Plant genomes contain large numbers of cell surface leucine-rich repeat (LRR) and intracellular nucleotide binding (NB)-LRR immune receptors encoded by resistance ( R ) genes that recognize specific pathogen effectors and trigger resistance responses. The unregulated expression of NB-LRR genes can trigger autoimmunity in the absence of pathogen infection and inhibit plant growth. Despite the potential serious consequence on agricultural production, the mechanisms regulating R -gene expression are not well understood. We identified microRNA (miRNA) progenitor genes precursor transcripts, and two miRNAs [nta-miR6019 (22-nt) and nta-miR6020 (21-nt)] that guide cleavage of transcripts of the Toll and Interleukin-1 receptor-NB-LRR immune receptor N from tobacco that confers resistance to tobacco mosaic virus (TMV). We further showed that cleavage by nta-miR6019 triggers RNA-dependent RNA polymerase 6- and ribonuclease Dicer-like 4-dependent biogenesis of 21-nt secondary siRNAs “in phase” with the 22-nt miR6019 cleavage site. Furthermore, we found that processing of the 22-nt nta-miR6019 depended on an asymmetric bulge caused by mismatch in the nta-miR6019 precursor. Interestingly, coexpression of N with nta-miR6019 and nta-miR6020 resulted in attenuation of N -mediated resistance to TMV, indicating that these miRNAs have functional roles in NB-LRR regulation. Using a bioinformatics approach, we identified six additional 22-nt miRNA and two 21-nt miRNA families from three Solanaceae species—tobacco, tomato, and potato. We show that members of these miRNA families cleave transcripts of predicted functional R genes and trigger production of phased secondary 21-nt siRNAs. Our results demonstrate a conserved role for miRNAs and secondary siRNAs in NB-LRR/LRR immune receptor gene regulation and pathogen resistance in Solanaceae .

Significance: Chloroplasts are critical, dynamic organelles in plant cells responsible for photosynthesis and myriad other aspects of metabolism. In recent years, plant cell biologists have increasingly focused on the formation of thin, long extensions from plastids called “stromules.” Although stromules have been observed in all land plant species and cell types investigated, we do not know why these projections form or what they do. Here we demonstrate that stromules form in response to light-related redox signals inside the chloroplast. We then show that chloroplasts extracted from plant cells can make stromules independently. These discoveries suggest that stromules may be involved in transmitting signals from within the chloroplast to other subcellular compartments.

The coordinated redistribution of sugars from mature “source” leaves to developing “sink” leaves requires tight regulation of sugar transport between cells via plasmodesmata (PD). Although fundamental to plant physiology, the mechanisms that control PD transport and thereby support development of new leaves have remained elusive. From a forward genetic screen for altered PD transport, we discovered that the conserved eukaryotic glucose-TOR (TARGET OF RAPAMYCIN) metabolic signaling network restricts PD transport in leaves. Genetic approaches and chemical or physiological treatments to either promote or disrupt TOR activity demonstrate that glucose-activated TOR decreases PD transport in leaves. We further found that TOR is significantly more active in mature leaves photosynthesizing excess sugars than in young, growing leaves, and that this increase in TOR activity correlates with decreased rates of PD transport. We conclude that leaf cells regulate PD trafficking in response to changing carbohydrate availability monitored by the TOR pathway.

Recent studies suggest that intercellular transport via plasmodesmata (PD) is regulated by cellular redox state. Until now, this relationship has been unclear, as increased production of reactive oxygen species (ROS) has been associated with both increased and decreased intercellular transport via PD. Here, we show that silencing two genes that both increase transport via PD, INCREASED SIZE EXCLUSION LIMIT1 (ISE1) and ISE2, alters organelle redox state. Using redox-sensitive green fluorescent proteins targeted to the mitochondria or plastids, we show that, relative to wild-type leaves, plastids are more reduced in both ISE1- and ISE2-silenced leaves, whereas mitochondria are more oxidized in ISE1-silenced leaves. We further show that PD transport is positively regulated by ROS production in mitochondria following treatment with salicylhydroxamic acid but negatively regulated by an oxidative shift in both chloroplasts and mitochondria following treatment with paraquat. Thus, oxidative shifts in the mitochondrial redox state positively regulate intercellular transport in leaves, but oxidative shifts in the plastid redox state counteract this effect and negatively regulate intercellular transport. This proposed model reconciles previous contradictory evidence relating ROS production to PD transport and supports accumulating evidence that mitochondria and plastids are crucial regulators of PD function.

Target of rapamycin (TOR) is a protein kinase that coordinates eukaryotic metabolism. In mammals, TOR specifically promotes translation of ribosomal protein (RP) mRNAs when amino acids are available to support protein synthesis. The mechanisms controlling translation downstream from TOR remain contested, however, and are largely unexplored in plants. To define these mechanisms in plants, we globally profiled the plant TOR-regulated transcriptome, translatome, proteome, and phosphoproteome. We found that TOR regulates ribosome biogenesis in plants at multiple levels, but through mechanisms that do not directly depend on 5′ oligopyrimidine tract motifs (5′TOPs) found in mammalian RP mRNAs. We then show that the TOR-LARP1-5′TOP signaling axis is conserved in plants and regulates expression of a core set of eukaryotic 5′TOP mRNAs, as well as new, plant-specific 5′TOP mRNAs. Our study illuminates ancestral roles of the TOR-LARP1-5′TOP metabolic regulatory network and provides evolutionary context for ongoing debates about the molecular function of LARP1.