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Abstract Plant viruses have evolved diverse strategies for cell-to-cell and systemic movement, utilizing various viral proteins and cellular components and pathways. They typically encode one or a small group of proteins called movement proteins that mediate their local cell-to-cell movement via plasmodesmata (PD). Other virus-encoded proteins also make important contributions to viral transit through PD. Movement and other viral proteins use various cellular pathways to mediate their localization to and transit through PD. This review summarizes current understanding of these pathways and mechanisms, with a focus on movement proteins and their interactions with host factors. 

ABSTRACT Intercellular communication via plasmodesmata (PD) is essential for plant growth, development, and defense, yet its regulation remains poorly understood. Chloroplasts communicate information about the environment and the physiological state of the plant cell to the nucleus. It has been proposed that chloroplast-generated signals also modulate the expression of nuclear genes related to PD to regulate the trafficking of photosynthetic products and metabolites along with other molecules that act non-cell-autonomously. In this study, we set out to identify the chloroplast retrograde signals that regulate intercellular trafficking via PD. Using a combination ofArabidopsis thalianamutants and gene silencing inNicotiana benthamiana,we found that the metabolites of the tetrapyrrole biosynthetic pathway, most likely heme, can act to modulate PD-mediated intercellular trafficking. We also identified genes that are potentially regulated by the heme signal to modify plasmodesmata function. Together, these findings strengthen the link between chloroplasts and PD in coordinating intercellular communication for optimal plant development and resource allocation. 

Abstract Intercellular communication in plants, as in other multicellular organisms, allows cells in tissues to coordinate their responses for development and in response to environmental stimuli. Much of this communication is facilitated by plasmodesmata (PD), consisting of membranes and cytoplasm, that connect adjacent cells to each other. PD have long been viewed as passive conduits for the movement of a variety of metabolites and molecular cargoes, but this perception has been changing over the last two decades or so. Research from the last few years has revealed the importance of PD as signaling hubs and as crucial players in hormone signaling. The adoption of advanced biochemical approaches, molecular tools, and high-resolution imaging modalities has led to several recent breakthroughs in our understanding of the roles of PD, revealing the structural and regulatory complexity of these ‘protoplasmic connecting threads’. We highlight several of these findings that we think well illustrate the current understanding of PD as functioning at the nexus of plant physiology, development, and acclimation to the environment. 

Summary Plasmodesmata (PD) allow direct communication across the cellulosic plant cell wall, facilitating the intercellular movement of metabolites and signaling molecules within the symplast. InArabidopsis thalianaembryos with reduced levels of the chloroplast RNA helicase ISE2, intercellular trafficking and the number of branched PD were increased. We therefore investigated the relationship between alteredISE2expression and intercellular trafficking.Gene expression analyses in Arabidopsis tissues whereISE2expression was increased or decreased identified genes associated with the metabolism of glucosinolates (GLSs) as highly affected.Concomitant with changes in the expression of GLS‐related genes, plants with abnormalISE2expression contained altered GLS metabolic profiles compared with wild‐type (WT) counterparts. Indeed, changes in the expression of GLS‐associated genes led to altered intercellular trafficking in Arabidopsis leaves. Exogenous application of GLSs but not their breakdown products also resulted in altered intercellular trafficking.These changes in trafficking may be mediated by callose levels at PD as exogenous GLS treatment was sufficient to modulate plasmodesmal callose in WT plants. Furthermore, auxin metabolism was perturbed in plants with increased indole‐type GLS levels. These findings suggest that GLSs, which are themselves transported between cells via PD, can act on PD to regulate plasmodesmal trafficking capacity. 

Abstract Plant viruses rely on intercellular trafficking via plasmodesmata (PD) to move between cells in their hosts. This ability is conferred by virus encoded movement proteins (MPs), which can increase plasmodesmal permeability and intercellular trafficking independent of other viral proteins. Callose dynamics in the cell walls surrounding PD have a critical role in determining plasmodesmal flux, with decreased callose levels correlates with increased trafficking. Notably, PD callose levels are both increased and decreased during virus infection, suggesting that there are regulatory responses to the virus. Here, we found that auxin and brassinosteroid (BR) exert opposing effects on PD connectivity. While auxin enhances intercellular trafficking primarily by promoting PD density, BR restricts connectivity by reducing reduced PD biogenesis and increasing callose accumulation. We identified genes involved in auxin and BR signaling as Most of those genes encode membrane-associated proteins. We identified the receptor-like protein RLP15 as a critical upstream regulator of intracellular auxin homeostasis through stabilization of the ER-localized auxin transporter PILS5. In parallel, negative regulators of PD permeability including ERECTA, PPI, CER3, and DEAL2 define a host connectivity restraint network. These findings point to an auxin-BR module as a nexus for determining the degree of changes in plasmodesmal permeability that is elicited by the viral MP. Together with changes in callose dynamics at PD, this regulatory node allows plants to maintain homeostasis of intercellular trafficking, possibly contributing to maintenance of cell and tissue integrity during infection. Significance StatementPlant viruses encode movement proteins (MP) which increase plasmodesmal permeability to allow the local cell-to-cell trafficking of viral entities. This study identifies a non-canonical auxin-BR antagonistic module that regulates plasmodesmal connectivity in response to the changes triggered by viral movement protein early in the infection cycle. We demonstrate that MP30, encoded by the tobacco mosaic virus, rewires BR’s and auxin’s roles in controlling intercellular communication by interfering with membrane proteins associated with these hormonal pathways. By defining this hormonal nexus, our findings reveal a sophisticated host-pathogen interface where plants attempt to maintain intercellular homeostasis during the onset of viral pathogenesis. 

Summary Intercellular communication via plasmodesmata (PD) is essential for plant growth, development, and defense, yet its regulation remains poorly understood. We set out to identify the chloroplast retrograde signals that regulate intercellular trafficking via PD.Using a combination ofArabidopsis thalianamutants and gene silencing inNicotiana benthamiana, we found that the metabolites of the tetrapyrrole biosynthetic pathway, most likely heme, can act to modulate PD‐mediated intercellular trafficking. Knockout or knockdown ofGUN5, encoding the catalytic subunit of the magnesium chelatase, revealed a correlation between reduced levels of total heme and severely decreased intercellular trafficking.Further, plants with reduced expression of genes involved in chloroplast RNA processing were shown to have decreased total heme levels and decreased intercellular trafficking. Like withGUN5, reduced expression ofFC2, but notFC1, led to reduced levels of total heme and drastically reduced intercellular trafficking.Thus, the results support a model where a specific pool of heme regulates plasmodesmata and intercellular trafficking. We also identified genes that are potentially regulated by the heme signal to modify plasmodesmal function. Together, these findings strengthen the link between chloroplasts and PD in coordinating intercellular communication for optimal plant development and resource allocation. 

Abstract In plants, cytidine-to-uridine (C-to-U) editing is a crucial step in processing mitochondria- and chloroplast-encoded transcripts. This editing requires nuclear-encoded proteins including members of the pentatricopeptide (PPR) family, especially PLS-type proteins carrying the DYW domain.IPI1/emb175/PPR103is a nuclear gene encoding a PLS-type PPR protein essential for survival inArabidopsis thalianaand maize. Arabidopsis IPI1 was identified as likely interacting with ISE2, a chloroplast-localized RNA helicase associated with C-to-U RNA editing in Arabidopsis and maize. Notably, while the Arabidopsis andNicotianaIPI1 orthologs possess complete DYW motifs at their C-termini, the maize homolog, ZmPPR103, lacks this triplet of residues which are essential for editing. In this study we examined the function of IPI1 in chloroplast RNA processing inN. benthamianato gain insight into the importance of the DYW domain to the function of the EMB175/PPR103/ IPI1 proteins. Structural predictions suggest that evolutionary loss of residues identified as critical for catalyzing C-to-U editing in other members of this class of proteins, were likely to lead to reduced or absent editing activity in theNicotianaand Arabidopsis IPI1 orthologs. Virus-induced gene silencing ofNbIPI1led to defects in chloroplast ribosomal RNA processing and changes to stability ofrpl16transcripts, revealing conserved function with its maize ortholog.NbIPI1-silenced plants also had defective C-to-U RNA editing in several chloroplast transcripts, a contrast from the finding that maize PPR103 had no role in editing. The results indicate that in addition to its role in transcript stability, NbIPI1 may contribute to C-to-U editing inN. benthamianachloroplasts. 

Callose, a beta-(1,3)-D-glucan polymer, is essential for regulating intercellular trafficking via plasmodesmata (PD). Pathogens manipulate PD-localized proteins to enable intercellular trafficking by removing callose at PD, or conversely by increasing callose accumulation at PD to limit intercellular trafficking during infection. Plant defense hormones like salicylic acid regulate PD-localized proteins to control PD and intercellular trafficking during immune defense responses such as systemic acquired resistance. Measuring callose deposition at PD in plants has therefore emerged as a popular parameter for assessing likely intercellular trafficking activity during plant immunity. Despite the popularity of this metric there is no standard for how these measurements should be made. In this study, we compared three commonly used methods for identifying and quantifying PD callose by aniline blue staining were evaluated to determine the most effective in the Nicotiana benthamiana leaf model. The results reveal that the most reliable method used aniline blue staining and fluorescent microscopy to measure callose deposition in fixed tissue. Manual or semi-automated workflows for image analysis were also compared and found to produce similar results although the semi-automated workflow produced a wider distribution of data points.