Leaves turn yellow: small RNAs from viruses silence plant genes such as those involved in photosynthesis. Vitantonio Pantaleo IPSP-CNR

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1 Consiglio Nazionale delle Ricerche Di.S.Ba Conferenza del Dipartimento di Scienze Bio-agroalimentari del CNR Accademia dei Georgofili, Firenze Ottobre 2016 Accademia dei Georgofili Leaves turn yellow: small RNAs from viruses silence plant genes such as those involved in photosynthesis Vitantonio Pantaleo IPSP-CNR Institute for Sustainable Plant Protection - CNR AGRI-FOOD Department CNR Award Price 2011 for Plant and animal genomics FP7 EU project

2 RNA silencing In eukariotes RNA silencing serves as sequence-specific gene inactivation system; It acts at either post-transcriptional or transcriptional levels and it is at the base of many epigenetics phenomena; Key effectors are small non-coding RNAs (sirnas) incorporated into ribonuclease protein effectors such as Argonautes; RNA silencing has a pivotal role in development, differentiation, resilience to stresses; Virus induced RNA silencing In plants and insects it can function as defence mechanisms against infectious entities including invading viruses and endogenous retro viruses; As counter defence, viruses deliver viral silencing suppressors (VSR)

3 Virus induced RNA silencing Virion particle Delivery of protein and nucleic acids in cells RNAs of viral origin are recognised as invaders by plant RNase like nt long viral sirnas Activation of silencing: sirnas guide Argonaute for viral inactivation in a sequence specific manner AGOs AGO 1 ANTIVIRAL RNA silencing AGO Viral genome

4 Viral sympthoms Developmental anomalies are often associated with plant virus infections. Symptoms on virus-infected plants are often very specific to the given virus or to a single viral sequence. Il Cucumber mosaic virus (CMV) is a model viral system since it is associated to Satellite non coding RNAs, determinants of specific symptoms. STUNTING LEAF DEFORMATION Healty CMV + Sat Fny NECROSYS STUNTING CMV + necrogenic Sat CMV + necrogenic Sat mutated (1 nucleotide!!) by Cillo F, IPSP

5 from Masuta et al, 1992, Genetics The CMV yellowing Y-Sat CMV CMV+YSat Y-sat induces a bright yellowing of the natural host Nicotiana tabacum and other related species such as N. benthamiana (a model plant system) and other Solanaceae, such as Pepper but not in Tomato or Arabidopsis CMV CMV+YSat CMV CMV+YSat from Shimura et al, 2011, Plos Pathogens By functional analysis, yellowing determinants were identified in a 22nt long stretch of Y-Sat sequence (SYR) By Y-Sat genome editing we generated a SYR matching with the corresponding gene in Arabidopsis and Tomato and we could inducea specific yellowing. Thanks to the genomic data we could identify a plant mrna complemetary to SYR, the YR CMV+YSat (ara) CMV+YSat (tom) from Shimura et al, 2011, Plos Pathogens The Magnesium Chelatase, subuntit I (ChlI) is the host target. It is the key enzyme for chlorophyll biosynthesis

6 The Y-Sat - induced silencing of host ChlI A Viral Satellite RNA Induces Yellow Symptoms on Tobacco by Targeting a Gene Involved in Chlorophyll Biosynthesis using the RNA Silencing Machinery NGS analysis revealed Sat sirna from SYR Hanako Shimura 1, Vitantonio Pantaleo 2, Takeaki Ishihara 1, Nobutoshi Myojo 1, Jun-ichi Inaba 1, Kae Sueda 1,József Burgyán 2, Chikara Masuta 1 * 1 Research Faculty of Agriculture, Hokkaido University, Kita-ku, Sapporo, Japan, 2 Istituto di Virologia Vegetale, CNR, Torino, Italy OFF targets plant host gene Antiviral silencing cleaved ChlI mrna in vivo

7 The virus - induced yellowing: beyond the curiosity a biological significance having strategic impacts Nature Reviews Microbiology AOP, published online 6 June 2011; doi: /nrmicro2600 infected with Y-sat CMV. Both groups demonstrated that silencing of CHLI leads to the yellowing phenotype using different methods; Smith and colleagues transformed N. tabacum with an RNA interference vector, and Shimura and colleagues used virusinduced gene silencing mediated by a Yellow leaves attract preferentially aphids, the vectors of CMV and SatRNAs; Thus Y-Sat would increase the ecological fitness of CMV (spread); In tomato, CMV infection attract bumblebees by changing scent: it induces changes in profiles of mirnas that regulate emission of pollinatorperceivable volatiles attractive for bumblebee; Infected plants show the greatest increase in seed yield; Groen et al., Sept 2016 Plos Pathogens

8 One main conclusion from recent studies Plant viruses are infectious entities known for their pathogenic properties, but they should considered as plant symbionts in the classical view. They can regulate host gene expression transiently or trans-generationally to increase plant fitness such as: -enhancing endurance to drought or cold; -protecting plants from herbivores; -attracting pollinators. We expect a large use of this information in developing strategies for crops cultivation Thanks for your attention