Inhibition cellular protein synthesis

Highly potent bacterial toxins such as ricin and diphtheria can completely inhibit cellular protein synthesis at very low levels [26]. The bacterial toxin exerts cytotoxicity through enzymatic inactivation of factors essential for protein synthesis (e.g., riboso-mal RNA, elongation factor 2 or EF2). Inactivation of these proteins, which are... 

Denileukin Diftitox. Denileukin Diftitox (Ontak) is formulated by combining interleukin-2 with diphtheria toxin.11 Certain leukemia and lymphoma cells have a surface receptor that has a high affinity for interleukin-2, thus attracting this drug directly to these cells. Upon binding with tbe receptor, the diphtheria toxin component of the drug inhibits cellular protein synthesis, which ultimately results in cell death. This... 

Diphtheria is an acute illness caused by the toxin released by a Corynebacterium diphtheriae infection. The toxin inhibits cellular protein synthesis, with membranes forming on mucosal surfaces. Systemic toxemia can result in myocarditis, neuritis, and thrombocytopenia. Membrane formation can cause respiratory obstruction, and significant toxin absorption can lead to severe illness and death. 

I. Mechanism of toxicity. Amatoxins are highly stable and resistant to heat and are not removed by any form of cooking. They are thought to act by inhibiting cellular protein synthesis by interfering with RNA polymerase. 

Not surprisingly, the toxin s liver toxicity is reflected by the ability of micromolar concentrations of cylindrospermopsin to kill in vitro liver cells such as rat hepatocytes and the human hepatoblastoma cell line HEP-G2. Cylindrospermopsin is known to potently inhibit cellular protein synthesis that can be measured in vitro using rabbit reticulocyte lysate. 

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. 

The VSY genome codes for only five proteins (33), and soon after infection the five YSY proteins represent the only translation products of the infected cell, since this virus also inhibits cellular protein synthesis (34) The five YSY monocistronic mRNAs serving as templates for these proteins have been relatively well characterized (35) and thus this system lends itself to studies on the mechanism of superinfection. Ehrenfeld and Lund (36) examined the consequences of superinfecting YSY infected HeLA cells at two hours after infection with poliovirus. By I.5 to 2 hours after poliovirus superinfection, virtually all YSY polysomes disaggregated. The pattern of protein synthesis as shown on polyacrylamide gels changed from YSY specific to poliovirus specific within 2.5 hours after infection. All five of the YSY proteins were inhibited to the same extent. 

Another way in which picomaviruses could inhibit cellular protein synthesis would be to inactivate an initiation factor needed for cellular, but not viral, mENA translation. If this were the case one might expect to find a decreased capacity of extracts from infected cells to initiate translation of cellular mENAs compared to extracts from uninfected cells. Such studies have yielded a variety of results. In some laboratories no differences in activity were detected between extracts from xininfected cells and from EMC infected plasmacytoma cells or mengovirus infected Ehrlich ascites tumor cells (26, 44) In one laboratory the ability of extracts from infected cells to translate exogenously added encephalomyocarditis (EMC) virus ENA and total Krebs II ascites cell mENA was markedly diminished, but no evidence for the selective inhibition of translation of host cell mENA was obtained (52). 

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 idea that each virus uses its own strategy to inhibit cellular protein synthesis is further illustrated by herpes simplex virus (66) infection of Friend eirythroleukemia cells. In this case cellular mENA (globin) is degraded, in contrast to the infection of these same cells by YSY (66), in which no cellular mENA degradation is seen. 

Also, we do not yet know how viruses such as YSY, which have capped messengers, inhibit cellular protein synthesis. Whatever the strategy that YSY uses to turn off the host, poliovirus is apparently resistent to this step, since poliovirus grows in YSY infected cells. If YSY does inactivate an initiation factor other than eIF-4B, then this would suggest that poliovirus mENA translation by-passes the requirement for several cellular initiation factors. This finding emphasizes our lack of knowledge of the exact initiation factor requirement for viral and cellular protein synthesis in cell-free systems. 

Both of the above analyses were conducted on cells infected with the Bearing strain of reovirus, type 3. Sharpe and Fields (1982) showed that type 2 reovirus inhibits cellular protein synthesis more effectively than type 3. By isolating recombinant viruses containing various combinations of double-stranded RNA segments derived from both strains of reovirus, they demonstrated that the S4 RNA segment, which encodes the major outer capsid protein of the virion, is responsible for the ability of type 2 reovirus to inhibit L cell ma-cromolecular synthesis. Inactivation of type 2 reovirus by ultraviolet... 

Superinfection with poliovirus has a striking effect on VSV-in-fected HeLa-Ss cells the evidence in these experiments was quite convincing that preformed VSV mRNA was not translated following infection with poliovirus (Ehrenfeld and Lund, 1977 Trachsel et al., 1980). Of course, the effect of VSV on cellular protein synthesis could not be determined in this experiment because poliovirus itself drastically inhibits cellular protein synthesis. As mentioned above, the ascendency of the mRNAs of two viruses may also depend on the type of cell which is doubly infected with the two competing viruses (Otto and Lucas-Lenard, 1980). 

A number of investigators had reported in earlier studies that 5 -DI particles of VSV, in which one-half or more of the 3 end of the genome is deleted, were fully capable of inhibiting cellular protein synthesis (Baxt and Bablanian, 1976 >). It seems likely that this apparent effect of DI particles on protein synthesis inhibition is due to difficult-to-detect contamination with the helper B virions (standard infection virus). Schnitzlein et al. (1983) detected little or no inhibition of BHK cell protein synthesis by various types of highly purified 5 -DI particles, even at high multiplicities of infection. These DI particles fully retained capacity to inhibit replication of infectious B particles and, as will be discussed later, the 5 -DI particles did not affect the capacity of infectious B virions to inhibit cellular protein synthesis (Schnitzlein et al., 1983). It seems safe to conclude that 5 -DI particles of VSV contain no structural components or biological activity that will inhibit cellular protein synthesis. 

A variety of chemical and physical agents have been used in an attempt to dissect and identify the VSV function(s) that inhibit cellular protein synthesis. 

Use of the general protein synthesis inhibitor, cycloheximide, led to the hypothesis that de novo VSV protein synthesis is required for inhibiting cellular protein synthesis because, when cycloheximide was removed from infected cells, viral protein synthesis preceded suppression of cellular protein synthesis inhibition (Wertz and Young-ner, 1972). In our experience, experiments such as these are difficult to interpret because cycloheximide has many side effects, such as inhibition of cellular RNA synthesis (Week and Wagner, unpublished data). 

From the foregoing analyses of data it seems safe to assume that a newly synthesized VSV product is the principal inhibitor of protein synthesis in infected cells. The fact that VSV transcription is required to inhibit cellular protein synthesis (McAllister and Wagner, 1976 Marvaldi et al., 1977) limited the search to newly synthesized viral RNAs and proteins. Circumstantial evidence obtained from very early studies with interferon and other inhibitors appeared to rule out viral proteins as primary inhibitors of cellular protein synthesis (Yamazaki and Wagner, 1970 Wertz and Youngner, 1970 reviewed by Bablan-ian, 1975). Careful studies by UV inactivation of selected VSV transcripts appear to rule out the mRNAs for the M, G, and L proteins (Marvaldi et al., 1977 Dunigan and Lucas-Lenard, 1983). This would seem to leave the VSV leader RNA, the N protein mRNA, and/or the NS protein mRNA as the principal candidates for inhibitors of cellular protein synthesis, or possibly one or all of these transcripts... 

Such a profound inhibition of cellular protein synthesis is not an uncommon feature of virus infections of mammalian cells. Several lines of research have recently converged to demonstrate that adenoviruses, like certain other viruses that inhibit cellular protein synthesis, such as the picornaviruses poliomyelitis virus and encephal-omyocarditis virus, have evolved a mechanism to permit the selective translation of viral mRNA species. Although both adenoviruses and picornaviruses can efficiently redirect the translational machinery of their host cells, the actual molecular mechanisms employed appear to be quite distinct. 

The mechanism by which the S4 dsRNA segment inhibits cellular protein synthesis is not known. The a3 polypeptide binds strongly to double-stranded regions of RNA (Huismans and Joklik, 1976). The binding of o3 to double-stranded regions of rRNA, tRNA, or mRNA... 

It is important to know whether viral-specified proteins are involved in the inhibition of host protein synthesis and ts mutants have been used to try to answer this question. RNA t mutants did not inhibit cellular protein synthesis at the restrictive temperature (Atkins, 1976). Under these conditions, RNA / mutants of alphaviruses make neither viral encoded proteins nor viral RNAs (Keranen and Kaariainen, 1975 Hashimoto and Simizu, 1978). From these findings, we can conclude that initiation of viral RNA replication is necessary for inhibition of host protein synthesis and also that viral components introduced into cells by the infecting virions are not directly responsible for this inhibition. 

TRICHOTHECENE MYCOTOXINS. A family of structurally related poisonous substances produced by various species of fungi, especially Acremorium (Cephalosporium), Fusarium, Myrothe-cium, Stachybotrys, Trichderme, and Verticumonosporium. Tri-chothecene mycotoxins are toxic to humans because they inhibit cellular protein synthesis. Prominent examples are deoxynivalenol (sometimes referred to as vomitoxin because it induces vomiting), diacetoxyscirpenol, HT-2, nivalenol, and T-2. These five toxins gained some notoriety in the so-caUed yellow rain events in Southeast Asia because of allegations that they were associated with Soviet-inspired use of chemical weapons (CW). 

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