scholarly journals The AU-Rich RNA Recombination Hot Spot Sequence of Brome Mosaic Virus Is Functional in Tombusviruses: Implications for the Mechanism of RNA Recombination

2004 ◽  
Vol 78 (5) ◽  
pp. 2288-2300 ◽  
Author(s):  
Natalia Shapka ◽  
Peter D. Nagy
Keyword(s):  
Mosaic Virus ◽  
Hot Spots ◽  
Hot Spot ◽  
Brome Mosaic Virus ◽  
Template Switching ◽  
Rna Recombination ◽  
Replicase Proteins ◽  
Recombination Hot Spots ◽  
Recombination Junction ◽  
Selection Of

ABSTRACT RNA recombination can be facilitated by recombination signals present in viral RNAs. Among such signals are short sequences with high AU contents that constitute recombination hot spots in Brome mosaic virus (BMV) and retroviruses. In this paper, we demonstrate that a defective interfering (DI) RNA, a model template associated with Tomato bushy stunt virus (TBSV), a tombusvirus, undergoes frequent recombination in plants and protoplast cells when it carries the AU-rich hot spot sequence from BMV. Similar to the situation with BMV, most of the recombination junction sites in the DI RNA recombinants were found within the AU-rich region. However, unlike BMV or retroviruses, where recombination usually occurred with precision between duplicated AU-rich sequences, the majority of TBSV DI RNA recombinants were imprecise. In addition, only one copy of the AU-rich sequence was essential to promote recombination in the DI RNA. The selection of junction sites was also influenced by a putative cis-acting element present in the DI RNA. We found that this RNA sequence bound to the TBSV replicase proteins more efficiently than did control nonviral sequences, suggesting that it might be involved in replicase “landing” during the template switching events. In summary, evidence is presented that a tombusvirus can use the recombination signal of BMV. This supports the idea that common AU-rich recombination signals might promote interviral recombination between unrelated viruses.

scholarly journals Silencing Homologous RNA Recombination Hot Spots with GC-Rich Sequences in Brome Mosaic Virus

1998 ◽  
Vol 72 (2) ◽  
pp. 1122-1130 ◽  
Author(s):  
Peter D. Nagy ◽  
Jozef J. Bujarski
Keyword(s):  
Homologous Recombination ◽  
Mosaic Virus ◽  
Hot Spots ◽  
Rna Virus ◽  
Hot Spot ◽  
Brome Mosaic Virus ◽  
Complex Nature ◽  
Template Switching ◽  
Rna Recombination ◽  
Recombination Hot Spots

ABSTRACT It has been observed that AU-rich sequences form homologous recombination hot spots in brome mosaic virus (BMV), a tripartite positive-stranded RNA virus of plants (P. D. Nagy and J. J. Bujarski, J. Virol. 71:3799–3810, 1997). To study the effect of GC-rich sequences on the recombination hot spots, we inserted 30-nucleotide-long GC-rich sequences downstream of AU-rich homologous recombination hot spot regions in parental BMV RNAs (RNA2 and RNA3). Although these insertions doubled the length of sequence identity in RNA2 and RNA3, the incidence of homologous RNA2 and RNA3 recombination was reduced markedly. Four different, both highly structured and nonstructured downstream GC-rich sequences had a similar “homologous recombination silencing” effect on the nearby hot spots. The GC-rich sequence-mediated recombination silencing mapped to RNA2, as it was observed when the GC-rich sequence was inserted at downstream locations in both RNA2 and RNA3 or only in the RNA2 component. On the contrary, when the downstream GC-rich sequence was present only in the RNA3 component, it increased the incidence of homologous recombination. In addition, upstream insertions of similar GC-rich sequences increased the incidence of homologous recombination within downstream hot spot regions. Overall, this study reveals the complex nature of homologous recombination in BMV, where sequences flanking the common hot spot regions affect recombination frequency. A replicase-driven template-switching model is presented to explain recombination silencing by GC-rich sequences.

scholarly journals RNA recombination in brome mosaic virus, a model plus strand RNA virus.

1998 ◽  
Vol 45 (4) ◽  
pp. 847-868 ◽  
Author(s):  
M Figlerowicz ◽  
J J Bujarski
Keyword(s):  
Mosaic Virus ◽  
Genetic Recombination ◽  
Rna Virus ◽  
Brome Mosaic Virus ◽  
Template Switching ◽  
Rna Recombination ◽  
Signal Sequences ◽  
Experimental Systems ◽  
Replicase Complex ◽  
Molecular Aspects

Studies on the molecular mechanism of genetic recombination in RNA viruses have progressed at the time when experimental systems of efficient recombination crossovers were established. The system of brome mosaic virus (BMV) represents one of the most useful and most advanced tools for investigation of the molecular aspects of the mechanism of RNA-RNA recombination events. By using engineered BMV RNA components, the occurrence of both homologous and nonhomologous crosses were demonstrated among the segments of the BMV RNA genome. Studies show that the two types of crossovers require different RNA signal sequences and that both types depend upon the participation of BMV replicase proteins. Mutations in the two BMV-encoded replicase polypeptides (proteins 1a and 2a) reveal that their different regions participate in homologous and in nonhomologous crossovers. Based on all these data, it is most likely that homologous and nonhomologous recombinant crosses do occur via two different types of template switching events (copy-choice mechanism) where viral replicase complex changes RNA templates during viral RNA replication at distinct signal sequences. In this review we discuss various aspects of the mechanism of RNA recombination in BMV and we emphasize future projections of this research.

scholarly journals Two types of non-homologous RNA recombination in brome mosaic virus.

2005 ◽  
Vol 52 (4) ◽  
pp. 833-844 ◽  
Author(s):  
Magdalena Alejska ◽  
Nelli Malinowska ◽  
Anna Urbanowicz ◽  
Marek Figlerowicz
Keyword(s):  
Mosaic Virus ◽  
Genetic Recombination ◽  
Genetic Material ◽  
Brome Mosaic Virus ◽  
Homologous Sequence ◽  
Template Switching ◽  
Rna Recombination ◽  
Genomic Rnas ◽  
Site Specific ◽  
Rna Interaction

Non-homologous RNA recombination is a process enabling the exchange of genetic material between various (related or unrelated) RNA-based viruses. Despite extensive investigations its molecular mechanism remains unclear. Studies on genetic recombination in brome mosaic virus (BMV) have shown that local hybridization between genomic RNAs induces frequent non-homologous crossovers. A detailed analysis of recombinant structures suggested that local complementary regions might be involved in two types of non-homologous recombination in BMV: site-specific and heteroduplex-mediated. To verify the above hypothesis and better recognize the mechanism of the phenomenon studied we have tested how the putative types of recombination are affected by a specific mutation in the BMV polymerase gene or by changes in RNA structure. The experiments undertaken revealed substantial differences between site-specific and heteroduplex-mediated recombination, indicating that they occur according to different mechanisms. The former can be classified as homology-assisted, and the latter as homology-independent. In addition to local RNA/RNA hybridization, short regions of homology are required for site-specific crossovers to occur. They are most efficiently mediated if one homologous sequence is located at the beginning of and the second just before a double-stranded region. At present it is difficult to state what is the mechanism of heteroduplex-mediated recombination. Earlier it was postulated that strong RNA/RNA interaction enforces template switching by the viral replicase. There are, however, several observations questioning this model and indicating that some other factors, which are still unknown, may influence heteroduplex-mediated crossovers.

scholarly journals Frequent Homologous Recombination Events between Molecules of One RNA Component in a Multipartite RNA Virus

2000 ◽  
Vol 74 (9) ◽  
pp. 4214-4219 ◽  
Author(s):  
A. Bruyere ◽  
M. Wantroba ◽  
S. Flasinski ◽  
A. Dzianott ◽  
J. J. Bujarski
Keyword(s):  
Homologous Recombination ◽  
Hot Spots ◽  
Rna Virus ◽  
Hot Spot ◽  
Rna Replication ◽  
Rna Recombination ◽  
Brome Mosaic Bromovirus ◽  
Local Lesion Host ◽  
Minor Sequence ◽  
Recombination Hot Spots

ABSTRACT Brome mosaic bromovirus (BMV), a tripartite plus-sense RNA virus, has been used as a model system to study homologous RNA recombination among molecules of the same RNA component. Pairs of BMV RNA3 variants carrying marker mutations at different locations were coinoculated on a local lesion host, and the progeny RNA3 in a large number of lesions was analyzed. The majority of doubly infected lesions accumulated the RNA3 recombinants. The distribution of the recombinant types was relatively even, indicating that both RNA3 counterparts could serve as donor or as acceptor molecules. The frequency of crossovers between one pair of RNA3 variants, which possessed closely located markers, was similar to that of another pair of RNA3 variants with more distant markers, suggesting the existence of an internal recombination hot spot. The majority of crossovers were precise, but some recombinants had minor sequence modifications, possibly marking the sites of imprecise homologous crossovers. Our results suggest discontinuous RNA replication, with the replicase changing among the homologous RNA templates and generating RNA diversity. This approach can be easily extended to other RNA viruses for identification of homologous recombination hot spots.

scholarly journals Foreign Complementary Sequences Facilitate Genetic RNA Recombination in Brome Mosaic Virus

Virology ◽  
1995 ◽  
Vol 208 (1) ◽  
pp. 370-375 ◽  
Author(s):  
Aleksandra Dzianott ◽  
Stanislaw Flasinski ◽  
Jozef J. Bujarski
Keyword(s):  
Mosaic Virus ◽  
Brome Mosaic Virus ◽  
Rna Recombination ◽  
Complementary Sequences
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