Effect of viral infection on host protein synthesis and mRNA association with the cytoplasmic cytoskeletal structure.

Abstract

We studied the association of several eucaryotic viral and cellular mRNAs with cytoskeletal fractions derived from normal and virus-infected cells. We found that all mRNAs appear to associate with the cytoskeletal structure during protein synthesis, irrespective of their 5' and 3' terminal structures: e.g., poliovirus that lacks a 5' cap structure or reovirus and histone mRNAs that lack a 3' poly A tail associated with the cytoskeletal framework to the same extent as capped, polyadenylated actin mRNA. Cellular (actin) and viral (vesicular stomatitis virus and reovirus) mRNAs were released from the cytoskeletal framework and their translation was inhibited when cells were infected with poliovirus. In contrast, actin mRNA was not released from the cytoskeleton during vesicular stomatitis virus infection although actin synthesis was inhibited. In addition, several other conditions under which protein synthesis is inhibited did not result in the release of mRNAs from the cytoskeletal framework. We conclude that the association of mRNA with the cytoskeletal framework is required but is not sufficient for protein synthesis in eucaryotes. Furthermore, the shut-off of host protein synthesis during poliovirus infection and not vesicular stomatitis virus infection occurs by a unique mechanism that leads to the release of host mRNAs from the cytoskeleton.Initiation of translation in eucaryotes is far more complex than in procaryotes. At least eight initiation factor activities along with several auxiliary factors and ATP have been demonstrated to be involved in the initiation process in eucaryotes in vitro, as compared with just three initiation factors and an ATP-independent mechanism in procaryotes. It seems plausible that this complex complement of eucaryotic initiation factors might reflect an involvement in the more intricate patterns of regulation of translation that occur in the eucaryotic system (1).Besides the greater complexity of the factors mentioned above, the cellular architecture might also play a significant role in eucaryotic translation. There is evidence that eucaryotic mRNAs are translated only when associated with an elaborate network of filaments which extends throughout the cell and is referred to as "the cytoskeleton" (2). This fibrous network is composed of three distinct but interconnected filament systems, namely, the micro filaments, microtubules, and intermediate filaments, and other components. Wolosewick and Porter (3) have shown by high-voltage electron microscopy of intact cells that polysomes are clustered in the vicinity of the cytoskeleton structure. Subsequently, Lenk et al. (4) developed a procedure to fractionate cells into a detergent-resistant fraction which contains the cytoskeletal elements and a soluble fraction (containing soluble proteins, tRNA, monosomes, and other components) and found that polysomes invariably were associated with the cytoskeleton fraction. In further experiments, Cervera et al. (5) have shown that vesicular stomatitis virus (VSV) ~ mRNAs are transl...

Beta-actin mRNA localization is regulated by signal transduction mechanisms.

“…When fibroblasts with localized fl-actin mRNA were detergent extracted, a majority of this mRNA remained associated with the cytoskeleton (Fig. 6), consistent with earlier reports (Bonneau et al, 1985;Singer et al, 1989). When cells were treated with forskolin under conditions which markedly delocalized fl-actin mRNA, there was no change in the extractability of the mRNA relative to untreated cells (Fig.…”

Interaction between mRNA, ribosomes and the cytoskeleton

“…70 % of cellular mRNA has been recovered in a socalled 'cytoskeletal-bound' form either retained in the cell matrix or released from such structures by Triton X-1 00/deoxycholate treatment. This has been observed in a variety of cells such as HeLa cells (Lenk & Penman, 1979;van Venrooij et al, 1981), myoblasts (Bag & Praminik, 1987;Meadus et al, 1990), virusinfected cells (Lenk & Penman, 1979;Cervera et al, 1981;van Venrooij et al, 1981;Lemieux & Beaud, 1982;Bonneau et al, 1985;Katze et al, 1989) and oocytes (Moon et al, 1983;Jeffery, 1984;Yisraeli & Melton, 1988). However, as with the analysis of polysome distribution, the main limitation of these experiments has been the definition of the insoluble cell matrix as a cytoskeletal fraction and any mRNAs associated with it or with material released by Triton X-100/deoxycholate treatment as cytoskeletalbound.…”

“…277 where disruption of the cytoskeleton does not occur, the host mRNAs are retained on the cytoskeleton (Katze et al, 1989). There is also evidence to suggest that during viral infection there is retention of some host mRNAs, such as those for actin and tubulin, in the cell matrix fraction even though these are not translated (Lemieux & Beaud, 1982;Bonneau et al, 1985;Katze et al, 1989). It is likely that these two mRNAs are cytoskeletalbound even during viral infection without being actively translated.…”

Splicing enhances translation in mammalian cells: an additional function of the exon junction complex

“…Alternatively, before their removal, EJCs may target mRNPs to subcellular locations highly active in translation. In eukaryotic cells, most cytosolic ribosomes are known to be associated with the actin cytoskeleton, and active translation is highly dependent on the integrity of this structure (Bonneau et al 1985;Negrutskii et al 1994;Hovland et al 1996;Stapulionis et al 1997). The Y14 and Magoh requirement for proper localization of oskar mRNA in developing Drosophila embryos (Hachet and Ephrussi 2001;Mohr et al 2001) could reflect a broader role for these proteins in targeting spliced mRNAs to particular locations in the cytoskeleton.…”