A Three-Dimensional RNA Motif inPotato spindle tuber viroidMediates Trafficking from Palisade Mesophyll to Spongy Mesophyll inNicotiana benthamiana 

Abstract

Cell-to-cell trafficking of RNA is an emerging biological principle that integrates systemic gene regulation, viral infection, antiviral response, and cell-to-cell communication. A key mechanistic question is how an RNA is specifically selected for trafficking from one type of cell into another type. Here, we report the identification of an RNA motif in Potato spindle tuber viroid (PSTVd) required for trafficking from palisade mesophyll to spongy mesophyll in Nicotiana benthamiana leaves. This motif, called loop 6, has the sequence 59-CGA-39...59-GAC-39 flanked on both sides by cis Watson-Crick G/C and G/U wobble base pairs. We present a three-dimensional (3D) structural model of loop 6 that specifies all non-Watson-Crick base pair interactions, derived by isostericity-based sequence comparisons with 3D RNA motifs from the RNA x-ray crystal structure database. The model is supported by available chemical modification patterns, natural sequence conservation/variations in PSTVd isolates and related species, and functional characterization of all possible mutants for each of the loop 6 base pairs. Our findings and approaches have broad implications for studying the 3D RNA structural motifs mediating trafficking of diverse RNA species across specific cellular boundaries and for studying the structure-function relationships of RNA motifs in other biological processes.

tRNA-Related Sequences Trigger Systemic mRNA Transport in Plants

“…This finds support in studies on viroid RNA structures that are specifically necessary for entering specific vascular tissues (Takeda et al, 2011) and in the observed interaction of phloem-delivered mRNAs such as GAI and BEL5 with a phloem-expressed polypyrimidine-tract binding protein (PTB) recognizing poly-cysteine (C)-uracil (U) nucleotide stretches present in the 39 UTRs of some phloem mobile transcripts (Ham et al, 2009;Cho et al, 2015). However, a predicted RNA sequence related to PTB binding motifs was not found in the tRNA fusion constructs used in this study and thus is unlikely to play the primary role in triggering their movement.…”

“…Nonconserved viral 39 UTR sequences, which interact with 59 UTRs, seem to play a role in facilitating viral RNA cell-to-cell transfer . Similarly, viroids (infectious, non-protein-coding small RNAs) form specific stem-loop structures not yet identified in other mobile RNAs, allowing them to enter the plant phloem long-distance transport system (Ding, 2009;Takeda et al, 2011).…”

Long-distance movement of ArabidopsisFLOWERING LOCUS TRNA participates in systemic floral regulation

“…In Potato spindle tuber viroid (PSTVd), cell-to-cell trafficking of viroid RNA was found to be mediated by an RNA structure motif. 36 Whether the endogenous mobile RNA adopts similar structure motifs for long-distance trafficking remains to be investigated.…”

The Nucleolar Fibrillarin Protein Is Required for Helper Virus-Independent Long-Distance Trafficking of a Subviral Satellite RNA in Plants

“…However, remorin, a Solanaceae protein resident in membrane rafts and plasmodesmata, can interact physically with the MP from Potato virus X and negatively regulates Potato virus X intercellular and phloem-mediated trafficking (Raffaele et al, 2009). Some viral RNAs and viroids also contain important 3D RNA motifs required for intercellular movement (Takeda et al, 2011).…”