or epithelial cells, the host cell protein synthesis is shut off by the tegument vhs protein immediately after infection. Subsequently, the viral capsid releases the viral DNA into the nucleus where it will circularize. 

How might chalone interfere with proper keratinization as well as with proliferation There are many conceivable ways in which this glycoprotein could shut off the synthesis of those enzymes required for DNA synthesis and cell division and at the same time (perhaps in a reciprocal manner) unblock the synthesis of those enzymes needed for synthesis of the keratohyalin protein. This would be the normal state. A lack of chalone would then lead to unblocking of DNA synthesis and blocking of formation of keratohyalin, i.e., psoriasis. 

Carrasco L, Smith AE (1976) Sodium ions and the shut-off of host ceU protein synthesis by picornaviruses. Nature 264 807-809... 

The protein kinase H RI (heme regulated eIF-2 kinase) was first identified in studies on the regulation of protein biosynthesis in erythroid cells. A decrease in the heme concentration in reticulocytes leads to inhibition of globin synthesis at the level of translation. This regulation mechanism ensures that only so much globin is produced as is heme available. If the level of heme drops, then HRI becomes activated. The activated HRI phosphorylates the eIF-2a subunit, which in turn shuts off protein biosynthesis (Fig. 1.48). The mechanism of regulation of HRI kinase by heme is not well understood. Heme binding sites have been identified on the N-terminus and the kinase domain of HRI. 

According to this model (see Fig. 4) two genes are involved in TAT synthesis one for the mRNA of TAT itself, and another for a labile, RNA or protein, repressor molecule which forms a reversible complex with TAT mRNA, and both prevents its translation and promotes its degradation. Steroids could increase the rate of TAT synthesis by one of two alternative, or complementary, mechanisms. They could work entirely at the transcriptional level by promoting the synthesis of TAT mRNA and shutting off the synthesis of the labile repressor RNA, or they could work entirely post-transcriptionally by inactivating the repressor and allowing TAT mRNA to accumulate (provided that it was al-... 

Although the effect of virus infection on cellular RNA and DNA synthesis will be considered, this review will be concerned primarily with the inhibition of cellular protein synthesis. Several reviews on virus-induced changes in the synthesis of macromolecules have been written in the past few years (1,2,5,4)5 and thus the reference list is selective rather than exhaustive. This review will consist of three parts the first will be concerned with the properties of the shut-off phenomenon. The second part will deal with theories that have been proposed to explain shut-off. 

It has been postulated that the 3 -poly(A) sequence on eukaryotic mRNAs mi t be involved in mRNA stability since it has been found that these sequences on mRNAs in HeLa cells (28) are successively degraded with increasing age of mRNA molecules. On this basis Koschei (29) examined whether infection of HeLA cells with poliovirus leads to a more rapid degradation of poly(A) sequences which in turn might explain the shut-off of cellular protein synthesis. No detectable differences were observed in the poly(A) sequences in cellular cytoplasmic RNA from uninfected cells and cells infected with poliovirus for 4 hours. Both amount and size of these sequences were identical. 

Apparently related to the problem of host cell protein synthesis inhibition by viruses is the phenomenon of viral interference. In some cases, superinfection of a cell already infected with one virus does not affect the yield of the original virus. In other cases, however, the superinfecting virus completely inhibits translation of the original viral mRRA. Such is the case for superinfection of vesicular stomatitis virus (VSV)-infected cells by poliovirus (32). The kinetics and general properties of the shut-off of VSY protein synthesis appear to be the same as the shut-off of cellular protein synthesis after poliovirus infection... 

Another theory to explain the shut-off, proposed by Cooper et al. (4Q) involves the synthesis of viral proteins which have an affinity for the small ribosomal subunit and the 5 end of viral mENA. The viral protein would repress the synthesis of cellular proteins by combining with the 4OS subunit, thereby blocking its link with host mENA. By also binding to the 5 end of viral ENA, the proteins would facilitate the attachment of viral ENA to the 40s subunit and increase the translation of viral ENA. Viral proteins have been found to co-sediment with the 4OS subunits of HeLa cells infected with poliovirus (49)> of Ehrlich ascites tumor cells infected with EMC virus (50) and of L-cells infected with mengovirus (5 ) poliovirus infected cells, the viral proteins co-sedimenting with ribosomes were identified as VPO, VP1 and VP3, all structural proteins (49) Both structural and non-structural proteins were found associated with ribosomes from EMC... 

The general consensus of researchers in the field is that ds RNA is not involved in shut-off, since its effect in extracts showed no specificity between viral and cellular protein synthesis, as would... 

THE ROLE OF INITIATION FACTORS IN THE SHUT-OFF OF PROTEIN SYNTHESIS... 

The shut-off of protein synthesis in virus-infected cells is a phenomenon that has attracted the attention of several investigators. In an attempt to provide an explanation at the molecular level for the mechanism of shut-off, studies were initially carried out with virus-infected cells. Since the results of this kind of studies are extensively discussed in the preceeding chapter of this book, we will concentrate on a particular aspect, namely the role of initiation factors on the virus-induced inhibition of host protein synthesis. 

Inhibition of protein synthesis in intact HeLa cells infected with EMC virus was observed at the same time that viral RNA synthesis reached its peak (Figure l). The shut-off of host protein synthesis, however, preceded this general inhibition of protein synthesis (see below). When cell extracts were prepared at different times after infection and tested for endogenous protein synthesis, a progressive loss of activity with time of infection was observed. A similar result was obtained in L-cells infected with EMC (Figure 2). 

Extracts prepared from infected cells may be a useful tool to investigate the mechanism of the shut-off of the host-cell protein synthesis. In the case of picornaviirus-infected cells, however, the cell extracts are relatively inactive in protein synthesis unless supplemented with initiation factors. This inhibition of protein synthesis in cell extracts is probably caused by dsENA of viral origin. 

Ribosomes phosphorylated vitro show no alteration in their translational activity (5, ) Trivial explanations such as the possibility that the ribosomes are dephosphorylated in the translation assay seem to have been eliminated. It is, of course, possible that current assay methods are inadequate to detect subtle changes caused by phosphorylation, but it is equally valid to draw the strai tforward conclusion that phosphorylation does not affect the activity of ribosomes. This is in accord with the fact that changes in the phosphorylation state of ribosomes vivo do not appear to correlate with alterations in the efficiency of the ribosomes in protein biosynthesis. The one possible exception is the phosphorylation of one of the small ribosomal subunit proteins (S2) that occurs after infection by vaccinia virus (8). The timing of the phosphorylation of S2 seems however to be related neither to the shut-off of host cell protein synthesis nor to the switch from early viral protein synthesis to late gene expression, and it remains to be proven whether the phosphorylation has any material effect on ribosome activity or specificity. 

There are two views currently in vogue to account for the shut-off. One proposes that as the translation of viral ENA in vitro is more resistant to high monovalent cation concentrations (particularly to added Na ) than is the translation of cellular mRNA, it might be the influx of Na into the infected cell (as a consequence of the membrane changes known to occur after viral infection) that suppresses host protein synthesis whilst viral protein synthesis is relatively unaffected or may even be enhanced (55> 54) It is generally the case that viral mRNA translation in infected cells is much more resistant to the inhibitory effects of additional extracellular NaCI (which causes inhibition of initiation) than is the translation of host cell mRNA either in infected or in uninfected cells (55) ... 

The other view, which is not necessarily in contradiction,stems from the observation that in a competitive situation the translation of EMC RNA outcompetes the translation of cellular mRNA in vitro under virtually all experimental conditions (56). The only exception is that the addition of extra eIF-4B tends to diminish the competitive advantage of the viral RNA (18). (According to the Basel group there is a requirement for eIF-4B for initiation on EMC RNA (17) Ihe conclusion which is the most consistent with all these observations is therefore that the optimiam ratio of eIF-4B to other factors may be lower for EMC ENA than for cellular mRNAs), The shut-off of host cell protein synthesis in this view is due to the competition by the viral RNA at the initiation step. The competitive effect might be enhanced if viral infection resulted in some inactivation or modification of eIP-4B, and some compelling evidence for this has recently been found (19)> althou the nature of the change in eIP-4B remains unknown. 

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