Abstract
Plant virus movement proteins (MPs) localize to plasmodesmata (PD) to facilitate virus cell-to-cell movement. Numerous studies have suggested that MPs use a pathway either through the ER or through the plasma membrane (PM). Furthermore, recent studies reported that ER-PM contact sites and PM microdomains, which are subdomains found in the ER and PM, are involved in virus cell-to-cell movement. However, functional relationship of these subdomains in MP traffic to PD has not been described previously. We demonstrate here the intracellular trafficking of fig mosaic virus MP (MPFMV) using live cell imaging, focusing on its ER-directing signal peptide (SPFMV). Transiently expressed MPFMV was distributed predominantly in PD and patchy microdomains of the PM. Investigation of ER translocation efficiency revealed that SPFMV has quite low efficiency compared with SPs of well-characterized plant proteins, calreticulin and CLAVATA3. An MPFMV mutant lacking SPFMV localized exclusively to the PM microdomains, whereas SP chimeras, in which the SP of MPFMV was replaced by an SP of calreticulin or CLAVATA3, localized exclusively to the nodes of the ER, which was labeled with Arabidopsis synaptotagmin 1, a major component of ER-PM contact sites. From these results, we speculated that the low translocation efficiency of SPFMV contributes to the generation of ER-translocated and the microdomain-localized populations, both of which are necessary for PD localization. Consistent with this hypothesis, SP-deficient MPFMV became localized to PD when co-expressed with an SP chimera. Here we propose a new model for the intracellular trafficking of a viral MP. A substantial portion of MPFMV that fails to be translocated is transferred to the microdomains, whereas the remainder of MPFMV that is successfully translocated into the ER subsequently localizes to ER-PM contact sites and plays an important role in the entry of the microdomain-localized MPFMV into PD.
Highlights
Plasmodesmata (PD), channels providing symplastic continuity of the ER and the plasma membrane (PM) between adjacent cells, play vital roles in intercellular communication in plants [1]
Measurement of the fluorescent intensity across plasmodesma shows that the fluorescence signal of MP of FMV (MPFMV):YFP coincided with that of aniline blue (Fig 1Aii), showing that MPFMV:YFP localized to PD
PM and PD localization can be distinguished in plasmolyzed cells because PM proteins are associated with Hechtian strands, which are stretched PMs connecting the retracted PM and the cell wall, whereas PD proteins are retained in PD even during plasmolysis [24]
Summary
Plasmodesmata (PD), channels providing symplastic continuity of the ER and the plasma membrane (PM) between adjacent cells, play vital roles in intercellular communication in plants [1]. Understanding how MPs reach PD will provide insight into the mechanism underlying virus cell-to-cell movement. MPs have been frequently proposed to use a membrane trafficking pathway either through the ER or through the PM to reach PD. Several viruses possess MPs that reportedly use endomembrane trafficking through the ER. These MPs are apparently associated with the ER and PD in infected cells [5,6,7,8] or in cells transiently expressing only MPs [9], even though the detailed mechanism by which these MPs traffic from the ER to PD is unclear. One recent study has proposed that cauliflower mosaic virus MP is transported from the PM to PD through the endocytic pathway [12]
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