Abstract
BackgroundCucurbit chlorotic yellows virus (CCYV) is a bipartite cucurbit-infecting crinivirus within the family Closteroviridae. The crinivirus genome varies among genera. P4.9 is the first protein encoded by CCYV RNA2. P5, which is encoded by LIYV, is necessary for efficient viral infectivity in plants; however, it remains unknown whether CCYV P4.9 is involved in movement.FindingIn this study, we used green fluorescent protein (GFP) to examine the intracellular distribution of P4.9-GFP in plant cells, and observed fluorescence in the cytoplasm and nucleus. Transient expression of P4.9 was localized to the plasmodesmata. Co-infiltration of agrobacterium carrying binary plasmids of P4.9 and GFP facilitated GFP diffusion between cells. Besides P4.9 was able to spread by itself to neighboring cells, and co-localized with a marker specific to the endoplasmic reticulum, HDEL-mCherry, but not with the Golgi marker Man49-mCherry.ConclusionsTogether, these results demonstrate that CCYV P4.9 is involved in cell–cell movement.
Highlights
Plant viruses are small obligate intracellular parasites that live in the symplast of their hosts
Fluorescence signals were individually visualized using laser scanning confocal microscopy (LSCM); we found that P4.9 accumulated in punctate spots along the cell periphery (Fig. 1c), which is characteristic of PD localization
P4.9-green fluorescent protein (GFP) was co-expressed with the endoplasmic reticulum (ER) marker HDEL-mCherry in N. benthamiana leaves; P4.9 was co-localized with the ER marker (Fig. 2c)
Summary
Plant viruses are small obligate intracellular parasites that live in the symplast of their hosts. Cucurbit chlorotic yellows virus (CCYV) is a cucurbitinfecting crinivirus within the family Closteroviridae [3,4,5] Members of this family possess the largest, most complex single-stranded RNA (ssRNA) genomes, which vary in size from ca. Beet yellows virus (BYV) is a well-studied closterovirus that requires five proteins for cell–cell movement: P6, HSP70h, P64, CP, and CPm [8,9,10]. Citrus tristeza virus (CTV) proteins involved in viral assembly have functions in viral spread; these include HSP70h, P61, CP, and minor CP (CPm). A non-virion protein, P26, which is encoded by lettuce infectious yellows crinivirus, is PD-localized and essential for viral systemic infection [13]. To construct a P4.9 binary expression vector, we amplified its PCR product using primer pair pGDP4.9PstIF/ pGDP4.9BamHIR (Additional file 1: Table S1), digested a
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