RNA, bacteriophage

A number of bacterial viruses have RNA genomes. The best-known bacterial RNA viruses have single-stranded RNA. Interestingly, the bacterial RNA viruses known in the enteric bacteria group infect only bacterial cells which behave as gene donors (males) in genetic recombination. This restriction to male bacterial cells arises because these viruses infect bacteria by attaching to male-specific pili. Since such pili are absent on female cells, these RNA viruses are unable to attach to the females, and hence do not initiate infection in females. 

The bacterial RNA viruses are all of quite small size, about 26 nm in size, and they are all icosahedral, with 180 copies of coat protein per virus particle. The complete nucleotide sequence of several RNA phages are known. In the RNA phage MS2, which infects Escherichia coli, the viral RNA is 3,569 nucleotides long. The virus RNA, although single stranded, has extensive regions of secondary and tertiary structure. The RNA strand in the virion has the plus (+) sense, acting directly as mRNA upon entry into the cell. 

The genetic map is shown and the flow of events of MS2 multiplication. The infecting RNA goes to the host ribosome, where it is translated into four (or more) proteins. The four proteins that have been recognized are maturation protein (A-protein present in 

As noted, the viral RNA is of the plus (+) sense. Replicase synthesizes RNA of minus (-) sense using the infecting RNA as template. After minus RNA has been synthesized, plus RNA is made from this minus RNA. The newly made plus RNA strands now serve as messengers for virus protein synthesis. The gene for the maturation protein is at the 5 end of the RNA. Translation of the gene coding for the maturation protein (needed in only one copy per virus particle), occurs only from the newly formed plus-strand RNA as 

Ultimately, assembly occurs and release of virions from die cell occurs as a result of cell lysis. The features of replication of these simple RNA viruses are themselves fairly simple. The viral RNA itself functions as an mRNA and regulation occurs primarily by way of controlling access of ribosomes to the appropriate start sites on the viral RNA. 

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One of the most striking results that has emerged from the high-resolution crystallographic studies of these icosahedral viruses is that their coat proteins have the same basic core structure, that of a jelly roll barrel, which was discussed in Chapter 5. This is true of plant, insect, and mammalian viruses. In the case of the picornaviruses, VPl, VP2, and VP3 all have the same jelly roll structure as the subunits of satellite tobacco necrosis virus, tomato bushy stunt virus, and the other T = 3 plant viruses. Not every spherical virus has subunit structures of the jelly roll type. As we will see, the subunits of the RNA bacteriophage, MS2, and those of alphavirus cores have quite different structures, although they do form regular icosahedral shells. 

O Toole, J., Sinclair, M., Diaper, C., and Leder, K. (2008). Comparative survival of enteric indicators, E. coli and somatic and F-RNA bacteriophages on turf-grass. Water Sci. Technol. 58, 513-518. 

The RNA replicase of most RNA bacteriophages has a molecular weight of -210,000 and consists of four subunits. 

RNA-dependent RNA polymerases, such as the replicases of RNA bacteriophages, are template-specific for the viral RNA. 

The first successful attempts to study RNA evolution in vitro were carried out in the late sixties by Sol Spiegelman9 and his group at Columbia University (Spiegel-man, 1971). They made use of an RNA replicase isolated from Escherichia coli cells infected by the RNA bacteriophage QP and prepared a medium for replication by adding the four ribonucleoside triphosphates (GTP, ATP, CTP, and UTP) in a suitable buffer solution. QP RNA, when transferred into this medium, instantaneously started to replicate. Evolutionary experiments were carried out by means of the serial transfer technique (Figure 4). Materials consumed in RNA replication... 

Mindich, L. (1999). Precise packaging of the three genomic segments of the double-stranded-RNA bacteriophage 06. Microbiol. Mol. Biol. Rev. 63, 149-160. 

Valegard, K., Murray, J. B., Stockley, P. G., Stonehouse, N. J., and Liljas, L. (1994). Crystal structure of an RNA bacteriophage coat protein-operator complex. Nature 371, 623-626. 

Wolf, S. et al., Detection and characterization of F-i- RNA bacteriophages in water and shellfish Application of a multiplex real-time reverse transcription PCR, J. Virol Methods, 149, 123, 2008. 

If one examines the pattern of proteins synthesized in cells infected for three to four hours with a picomavirus, the only proteins synthesized are viral, for by this time the synthesis of most host proteins has ceased. In the earliest studies carried out by Summers et al. (I4) in poliovirus-infected HeLa cells, about I4 different virus-specified polypeptides were identified by SDS-polyacrylamide gel electrophoresis. The net mass of these viral polypeptides, however, exceeded the coding capacity of the viral genome by a factor of 2. This finding was incompatible with what was then known about the translation of the small RNA bacteriophages such as Q6 and R17 In these cases, each of the three cistrons was found to have its own initiation and termination signals (15) ... 

Infection of animal cells with a picomavirus results in the formation of an RNA dependent RNA polymerase (replicase) which is apparently responsible for the biosynthesis of the viral RNA genome. The discovery of the picomavirus replicase (l-3) and similar enzymatic activities in E, coli infected with RNA bacteriophages (4-6) were made about fifteen years ago. Today, there is detailed knowledge of the structure and properties of the replicases of bacteriophage Qg (7i 8) and f2 (9)j but only partial infoimation on the picomavirus replicase (10-14) This rather slow progress is due mainly to the difficulties encountered in the isolation of a stable ENA dependent replicase from a eukaryotic cell-vims system. 

The RNA replication complex of a picornavims is bound to smooth cytoplasmic membranes (10, 16). In order to obtain a soluble RNA dependent activity it is necessary to dissociate the enzyme from the membranes by means which adversely affect the activity of the enzyme. It is also more laborious and costly to grow and infect large quantities of animal cells than to carry out a large scale infection of E. coli with an RNA bacteriophage. So far, because of these limitations, the isolation of a picornavims replicase was carried out with very dilute solutions of enzyme-protein, a... 

Let us now consider the physics of catalytic interaction between individual polynucleotides. The most simple example for higher order catalytic action can be observed with primitive RNA bacteriophages. We consider the replication of Q0 in the host cell (Fig. 15). Plus and minus strands act as templates in replication. 

Hattman, S. Influence of T4 superinfection on the formation of RNA bacteriophage coat protein. J. molec. Biol. 47, 599-603 (1970). 

Yarosh, E., Levinthal, C. Exclusion of RNA bacteriophages and interference with their RNA replication by bacteriophage T4. J. molec. Biol. 30, 329-348 (1967). 

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