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The Citrus leaf blotch virus movement protein acts as silencing suppressor

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Abstract

To counteract plant antiviral defense based on RNA silencing, many viruses express proteins that inhibit this mechanism at different levels. The genome of Citrus leaf blotch virus (CLBV) encodes a 227-kDa protein involved in replication, a 40-kDa movement protein (MP), and a 41-kDa coat protein (CP). To determine if any of these proteins might have RNA silencing suppressor activities, we have used Agrobacterium-mediated transient assays in the green fluorescent protein (GFP)-expressing Nicotiana benthamiana line 16c. Only CLBV MP was able to suppress intracellular GFP silencing induced by expression of either single- or double-stranded (ds) GFP RNA, but not cell-to-cell or long distance spread of the silencing signal. The MP suppressor activity was weak compared to other characterized viral suppressor proteins. Overall our data indicate that MP acts as a suppressor of local silencing probably by interfering in the silencing pathway downstream of the steps of dsRNA and small RNAs generation.

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Acknowledgments

Águeda Renovell was recipient of a doctoral fellowship from the Ministerio de Educación y Ciencia (MEC). Mª Carmen Vives was recipient of a contract from IVIA. This study was supported by grants AGL2006-0316 and AGL2009-08226, co-financed by FEDER funds, and by the MEC and the Ministerio de Ciencia e Innovación (MICINN). We thank Maria Boil for excellent lab assistance and José Juárez for excellent technical support to prepare photographs.

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  1. Centro de Protección Vegetal y Biotecnología, Instituto Valenciano de Investigaciones Agrarias, Ctra. Moncada-Náquera Km. 4.5, 46113, Moncada, Valencia, Spain

    Águeda Renovell, Mari Carmen Vives, Susana Ruiz-Ruiz, Luis Navarro, Pedro Moreno & José Guerri

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  1. Águeda Renovell
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  2. Mari Carmen Vives
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Correspondence to Mari Carmen Vives or José Guerri.

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11262_2011_674_MOESM1_ESM.tif

Fig. S1. Suppression by CLBV MP of RNA silencing triggered by dsGFP. a Leaves of N. benthamiana 16c plants infiltrated with a mixture of A. tumefaciens cultures harbouring binary vectors pBI-dsGFP and pBI-p19, pBI-MP or the empty vector (Ø). The green fluorescence images of the co-infiltrated leaves were taken at 5 days pos-infiltration (dpi) under a long-wavelength UV lamp. b Northern blot analysis of GFP mRNA and gfp-derived sRNAs extracted from the agro-infiltrated leaf patches shown in (a) or from non-infiltrated leaves (c) using a DIG-riboprobe specific for the GFP mRNA. Ethidium bromide staining of 5S rRNA and tRNAs are shown as loading controls for mRNA and sRNAs, respectively. Supplementary material 1 (TIFF 412 kb)

11262_2011_674_MOESM2_ESM.tif

Fig. S2. Effect of CLBV-encoded proteins on systemic spread of the GFP silencing signal in N. benthamiana 16c plants. Basal leaves were agro-infiltrated with pBI-GFP at the leaf tip and with pBI-Ø, pBI-MET, pBI-REP, pBI-MP, pBI-CP or pBI-p25 at the leaf base. Plants were photographed at 14 dpi under long-wavelength UV light with a yellow filter. Only the p25 protein of CTV prevented systemic silencing of the upper leaves. Supplementary material 2 (TIFF 1302 kb)

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Renovell, Á., Vives, M.C., Ruiz-Ruiz, S. et al. The Citrus leaf blotch virus movement protein acts as silencing suppressor. Virus Genes 44, 131–140 (2012). https://doi.org/10.1007/s11262-011-0674-9

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