JIA-2018-09

2037 PAN Yuan et al. Journal of Integrative Agriculture 2018, 17(9): 2031–2041 tissues are involved in replication and assembly of virons. Plant viruses encode movement proteins (MP) that localize to the PD and modify the PD structure to facilitate cell to cell movement of virons or nucleic acids (Rodriguez et al . 2014). Recently, it has been reported that other virus proteins can also participate in virus intracellular and intercellular movement. For example, the tomato yellow leaf curl virus (TYLCV) C4 protein is able to interact with PD to modify their diameter and promote the intercellular movement of the viral DNA (Rojas et al . 2001). In addition, the potyviral P3N-PIPO and CI encoded protein can form a complex that coordinates the formation of PD-associated structures that can facilitate the intercellular movement (Wei et al . 2010). The cytoskeleton is also involved in transferring virus particles from replication sites to PD where they can be transported to adjacent cells. Ectopic expression of CaMV P6 in N . benthamiana leaves resulted in the formation of IBs that were capable of moving along actin microfilaments. When CaMV-infected Nicotiana edwardsonii leaves are treated with latrunculin B (Lat B), a chemical agent used to destabilize microfilaments, local lesion formation is abbrograted, suggesting that microfilament functions are required for CaMV infection (Harries et al . 2008). The 126-kD protein encoded by TMV also can form IBs that traffic along actin microfilaments. TMV IBs form in Lat B-treated cells, but the drug suppresses intracellular movement of IBs. Virus-induced gene silencing of actin and Lat B can Fig. 4 Fluorescence distribution of P6 mutants. A, schematic diagrams of P6-GFP and the P6 mutant derivatives. The blue and red regions in the top diagram represent the Strawberry vein banding virus (SVBV) P6 514 amino acid sequence. The single black lines identify deleted regions of the P6 mutants. The red regions designated a and b represent the nuclear location signals predicted by software. The purple region designated c represents the substituted SV40 nuclear location signal. The constructs were agroinfiltrated into Nicotiana benthamiana leaves and examined by fluorescence microscopy 3 days after infiltration. B, M1 inclusion body (IB) formation and nuclear accumulation. a, M1-GFP; b, bright field; c, overlay of a and b; d, M1-GFP; e, DAPI stained nucleus; f, overlay of d and e. C, restoration of M2NLS nuclear localization by insertion of the SV40 nuclear localization signal. a, M2-GFP; b, bright field; c, overlay of a and b; d, M5-GFP; e, bright field; f, overlay of d and e; g, M5-GFP; h, DAPI stained nucleus; I, overlay of g and h. D, disruption of M3 and M4 nuclear import. a, M3-GFP; b, bright field; c, overlay of a and b; d, M4-GFP; e, bright field; f, overlay of d and e. Magnification bar equals 20 μm. P6-GFP A B C D 193 223 a b c 402 426 514 GFP GFP GFP GFP YFP YFP 1 SV40-NLS M1-GFP M2-GFP M3-GFP M4-GFP M5-GFP a b c a b c d e f a b c d e f d e f g h i