The Importance of the KR-Rich Region of the Coat Protein of Ourmia melon virus for Host Specificity, Tissue Tropism, and Interference With Antiviral Defense

Abstract

The N-terminal region of the Ourmia melon virus (OuMV) coat protein (CP) contains a short lysine/arginine-rich (KR) region. By alanine scanning mutagenesis, we showed that the KR region influences pathogenicity and virulence of OuMV without altering viral particle assembly. A mutant, called OuMV 6710 , with three basic residue substitutions in the KR region, was impaired in the ability to maintain the initial systemic infection in Nicotiana benthamiana and to infect both cucumber and melon plants systemically. The integrity of this protein region was also crucial for encapsidation of viral genomic RNA; in fact, certain mutations within the KR region partially compromised the RNA encapsidation efficiency of the CP. In Arabidopsis thaliana Col-0, OuMV 6710 was impaired in particle accumulation; however, this phenotype was abolished in dcl2/dcl4 and dcl2/dcl3/dcl4 Arabidopsis mutants defective for antiviral silencing. Moreover, in contrast to CP wt , in situ immunolocalization experiments indicated that CP 6710 accumulates efficiently in the spongy mesophyll tissue of infected N. benthamiana and A. thaliana leaves but only occasionally infects palisade tissues. These results provided strong evidence of a crucial role for OuMV CP during viral infection and highlighted the relevance of the KR region in determining tissue tropism, host range, pathogenicity, and RNA affinity, which may be all correlated with a possible CP silencing-suppression activity.Ourmia melon virus (OuMV) is the best-characterized species of the Ourmiaviridae family, which also consists of Epirus cherry virus (EpCV) and Cassava virus C (CaVC) species (King et al. 2012). Three plus-strand RNA molecules form the OuMV genome and each segment encodes a single protein: RNA1 directs synthesis of an RNA-dependent RNA polymerase (RdRp), RNA2 encodes a movement protein (MP), and RNA3 specifies the 22-kDa coat protein (CP) (Rastgou et al. 2009). Ourmiaviruses have affinities with other postive single stranded RNA multipartite viruses, such as the members of the family Bromoviridae, but their unique structural and molecular features have resulted in their classification in a novel viral family. In fact, phylogenetic analyses showed that the members of the family likely went through a very unusual evolutionary process based on a possible reassortment between a plant virus and a mycovirus. In particular, OuMV RdRp shares distant similarity only with RdRp of narnaviruses, a group of viruses which mainly infects fungi; however, the MP is distantly related to MP of plant tombusviruses. The CP has limited but significant similarity with CP of plant and mammalian viruses, although its phylogenetic origin remains uncertain (Rastgou et al. 2009). Another peculiarity of the OuMV CP is the unique morphology of the virions among plant viruses: bacilliform particles of three different lengths with pointed ends (Lisa et al. 1988). Ectopic expression of RNA3 in Nicotiana benthamiana does not result in virion formation because the active replication of OuMV is necessary fo...

Identification of Diverse Mycoviruses through Metatranscriptomics Characterization of the Viromes of Five Major Fungal Plant Pathogens

“…It is not uncommon for deletions to occur in the CP coding regions of viruses that are normally vectored by Olpidium, Polymyxa, or Spongospora when they are repeatedly transmitted mechanically from plant to plant that result in loss of transmission by fungi or protists (83,84). In plant benyviruses and ourmiaviruses, entire CP-encoding genome segments can be lost as a result of maintenance in plants for extended periods of time (85)(86)(87). The retention of CP coding regions in genomes similar to those of ophioviruses and tobamoviruses suggests that these sequences are required for virus function.…”

The Torradovirus‐specific RNA2‐ORF1 protein is necessary for plant systemic infection

“…Sections were then processed as described in Rossi et al . (), using the polyclonal anti‐CP primary antibody (A405 II from the IPSP [Institute for Sustainable Plant Protection, Italy] collection). As negative controls, some sections were processed omitting either the primary or secondary antibody (not shown), or staining sections agroinfiltrated only with the R1 construct.…”

“…After washing with PBS, and embedding in 8% (w/v) low-melting agarose, 100lm-thick transverse sections were obtained with a vibrating blade microtome (Leica VT1000S, Leica Microsystems, Wetzlar, Germany). Sections were then processed as described in Rossi et al (2015), using the polyclonal anti-CP primary antibody (A405 II from the IPSP [Institute for Sustainable Plant Protection, Italy] collection). As negative controls, some sections were processed omitting either the primary or secondary antibody (not shown), or staining sections agroinfiltrated only with the R1 construct.…”

Transmission of Penicillium aurantiogriseum partiti‐like virus 1 to a new fungal host (Cryphonectria parasitica) confers higher resistance to salinity and reveals adaptive genomic changes

“…SDS-PAGE was carried out on the band obtained from sucrose gradient (on the negative control we took the same volume from the corresponding area in the sucrose density gradient) following standard protocol (Turina et al, 2000). From the same virus particles, we extracted RNA with a proteinase K treatment and Trizol extraction as previously detailed (Rossi et al, 2015). Northern blot was carried out as detailed above.…”