Potyvirus-induced gene silencing: the dynamic process of systemic silencing and silencing suppression.
Potato virus A (PVA; genus Potyvirus) was used for virus-induced gene silencing in a model system that included transgenic Nicotiana benthamiana (line 16c) expressing the gfp transgene for green fluorescent protein (GFP) and chimeric PVA (PVA-GFP) carrying gfp in the P1-encoding region. Infection of the 16c plants with PVA-GFP in five experiments resulted in a reproducible pattern of systemic gfp transgene silencing, despite the presence of the strong silencing-suppressor protein, HC-Pro, produced by the virus. PVA-GFP was also targeted by silencing, and virus-specific short interfering RNA accumulated from the length of the viral genome. Viral deletion mutants lacking the gfp insert appeared in systemically infected leaves and reversed silencing of the gfp transgene in limited areas. However, systemic gfp silencing continued in newly emerging leaves in the absence of the gfp-carrying virus, which implicated a systemic silencing signal that moved from lower leaves without interference by HC-Pro. Use of GFP as a visual marker revealed a novel, mosaic-like recovery phenotype in the top leaves. The leaf areas appearing red or purple under UV light (no GFP expression) contained little PVA and gfp mRNA, and corresponded to the dark-green islands observed under visible light. The surrounding green fluorescent tissues contained actively replicating viral deletion mutants that suppressed GFP silencing. Taken together, systemic progression of gene silencing and antiviral defence (RNA silencing) and circumvention of the silencing by the virus could be visualized and analysed in a novel manner.
Gammelgård
Elin
E
Department of Plant Biology and Forest Genetics, Swedish University of Agricultural Sciences (SLU), PO Box 7080, SE-750 07 Uppsala, Sweden.
Mohan
Maradumane
M
Department of Plant Biology and Forest Genetics, Swedish University of Agricultural Sciences (SLU), PO Box 7080, SE-750 07 Uppsala, Sweden.
Valkonen
Jari P T
JPT
Department of Applied Biology, PO Box 27, FIN-00014 University of Helsinki, Finland.|Department of Plant Biology and Forest Genetics, Swedish University of Agricultural Sciences (SLU), PO Box 7080, SE-750 07 Uppsala, Sweden.
D003546:Cysteine Endopeptidases / Q000235:genetics / Q000378:metabolism; D020868:Gene Silencing; D018390:Gene Targeting; D049452:Green Fluorescent Proteins / Q000235:genetics; D010935:Plant Diseases / Q000821:virology*; D018515:Plant Leaves / Q000821:virology; D030821:Plants, Genetically Modified / Q000235:genetics* / Q000821:virology; D017800:Potyvirus / Q000235:genetics* / Q000378:metabolism; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D016865:Reassortant Viruses; D014026:Nicotiana / Q000821:virology*; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism
D034741:RNA, Small Interfering; D014764:Viral Proteins; D049452:Green Fluorescent Proteins; D003546:Cysteine Endopeptidases; C072775:HC-Pro protein, potyvirus