Molecular signatures of silencing suppression degeneracy from a complex RNA virus.

Silvia Ambrós,Javier Sánchez-Vicente,Guillermo Rodrigo,José-Antonio Daròs,Neus Gómez-Muñoz,Fernando Martínez,Josep Navarro-López,Silvia Giménez-Santamarina

doi:10.1371/journal.pcbi.1009166

Abstract

As genomic architectures become more complex, they begin to accumulate degenerate and redundant elements. However, analyses of the molecular mechanisms underlying these genetic architecture features remain scarce, especially in compact but sufficiently complex genomes. In the present study, we followed a proteomic approach together with a computational network analysis to reveal molecular signatures of protein function degeneracy from a plant virus (as virus-host protein-protein interactions). We employed affinity purification coupled to mass spectrometry to detect several host factors interacting with two proteins of Citrus tristeza virus (p20 and p25) that are known to function as RNA silencing suppressors, using an experimental system of transient expression in a model plant. The study was expanded by considering two different isolates of the virus, and some key interactions were confirmed by bimolecular fluorescence complementation assays. We found that p20 and p25 target a common set of plant proteins including chloroplastic proteins and translation factors. Moreover, we noted that even specific targets of each viral protein overlap in function. Notably, we identified argonaute proteins (key players in RNA silencing) as reliable targets of p20. Furthermore, we found that these viral proteins preferentially do not target hubs in the host protein interactome, but elements that can transfer information by bridging different parts of the interactome. Overall, our results demonstrate that two distinct proteins encoded in the same viral genome that overlap in function also overlap in their interactions with the cell proteome, thereby highlighting an overlooked connection from a degenerate viral system.

Highlights

Modern natural genomic architectures are the result of millions of years of evolution that have subjected them to a tinkering process with no rational a priori direction [1,2]
We began with the transient expression of the four viral proteins of interest in N. benthamiana plants using agroinfiltration (Fig 1C)
The tagged viral proteins and the host plant proteins that formed complexes with them were purified by affinity chromatography, and the resulting samples were analyzed by mass spectrometry to obtain lists of interacting proteins (Fig 1C)

Summary

Introduction

Modern natural genomic architectures are the result of millions of years of evolution that have subjected them to a tinkering process with no rational a priori direction [1,2]. Degeneracy is a genetic architecture feature that has been long assumed to go in parallel with complexity [3], so the occurrence of degeneracy remains elusive in minimal genomes, such as those of viruses. These genomes are highly constrained in size and evolve rapidly as a consequence of high mutation rates [13]. A plausible scenario would be that degeneracy contributed to achieve better adaption to new hosts by the virus This is because in such systems there is higher freedom to functionally diverge over time, and the probability of finding genetic solutions to efficiently perform in different environments increases, as previous theoretical work pointed out [6]. Without a precise characterization of the mechanistic mode of action of the different structural elements at play (e.g., through virus-host proteomics), it is difficult to derive any conclusion

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Discussion

Conclusion