Molecular variability, genetic relatedness and a novel open reading frame (pispo) of sweet potato-infecting potyviruses

Sweet potato feathery mottle virus (SPFMV, genus Potyvirus) infects sweet potato wherever it is cultivated. In single infections, SPFMV often causes only mild symptoms a situation that changes dramatically when it co-infects the plants with Sweet potato chlorotic stunt virus (SPCSV, genus Crinivirus). Co-infection generates the sweet potato virus disease (SPVD), the most devastating viral disease of sweet potato that can cause total loss of yield. Previous studies have described RNase3 as the SPCSV protein responsible for the occurrence of SPVD. However, attempts to find resistance to SPVD based on this knowledge have failed, suggesting possible SPFMV determinants involved in the development of SPVD. This study aimed to contribute to the molecular understanding of SPFMV, in particular regarding its phylogenetic relationships, genomic structure, and ability to suppress RNA interference (RNAi). Deployment of adequate control measures requires proper characterization of the pathogen. Phylogenetic relationship of SPFMV isolates and closely related potyviruses was ambiguous due to misidentification of viruses often found in mixed infections, and lack of specific methods of detection. In this study, we sequenced the complete genome of SPFMV strain C and found phylogenetic evidence to support its reclassification as a different species, Sweet potato virus C (SPVC). Furthermore, we demonstrated that Sweet potato feathery mottle virus and Sweet potato virus C together with Sweet potato virus G (SPVG) and Sweet potato virus 2 (SPV2) were found to cluster into one unique monophyletic subgroup among the potyviruses, the so-called SPFMV-group . Members of the SPFMV-group contain some unique genomic features. Based on pairwise comparisons of partial and complete genome sequences we found recombinant isolates in SPFMV and SPVC. We also identified, novel viral determinants characteristic for the SPFMV-group of potyviruses, mainly in the P1 cistron, a region known for its high variability among potyviruses. An additional domain at the N terminus of P1 (P1-N) which is not found in other potyvirus, but is found in sweet potato mild mottle virus (genus Ipomovirus) is invariably found in members of the SPFMV-group . The P1-N domain is followed by a hypervariable region which contains specific hallmarks for each member of the SPFMV-group , and is becoming a promising region for their rapid detection and characterization. However, perhaps the most remarkable finding was the identification of an extra open reading frame (ORF) overlapping the C-terminal part of P1, and which was designated as pispo (Pretty Interesting Sweet potato Potyvirus ORF). pispo is translated as a result of a transcriptional slippage mechanism occurring in the members of the SPFMV-group of potyviruses and results in a novel protein P1N-PISPO. Plants rely on RNAi, an antiviral defense machinery, to destroy viral ribonucleic acid (RNA) molecules recognized inside cells, and spread the alert signal to neighboring cells. To evade RNAi, viruses encode proteins termed suppressors. Our analyses revealed that both P1 and P1N-PISPO contain RNAi suppressor (RSS) activity. Hence, SPFMV utilizes novel suppressors expressed from the same P1 region in different reading frames. The protein P1N-PISPO suppresses cell-to-cell movement of silencing, probably by blocking the spread of signaling to neighboring cells. In contrast, P1 protein suppresses silencing only locally. In both cases, a conserved Glycine/Tryptophan (GW) motif located in the P1N part of P1 plays a crucial role in the RSS activity. On the other hand, HC-Pro, a widely known RSS protein of potyviruses was not able to suppress silencing in SPFMV. This particular arrangement, where the RSS activity resides on P1 region and not in HC-Pro, is not reported in potyviruses, but is known, e.g., in the members of the genus Ipomovirus of the Potyviridae family. Taxonomic, structural and functional findings of this study will contribute greatly to the understanding of the evolution of SPFMV, and characterization of diseases it causes.Bataatin höyhenlaikkavirus (SPFMV, suku Potyvirus) tartuttaa bataattia kaikkialla, missä bataattia viljellään. Yksinään SPFMV aiheuttaa useimmiten vain lieviä oireita, mutta jos kasveilla on samanaikaisesti bataatin kitukasvuvirus (SPCSV , suku Crinivirus) tartunta, tilanne muuttaa dramaattisesti. Kahden viruksen yhteistartunta johtaa ns. bataatin virustautiin (SPVD-tauti), joka on bataatin pahin virustauti, ja se voi johtaa jopa täydelliseen sadonmenetykseen. Aikaisemmat tutkimukset ovat osoittaneet, että SPCSV:n RNase3-proteiini yhdessä SPFMV:n kanssa on riittävä aiheuttumaan SPVD-taudin. Yritykset kehittää kestävyyttä bataatin SPVD-tautia vastaan tämän tiedon perusteella ovat kuitenkin epäonnistuneet, mikä voi johtua siitä, että myös SPFMV tarvitaan SPVD-taudin kehittymiseen. Tämän tutkimuksen tarkoituksena oli lisätä molekyylitason ymmärrystä SPFMV:sta ja erityisesti sen fylogeneettisistä sukulaissuhteista, genomirakenteesta ja kyvystä estää RNA-hiljennystä