DIR1 is a lipid transfer protein (LTP) postulated to complex with and/or chaperone a signal(s) to distant leaves during Systemic Acquired Resistance (SAR) in Arabidopsis. DIR1 was detected in phloem sap-enriched petiole exudates collected from wild-type leaves induced for SAR, suggesting that DIR1 gains access to the phloem for movement from the induced leaf. Occasionally the defective in induced resistance1 (dir1-1) mutant displayed a partially SAR-competent phenotype and a DIR1-sized band in protein gel blots was detected in dir1-1 exudates suggesting that a highly similar protein, DIR1-like (At5g48490), may contribute to SAR. Recombinant protein studies demonstrated that DIR1 polyclonal antibodies recognize DIR1 and DIR1-like. Homology modeling of DIR1-like using the DIR1-phospholipid crystal structure as template, provides clues as to why the dir1-1 mutant is rarely SAR-competent. The contribution of DIR1 and DIR1-like during SAR was examined using an Agrobacterium-mediated transient expression-SAR assay and an estrogen-inducible DIR1-EGFP/dir1-1 line. We provide evidence that upon SAR induction, DIR1 moves down the leaf petiole to distant leaves. Our data also suggests that DIR1-like displays a reduced capacity to move to distant leaves during SAR and this may explain why dir1-1 is occasionally SAR-competent.

BackgroundThe investigation of extremophile plant species growing in their natural environment offers certain advantages, chiefly that plants adapted to severe habitats have a repertoire of stress tolerance genes that are regulated to maximize plant performance under physiologically challenging conditions. Accordingly, transcriptome sequencing offers a powerful approach to address questions concerning the influence of natural habitat on the physiology of an organism. We used RNA sequencing of Eutrema salsugineum, an extremophile relative of Arabidopsis thaliana, to investigate the extent to which genetic variation and controlled versus natural environments contribute to differences between transcript profiles.ResultsUsing 10 million cDNA reads, we compared transcriptomes from two natural Eutrema accessions (originating from Yukon Territory, Canada and Shandong Province, China) grown under controlled conditions in cabinets and those from Yukon plants collected at a Yukon field site. We assessed the genetic heterogeneity between individuals using single-nucleotide polymorphisms (SNPs) and the expression patterns of 27,016 genes. Over 39,000 SNPs distinguish the Yukon from the Shandong accessions but only 4,475 SNPs differentiated transcriptomes of Yukon field plants from an inbred Yukon line. We found 2,989 genes that were differentially expressed between the three sample groups and multivariate statistical analyses showed that transcriptomes of individual plants from a Yukon field site were as reproducible as those from inbred plants grown under controlled conditions. Predicted functions based upon gene ontology classifications show that the transcriptomes of field plants were enriched by the differential expression of light- and stress-related genes, an observation consistent with the habitat where the plants were found.ConclusionOur expectation that comparative RNA-Seq analysis of transcriptomes from plants originating in natural habitats would be confounded by uncontrolled genetic and environmental factors was not borne out. Moreover, the transcriptome data shows little genetic variation between laboratory Yukon Eutrema plants and those found at a field site. Transcriptomes were reproducible and biological associations meaningful whether plants were grown in cabinets or found in the field. Thus RNA-Seq is a valuable approach to study native plants in natural environments and this technology can be exploited to discover new gene targets for improved crop performance under adverse conditions.

BackgroundThellungiella salsuginea is an important model plant due to its natural tolerance to abiotic stresses including salt, cold, and water deficits. Microarray and metabolite profiling have shown that Thellungiella undergoes stress-responsive changes in transcript and organic solute abundance when grown under controlled environmental conditions. However, few reports assess the capacity of plants to display stress-responsive traits in natural habitats where concurrent stresses are the norm.ResultsTo determine whether stress-responsive changes observed in cabinet-grown plants are recapitulated in the field, we analyzed leaf transcript and metabolic profiles of Thellungiella growing in its native Yukon habitat during two years of contrasting meteorological conditions. We found 673 genes showing differential expression between field and unstressed, chamber-grown plants. There were comparatively few overlaps between genes expressed under field and cabinet treatment-specific conditions. Only 20 of 99 drought-responsive genes were expressed both in the field during a year of low precipitation and in plants subjected to drought treatments in cabinets. There was also a general pattern of lower abundance among metabolites found in field plants relative to control or stress-treated plants in growth cabinets. Nutrient availability may explain some of the observed differences. For example, proline accumulated to high levels in cold and salt-stressed cabinet-grown plants but proline content was, by comparison, negligible in plants at a saline Yukon field site. We show that proline accumulated in a stress-responsive manner in Thellungiella plants salinized in growth cabinets and in salt-stressed seedlings when nitrogen was provided at 1.0 mM. In seedlings grown on 0.1 mM nitrogen medium, the proline content was low while carbohydrates increased. The relatively higher content of sugar-like compounds in field plants and seedlings on low nitrogen media suggests that Thellungiella shows metabolic plasticity in response to environmental stress and that resource availability can influence the expression of stress tolerance traits under field conditions.ConclusionComparisons between Thellungiella plants responding to stress in cabinets and in their natural habitats showed differences but also overlap between transcript and metabolite profiles. The traits in common offer potential targets for improving crops that must respond appropriately to multiple, concurrent stresses.

Methylation has frequently been implicated in gender determination in plants. The recent discovery of the sex determining region (SDR) of balsam poplar, Populus balsamifera, pinpointed 13 genes with differentiated X and Y copies. We tested these genes for differential methylation using whole methylome sequencing of xylem tissue of multiple individuals grown under field conditions in two common gardens. The only SDR gene to show a marked pattern of gender-specific methylation is PbRR9, a member of the two component response regulator (type-A) gene family, involved in cytokinin signalling. It is an ortholog of Arabidopsis genes ARR16 and ARR17. The strongest patterns of differential methylation (mostly male-biased) are found in the putative promoter and the first intron. The 4th intron is strongly methylated in both sexes and the 5th intron is unmethylated in both sexes. Using a statistical learning algorithm we find that it is possible accurately to assign trees to gender using genome-wide methylation patterns alone. The strongest predictor is the region coincident with PbRR9, showing that this gene stands out against all genes in the genome in having the strongest sex-specific methylation pattern. We propose the hypothesis that PbRR9 has a direct, epigenetically mediated, role in poplar sex determination.