Selective Targeting of Mobile mRNAs to Plasmodesmata for Cell-to-Cell Movement

Abstract

Many plant mRNAs move from cell to cell or long distance to execute non-cell-autonomous functions. These mobile mRNAs traffic through the phloem to regulate many developmental processes, but despite the burgeoning discovery of mobile mRNAs, little is known about the mechanism underlying the intracellular sorting of these mRNAs. Here, we exploited a fluorescence-based mRNA labeling system, using the bacteriophage coat protein MS2, fused to GFP (MS2-GFP) and an MS2 recognition site in the RNA of interest, to visualize the intracellular trafficking of mobile mRNAs in living plant cells of Nicotiana benthamiana We first improved this system by using the nuclear localization sequence from FD, which substantially reduced the fluorescent background of MS2-GFP in the cytoplasm. The modified system allowed us to observe the cytoplasmic fluorescent foci dependent on MS2-binding sites. Coexpressing the MS2-GFP system with a virus movement protein, which is a plasmodesmata (PD)-localized nonspecific RNA-binding protein, targeted cytoplasmic fluorescent foci to the PD, suggesting that the cytoplasmic fluorescent foci contain mRNA and MS2-GFP. Our ex vivo RNA imaging revealed that mobile but not nonmobile mRNAs were selectively targeted to PD. Real-time images of intracellular translocation revealed that the translocation of mRNA and organelles in the transvacuolar strands may be governed by the same mechanism. Our study suggests that PD targeting of mRNA is a selective step in determining mRNA cell-to-cell movement of mRNAs.