The plant cuticle is often considered a passive barrier from the environment. We show that the cuticle regulates active transport of the defense hormone salicylic acid (SA). SA, an important regulator of systemic acquired resistance (SAR), is preferentially transported from pathogen-infected to uninfected parts via the apoplast. Apoplastic accumulation of SA, which precedes its accumulation in the cytosol, is driven by the pH gradient and deprotonation of SA. In cuticle-defective mutants, increased transpiration and reduced water potential preferentially routes SA to cuticle wax rather than to the apoplast. This results in defective long-distance transport of SA, which in turn impairs distal accumulation of the SAR-inducer pipecolic acid. High humidity reduces transpiration to restore systemic SA transport and, thereby, SAR in cuticle-defective mutants. Together, our results demonstrate that long-distance mobility of SA is essential for SAR and that partitioning of SA between the symplast and cuticle is regulated by transpiration.

INCREASED SIZE EXCLUSION LIMIT2 (ISE2) is a chloroplast-localized RNA helicase that is indispensable for proper plant development. Chloroplasts in leaves with reduced ISE2 expression have previously been shown to exhibit reduced thylakoid contents and increased stromal volume, indicative of defective development. It has recently been reported that ISE2 is required for the splicing of group II introns from chloroplast transcripts. The current study extends these findings, and presents evidence for ISE2's role in multiple aspects of chloroplast RNA processing beyond group II intron splicing. Loss of ISE2 from Arabidopsis thaliana leaves resulted in defects in C-to-U RNA editing, altered accumulation of chloroplast transcripts and chloroplast-encoded proteins, and defective processing of chloroplast ribosomal RNAs. Potential ISE2 substrates were identified by RNA immunoprecipitation followed by next-generation sequencing (RIP-seq), and the diversity of RNA species identified supports ISE2's involvement in multiple aspects of chloroplast RNA metabolism. Comprehensive phylogenetic analyses revealed that ISE2 is a non-canonical Ski2-like RNA helicase that represents a separate sub-clade unique to green photosynthetic organisms, consistent with its function as an essential protein. Thus ISE2's evolutionary conservation may be explained by its numerous roles in regulating chloroplast gene expression.

OATAO is an open access repository that collects the work of Toulouse researchers and makes it freely available over the web where possible. This is an author-deposited version published in : http://oatao.univ-toulouse.fr/ Eprints ID : 19433To link to this article :

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Chloroplasts retain part of their ancestral genomes and the machinery for expression of those genomes. The nucleus-encoded chloroplast RNA helicase INCREASED SIZE EXCLUSION LIMIT2 (ISE2) is required for chloroplast ribosomal RNA processing and chloro-ribosome assembly. To further elucidate ISE2's role in chloroplast translation, two independent approaches were used to identify its potential protein partners. Both a yeast two-hybrid screen and a pull-down assay identified plastid ribosomal protein L15, uL15c (formerly RPL15), as interacting with ISE2. The interaction was confirmed in vivo by co-immunoprecipitation. Interestingly, we found that rpl15 null mutants do not complete embryogenesis, indicating that RPL15 is an essential gene for autotrophic growth of Arabidopsis thaliana. Arabidopsis and Nicotiana benthamiana plants with reduced expression of RPL15 developed chlorotic leaves, had reduced photosynthetic capacity and exhibited defective chloroplast development. Processing of chloroplast ribosomal RNAs and assembly of ribosomal subunits were disrupted by reduced expression of RPL15. Chloroplast translation was also decreased, reducing accumulation of chloroplast-encoded proteins, in such plants compared to wild-type plants. Notably, knockdown of RPL15 expression increased intercellular trafficking, a phenotype also observed in plants with reduced ISE2 expression. This finding provides further evidence for chloroplast function in modulating intercellular trafficking via plasmodesmata.

The signalling pathways that regulate intercellular trafficking via plasmodesmata (PD) remain largely unknown. Analyses of mutants with defects in intercellular trafficking led to the hypothesis that chloroplasts are important for controlling PD, probably by retrograde signalling to the nucleus to regulate expression of genes that influence PD formation and function, an idea encapsulated in the organelle-nucleus-PD signalling (ONPS) hypothesis. ONPS is supported by findings that point to chloroplast redox state as also modulating PD. Here, we have attempted to further elucidate details of ONPS. Through reverse genetics, expression of select nucleus-encoded genes with known or predicted roles in chloroplast gene expression was knocked down, and the effects on intercellular trafficking were then assessed. Silencing most genes resulted in chlorosis, and the expression of several photosynthesis and tetrapyrrole biosynthesis associated nuclear genes was repressed in all silenced plants. PD-mediated intercellular trafficking was changed in the silenced plants, consistent with predictions of the ONPS hypothesis. One striking observation, best exemplified by silencing the PNPase homologues, was that the degree of chlorosis of silenced leaves was not correlated with the capacity for intercellular trafficking. Finally, we measured the distribution of PD in silenced leaves and found that intercellular trafficking was positively correlated with the numbers of PD. Together, these results not only provide further support for ONPS but also point to a genetic mechanism for PD formation, clarifying a longstanding question about PD and intercellular trafficking. This article is part of the theme issue ‘Retrograde signalling from endosymbiotic organelles'.

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