Arsenite provides a selective signal that coordinates arsenate uptake and detoxification through the regulation of PHR1 stability in Arabidopsis.

In nature, plants acquire nutrients from soils to sustain growth, and at the same time, they need to avoid the uptake of toxic compounds and/or possess tolerance systems to cope with them. This is particularly challenging when the toxic compound and the nutrient are chemically similar, as in the case of phosphate and arsenate. In this study, we demonstrated that regulatory elements of the phosphate starvation response (PSR) coordinate the arsenate detoxification machinery in the cell. We showed that arsenate repression of the phosphate transporter PHT1;1 is associated with the degradation of the PSR master regulator PHR1. Once arsenic is sequestered into the vacuole, PHR1 stability is restored and PHT1;1 expression is recovered. Furthermore, we identified an arsenite responsive SKP1-like protein and a PHR1 interactor F-box (PHIF1) as constituents of the SCF complex responsible for PHR1 degradation.We found that arsenite, the form to which arsenate is reduced for compartmentalization in vacuoles, represses PHT1;1 expression, providing a highly selective signal versus phosphate to control PHT1;1 expression in response to arsenate. Collectively, our results provide molecular insights into a sensing mechanism that regulates arsenate/phosphate uptake depending on the plant's detoxification capacity.

Targeted expression of the arsenate reductase HAC1 identifies cell type specificity of arsenic metabolism and transport in plant roots.

High Arsenic Concentration 1 (HAC1), an Arabidopsis thaliana arsenate reductase, plays a key role in arsenate [As(V)] tolerance. Through conversion of As(V) to arsenite [As(III)], HAC1 enables As(III) export from roots, and restricts translocation of As(V) to shoots. To probe the ability of different root tissues to detoxify As(III) produced by HAC1, we generated A. thaliana lines expressing HAC1 in different cell types. We investigated the As(V) tolerance phenotypes: root growth, As(III) efflux, As translocation, and As chemical speciation. We showed that HAC1 can function in the outer tissues of the root (epidermis, cortex, and endodermis) to confer As(V) tolerance, As(III) efflux, and limit As accumulation in shoots. HAC1 is less effective in the stele at conferring As(V) tolerance phenotypes. The exception is HAC1 activity in the protoxylem, which we found to be sufficient to restrict As translocation, but not to confer As(V) tolerance. In conclusion, we describe cell type-specific functions of HAC1 that spatially separate the control of As(V) tolerance and As translocation. Further, we identify a key function of protoxylem cells in As(V) translocation, consistent with the model where endodermal passage cells, above protoxylem pericycle cells, form a 'funnel' loading nutrients and potentially toxic elements into the vasculature.

A Genetic Dissection of Natural Variation for Stomatal Abundance Traits in Arabidopsis.

Stomatal abundance varies widely across natural populations of Arabidopsis thaliana, and presumably affects plant performance because it influences water and CO2 exchange with the atmosphere and thence photosynthesis and transpiration. In order to determine the genetic basis of this natural variation, we have analyzed a recombinant inbred line (RIL) population derived from the wild accession Ll-0 and the reference strain Landsberg erecta (Ler), which show low and high stomatal abundance, respectively. Quantitative trait locus (QTL) analyses of stomatal index, stomatal density, and pavement cell density measured in the adaxial cotyledon epidermis, identified five loci. Three of the genomic regions affect all traits and were named MID (Modulator of Cell Index and Density) 1 to 3. MID2 is a large-effect QTL overlapping with ERECTA (ER), the er-1 allele from Ler increasing all trait values. Additional analyses of natural and induced loss-of-function er mutations in different genetic backgrounds revealed that ER dysfunctions have differential and opposite effects on the stomatal index in adaxial and abaxial cotyledon epidermis and confirmed that ER is the gene underlying MID2. Ll-0 alleles at MID1 and MID3 displayed moderate and positive effects on the various traits. Furthermore, detailed developmental studies tracking primary and satellite stomatal lineages show that MID3-Ll-0 allele promotes the spacing divisions that initiate satellite lineages, while the ER allele limits them. Finally, expression analyses suggest that ER and MID3 modulate satellization through partly different regulatory pathways. Our characterization of MID3 indicates that genetic modulation of satellization contributes to the variation for stomatal abundance in natural populations, and subsequently that this trait might be involved in plant adaptation.

A Polynucleotide Repeat Expansion Causing Temperature-Sensitivity Persists in Wild Irish Accessions of Arabidopsis thaliana.

Triplet repeat expansions underlie several human genetic diseases such as Huntington's disease and Friedreich's ataxia. Although such mutations are primarily known from humans, a triplet expansion associated genetic defect has also been reported at the IIL1 locus in the Bur-0 accession of the model plant Arabidopsis thaliana. The IIL1 triplet expansion is an example of cryptic genetic variation as its phenotypic effects are seen only under genetic or environmental perturbation, with high temperatures resulting in a growth defect. Here we demonstrate that the IIL1 triplet expansion associated growth defect is not a general stress response and is specific to particular environmental perturbations. We also confirm and map genetic modifiers that suppress the effect of IIL1 triplet repeat expansion. By collecting and analyzing accessions from the island of Ireland, we recover the repeat expansion in wild populations suggesting that the repeat expansion has persisted at least 60 years in Ireland. Through genome-wide genotyping, we show that the repeat expansion is present in diverse Irish populations. Our findings indicate that even deleterious alleles can persist in populations if their effect is conditional. Our study demonstrates that analysis of groups of wild populations is a powerful tool for understanding the dynamics of cryptic genetic variation.

Natural variation involving deletion alleles of FRIGIDA modulate temperature-sensitive flowering responses in Arabidopsis thaliana.

Ambient temperature is one of the major environmental factors that modulate plant growth and development. There is extensive natural genetic variation in thermal responses of plants exemplified by the variation exhibited by the accessions of Arabidopsis thaliana. In this work we have studied the enhanced temperature response in hypocotyl elongation and flowering shown by the Tsu-0 accession in long days. Genetic mapping in the Col-0 × Tsu-0 recombinant inbred line (RIL) population identified several QTLs for thermal response including three major effect loci encompassing candidate genes FRIGIDA (FRI), FLOWERING LOCUS C (FLC) and FLOWERING LOCUS T (FT). We confirm and validate these QTLs. We show that the Tsu-0 FRI allele, which is the same as FRI-Ler is associated with late flowering but only at lower temperatures in long days. Using transgenic lines and accessions, we show that the FRI-Ler allele confers temperature-sensitive late flowering confirming a role for FRI in photoperiod-dependent thermal response. Through quantitative complementation with heterogeneous inbred families, we further show that cis-regulatory variation at FT contributes to the observed hypersensitivity of Tsu-0 to ambient temperature. Overall our results suggest that multiple loci that interact epistatically govern photoperiod-dependent thermal responses of A. thaliana.

Genetic Architecture of Natural Variation in Thermal Responses of Arabidopsis.

Wild strains of Arabidopsis (Arabidopsis thaliana) exhibit extensive natural variation in a wide variety of traits, including response to environmental changes. Ambient temperature is one of the major external factors that modulates plant growth and development. Here, we analyze the genetic architecture of natural variation in thermal responses of Arabidopsis. Exploiting wild accessions and recombinant inbred lines, we reveal extensive phenotypic variation in response to ambient temperature in distinct developmental traits such as hypocotyl elongation, root elongation, and flowering time. We show that variation in thermal response differs between traits, suggesting that the individual phenotypes do not capture all the variation associated with thermal response. Genome-wide association studies and quantitative trait locus analyses reveal that multiple rare alleles contribute to the genetic architecture of variation in thermal response. We identify at least 20 genomic regions that are associated with variation in thermal response. Further characterizations of temperature sensitivity quantitative trait loci that are shared between traits reveal a role for the blue-light receptor CRYPTOCHROME2 (CRY2) in thermosensory growth responses. We show the accession Cape Verde Islands is less sensitive to changes in ambient temperature, and through transgenic analysis, we demonstrate that allelic variation at CRY2 underlies this temperature insensitivity across several traits. Transgenic analyses suggest that the allelic effects of CRY2 on thermal response are dependent on genetic background suggestive of the presence of modifiers. In addition, our results indicate that complex light and temperature interactions, in a background-dependent manner, govern growth responses in Arabidopsis.

Natural variation in arsenate tolerance identifies an arsenate reductase in Arabidopsis thaliana.

The enormous amount of environmental arsenic was a major factor in determining the biochemistry of incipient life forms early in the Earth's history. The most abundant chemical form in the reducing atmosphere was arsenite, which forced organisms to evolve strategies to manage this chemical species. Following the great oxygenation event, arsenite oxidized to arsenate and the action of arsenate reductases became a central survival requirement. The identity of a biologically relevant arsenate reductase in plants nonetheless continues to be debated. Here we identify a quantitative trait locus that encodes a novel arsenate reductase critical for arsenic tolerance in plants. Functional analyses indicate that several non-additive polymorphisms affect protein structure and account for the natural variation in arsenate reductase activity in Arabidopsis thaliana accessions. This study shows that arsenate reductases are an essential component for natural plant variation in As(V) tolerance.

WRKY6 transcription factor restricts arsenate uptake and transposon activation in Arabidopsis.

Stress constantly challenges plant adaptation to the environment. Of all stress types, arsenic was a major threat during the early evolution of plants. The most prevalent chemical form of arsenic is arsenate, whose similarity to phosphate renders it easily incorporated into cells via the phosphate transporters. Here, we found that arsenate stress provokes a notable transposon burst in plants, in coordination with arsenate/phosphate transporter repression, which immediately restricts arsenate uptake. This repression was accompanied by delocalization of the phosphate transporter from the plasma membrane. When arsenate was removed, the system rapidly restored transcriptional expression and membrane localization of the transporter. We identify WRKY6 as an arsenate-responsive transcription factor that mediates arsenate/phosphate transporter gene expression and restricts arsenate-induced transposon activation. Plants therefore have a dual WRKY-dependent signaling mechanism that modulates arsenate uptake and transposon expression, providing a coordinated strategy for arsenate tolerance and transposon gene silencing.

Role of actin cytoskeleton in brassinosteroid signaling and in its integration with the auxin response in plants.

In plants, developmental programs and tropisms are modulated by the phytohormone auxin. Auxin reconfigures the actin cytoskeleton, which controls polar localization of auxin transporters such as PIN2 and thus determines cell-type-specific responses. In conjunction with a second growth-promoting phytohormone, brassinosteroid (BR), auxin synergistically enhances growth and gene transcription. We show that BR alters actin configuration and PIN2 localization in a manner similar to that of auxin. We describe a BR constitutive-response mutant that bears an allele of the ACTIN2 gene and shows altered actin configuration, PIN2 delocalization, and a broad array of phenotypes that recapitulate BR-treated plants. Moreover, we show that actin filament reconfiguration is sufficient to activate BR signaling, which leads to an enhanced auxin response. Our results demonstrate that the actin cytoskeleton functions as an integration node for the BR signaling pathway and auxin responsiveness.

Novel natural alleles at FLC and LVR loci account for enhanced vernalization responses in Arabidopsis thaliana.

Vernalization, the induction of flowering by low winter temperatures, is likely to be involved in plant climatic adaptation. However, the genetic, molecular and ecological bases underlying the quantitative variation that tunes vernalization sensitivity to natural environments are largely unknown. To address these questions, we have studied the enhanced vernalization response shown by the Ll-0 accession of Arabidopsis thaliana. Quantitative trait locus (QTL) mapping for several flowering initiation traits in relation to vernalization, in a new Ler × Ll-0 recombinant inbred line (RIL) population, identified large effect alleles at FRI, FLC and HUA2, together with two small effect loci named as Llagostera vernalization response (LVR) 1 and 2. Phenotypic analyses of near isogenic lines validated LVR1 effect on flowering vernalization responses. To further characterize the FLC allele from Ll-0, we carried out genetic association analyses using a regional collection of wild genotypes. FLC-Ll-0 appeared as a low-frequency allele that is distinguished by polymorphism Del(-57), a 50-bp-deletion in the 5'-UTR. Del(-57) was significantly associated with enhanced vernalization responses and FLC RNA expression, as well as with altitude and minimum temperatures. These results are consistent with Del(-57) acting as a novel cis-regulatory FLC polymorphism that may confer climatic adaptation by increasing vernalization sensitivity.