Nutritional and tissue-specific regulation of cytochrome P450 CYP711A MAX1 homologues and strigolactone biosynthesis in wheat.

Strigolactones (SLs) are a class of phytohormones regulating branching/tillering, and their biosynthesis has been associated with nutritional signals and plant adaptation to nutrient-limiting conditions. The enzymes in the SL biosynthetic pathway downstream of carlactone are of interest as they are responsible for structural diversity in SLs, particularly cytochrome P450 CYP711A subfamily members, such as MORE AXILLARY GROWTH1 (MAX1) in Arabidopsis. We identified 13 MAX1 homologues in wheat, clustering in four clades and five homoeologous subgroups. The utilization of RNA-sequencing data revealed a distinct expression pattern of MAX1 homologues in above- and below-ground tissues, providing insights into the distinct roles of MAX1 homologues in wheat. In addition, a transcriptional analysis showed that SL biosynthetic genes were systematically regulated by nitrogen supply. Nitrogen limitation led to larger transcriptional changes in the basal nodes than phosphorus limitation, which was consistent with the observed tillering suppression, as wheat showed higher sensitivity to nitrogen. The opposite was observed in roots, with phosphorus limitation leading to stronger induction of most SL biosynthetic genes compared with nitrogen limitation. The observed tissue-specific regulation of SL biosynthetic genes in response to nutritional signals is likely to reflect the dual role of SLs as rhizosphere signals and branching inhibitors.

The Functional Diversity of the High-Affinity Nitrate Transporter Gene Family in Hexaploid Wheat: Insights from Distinct Expression Profiles.

High-affinity nitrate transporters (NRT) are key components for nitrogen (N) acquisition and distribution within plants. However, insights on these transporters in wheat are scarce. This study presents a comprehensive analysis of the NRT2 and NRT3 gene families, where the aim is to shed light on their functionality and to evaluate their responses to N availability. A total of 53 NRT2s and 11 NRT3s were identified in the bread wheat genome, and these were grouped into different clades and homoeologous subgroups. The transcriptional dynamics of the identified NRT2 and NRT3 genes, in response to N starvation and nitrate resupply, were examined by RT-qPCR in the roots and shoots of hydroponically grown wheat plants through a time course experiment. Additionally, the spatial expression patterns of these genes were explored within the plant. The NRT2s of clade 1, TaNRT2.1-2.6, showed a root-specific expression and significant upregulation in response to N starvation, thus emphasizing a role in N acquisition. However, most of the clade 2 NRT2s displayed reduced expression under N-starved conditions. Nitrate resupply after N starvation revealed rapid responsiveness in TaNRT2.1-2.6, while clade 2 genes exhibited gradual induction, primarily in the roots. TaNRT2.18 was highly expressed in above-ground tissues and exhibited distinct nitrate-related response patterns for roots and shoots. The TaNRT3 gene expression closely paralleled the profiles of TaNRT2.1-2.6 in response to nitrate induction. These findings enhance the understanding of NRT2 and NRT3 involvement in nitrogen uptake and utilization, and they could have practical implications for improving nitrogen use efficiency. The study also recommends a standardized nomenclature for wheat NRT2 genes, thereby addressing prior naming inconsistencies.

The relative abundance of wheat Rubisco activase isoforms is post-transcriptionally regulated.

Diurnal rhythms and light availability affect transcription-translation feedback loops that regulate the synthesis of photosynthetic proteins. The CO2-fixing enzyme Rubisco is the most abundant protein in the leaves of major crop species and its activity depends on interaction with the molecular chaperone Rubisco activase (Rca). In Triticum aestivum L. (wheat), three Rca isoforms are present that differ in their regulatory properties. Here, we tested the hypothesis that the relative abundance of the redox-sensitive and redox-insensitive Rca isoforms could be differentially regulated throughout light-dark diel cycle in wheat. While TaRca1-ß expression was consistently negligible throughout the day, transcript levels of both TaRca2-ß and TaRca2-α were higher and increased at the start of the day, with peak levels occurring at the middle of the photoperiod. Abundance of TaRca-ß protein was maximal 1.5 h after the peak in TaRca2-ß expression, but the abundance of TaRca-α remained constant during the entire photoperiod. The redox-sensitive TaRca-α isoform was less abundant, representing 85% of the redox-insensitive TaRca-ß at the transcript level and 12.5% at the protein level. Expression of Rubisco large and small subunit genes did not show a consistent pattern throughout the diel cycle, but the abundance of Rubisco decreased by up to 20% during the dark period in fully expanded wheat leaves. These results, combined with a lack of correlation between transcript and protein abundance for both Rca isoforms and Rubisco throughout the entire diel cycle, suggest that the abundance of these photosynthetic enzymes is post-transcriptionally regulated.

Wheat amino acid transporters highly expressed in grain cells regulate amino acid accumulation in grain.

Amino acids are delivered into developing wheat grains to support the accumulation of storage proteins in the starchy endosperm, and transporters play important roles in regulating this process. RNA-seq, RT-qPCR, and promoter-GUS assays showed that three amino acid transporters are differentially expressed in the endosperm transfer cells (TaAAP2), starchy endosperm cells (TaAAP13), and aleurone cells and embryo of the developing grain (TaAAP21), respectively. Yeast complementation revealed that all three transporters can transport a broad spectrum of amino acids. RNAi-mediated suppression of TaAAP13 expression in the starchy endosperm did not reduce the total nitrogen content of the whole grain, but significantly altered the composition and distribution of metabolites in the starchy endosperm, with increasing concentrations of some amino acids (notably glutamine and glycine) from the outer to inner starchy endosperm cells compared with wild type. Overexpression of TaAAP13 under the endosperm-specific HMW-GS (high molecular weight glutenin subunit) promoter significantly increased grain size, grain nitrogen concentration, and thousand grain weight, indicating that the sink strength for nitrogen transport was increased by manipulation of amino acid transporters. However, the total grain number was reduced, suggesting that source nitrogen remobilized from leaves is a limiting factor for productivity. Therefore, simultaneously increasing loading of amino acids into the phloem and delivery to the spike would be required to increase protein content while maintaining grain yield.

Phylogeny and gene expression of the complete NITRATE TRANSPORTER 1/PEPTIDE TRANSPORTER FAMILY in Triticum aestivum.

NPF genes encode membrane transporters involved in the transport of a large variety of substrates including nitrate and peptides. The NPF gene family has been described for many plants, but the whole NPF gene family for wheat has not been completely identified. The release of the wheat reference genome has enabled the identification of the entire wheat NPF gene family. A systematic analysis of the whole wheat NPF gene family was performed, including responses of specific gene expression to development and nitrogen supply. A total of 331 NPF genes (113 homoeologous groups) have been identified in wheat. The chromosomal location of the NPF genes is unevenly distributed, with predominant occurrence in the long arms of the chromosomes. The phylogenetic analysis indicated that wheat NPF genes are closely clustered with Arabidopsis, Brachypodium, and rice orthologues, and subdivided into eight subfamilies. The expression profiles of wheat NPF genes were examined using RNA-seq data, and a subset of 44 NPF genes (homoeologous groups) with contrasting expression responses to nitrogen and/or development in different tissues were identified. The systematic identification of gene composition, chromosomal locations, evolutionary relationships, and expression profiles contributes to a better understanding of the roles of the wheat NPF genes and lays the foundation for further functional analysis in wheat.

Foliar N Application at Anthesis Stimulates Gene Expression of Grain Protein Fractions and Alters Protein Body Distribution in Winter Wheat (Triticum aestivum L.).

The major components of wheat storage proteins are gliadins and glutenins, and as they contribute differently to baking quality, a balanced mixture of these components is essential. The application of foliar nitrogen (N) at anthesis is a common practice to improve protein concentration and composition. The aim of this study was to investigate the effects of a foliar N application at anthesis on storage protein gene expression during grain development and on the distribution of protein concentration and protein body size within the grain. In this experiment, an additional N application at anthesis stimulated the expression of genes of the majority of storage proteins when the N supply was low. Furthermore, it led to higher protein concentrations in the subaleurone layers, while in the center of the lobes, the protein concentrations were decreased. These changes will affect the protein recovery in white flours, as proportionally more protein might be lost during milling processes.

Temperature and nitrogen supply interact to determine protein distribution gradients in the wheat grain endosperm.

Gradients exist in the distribution of storage proteins in the wheat (Triticum aestivum) endosperm and determine the milling properties and protein recovery rate of the grain. A novel image analysis technique was developed to quantify both the gradients in protein concentration, and the size distribution of protein bodies within the endosperm of wheat plants grown under two different (20 or 28 °C) post-anthesis temperatures, and supplied with a nutrient solution with either high or low nitrogen content. Under all treatment combinations, protein concentration was greater in the endosperm cells closest to the aleurone layer and decreased towards the centre of the two lobes of the grain, i.e. a negative gradient. This was accompanied by a decrease in size of protein bodies from the outer to the inner endosperm layers in all but one of the treatments. Elevated post-anthesis temperature had the effect of increasing the magnitude of the negative gradients in both protein concentration and protein body size, whilst limiting nitrogen supply decreased the gradients.

Characterization of the Wheat Leaf Metabolome during Grain Filling and under Varied N-Supply.

Progress in improving crop growth is an absolute goal despite the influence multifactorial components have on crop yield and quality. An Avalon × Cadenza doubled-haploid wheat mapping population was used to study the leaf metabolome of field grown wheat at weekly intervals during the time in which the canopy contributes to grain filling, i.e., from anthesis to 5 weeks post-anthesis. Wheat was grown under four different nitrogen supplies reaching from residual soil N to a luxury over-fertilization (0, 100, 200, and 350 kg N ha-1). Four lines from a segregating doubled haploid population derived of a cross of the wheat elite cvs. Avalon and Cadenza were chosen as they showed pairwise differences in either N utilization efficiency (NUtE) or senescence timing. 108 annotated metabolites of primary metabolism and ions were determined. The analysis did not provide genotype specific markers because of a remarkable stability of the metabolome between lines. We speculate that the reason for failing to identify genotypic markers might be due to insufficient genetic diversity of the wheat parents and/or the known tendency of plants to keep metabolome homeostasis even under adverse conditions through multiple adaptations and rescue mechanism. The data, however, provided a consistent catalogue of metabolites and their respective responses to environmental and developmental factors and may bode well for future systems biology approaches, and support plant breeding and crop improvement.

Three new species and one new subspecies of Depressariinae (Lepidoptera) from Europe.

The species Depressaria albarracinella Corley, sp. n., Agonopterix carduncelli Corley, sp. n. and Agonopterix pseudoferulae Buchner & Junnilainen, sp. n. and the subspecies Depressaria saharae Gastón & Vives ssp. tabelli Buchner, ssp. n. are described. Depressaria albarracinella was first found in Spain in 1969 and recognised as apparently new but the specimens in NHMUK have remained undescribed. Additional Spanish material has been located in ZMUC and other collections and three specimens have been found from Greece. Agonopterix carduncelli. A single male of an unidentified Agonopterix of the pallorella group was found in Algarve, Portugal in 2010. A search for larvae in March 2011 was successful and one male and one female were reared from Carthamus caeruleus. Additional specimens of the new species have been located in collections from Spain, Greece and Morocco. Agonopterix pseudoferulae. A specimen from Greece with the name Agonopterix ferulae (Zeller, 1847) found in the Klimesch collection in ZSM had forewing markings which suggested that it might be a different species. Further specimens from Italy and Greece have been examined, among them two reared from Elaeoselinum asclepium (Apiaceae). Both genitalia and barcode show that this is an undescribed species. Depressaria saharae Gastón & Vives, 2017 was described very recently (Gastón and Vives 2017) from northern Spain with a brief description, and figures of two males and male genitalia. Here the new species is redescribed, and additional data on distribution and relationships of the new species added. The opportunity is also taken to show that Canary Islands specimens with the same male genitalia should be treated as a new subspecies D. saharae ssp. tabelli Buchner, ssp. n.

The role of ZIP transporters and group F bZIP transcription factors in the Zn-deficiency response of wheat (Triticum aestivum).

Understanding the molecular basis of zinc (Zn) uptake and transport in staple cereal crops is critical for improving both Zn content and tolerance to low-Zn soils. This study demonstrates the importance of group F bZIP transcription factors and ZIP transporters in responses to Zn deficiency in wheat (Triticum aestivum). Seven group F TabZIP genes and 14 ZIPs with homeologs were identified in hexaploid wheat. Promoter analysis revealed the presence of Zn-deficiency-response elements (ZDREs) in a number of the ZIPs. Functional complementation of the zrt1/zrt2 yeast mutant by TaZIP3, -6, -7, -9 and -13 supported an ability to transport Zn. Group F TabZIPs contain the group-defining cysteine-histidine-rich motifs, which are the predicted binding site of Zn2+ in the Zn-deficiency response. Conservation of these motifs varied between the TabZIPs suggesting that individual TabZIPs may have specific roles in the wheat Zn-homeostatic network. Increased expression in response to low Zn levels was observed for several of the wheat ZIPs and bZIPs; this varied temporally and spatially suggesting specific functions in the response mechanism. The ability of the group F TabZIPs to bind to specific ZDREs in the promoters of TaZIPs indicates a conserved mechanism in monocots and dicots in responding to Zn deficiency. In support of this, TabZIPF1-7DL and TabZIPF4-7AL afforded a strong level of rescue to the Arabidopsis hypersensitive bzip19 bzip23 double mutant under Zn deficiency. These results provide a greater understanding of Zn-homeostatic mechanisms in wheat, demonstrating an expanded repertoire of group F bZIP transcription factors, adding to the complexity of Zn homeostasis.

Two new species of Agonopterix (Depressariidae, Lepidoptera) from Europe.

The species Agonopterix tripunctaria sp. nov. and Agonopterix medelichensis sp. nov. are described. A. tripunctaria, previously misidentified as Agonopterix nodiflorella (Millière, 1866), was recognized as specifically different by the distinctive male genitalia. 19 specimens have been examined, DNA-barcoding yielded full 658 bp fragment of COI from two specimens and a 639 bp sequence from a third, confirming the impression of a rather isolated species. Specimens from Italy, Slovenia, Croatia and Greece had been checked, among them one reared from Ferulago campestris. A. medelichensis was misidentified as Agonopterix rotundella (Douglas, 1846) in NHMV; its male genitalia are erroneously depicted as A. rotundella in Hannemann (1953) and (1995). 20 specimens have been examined, from one a 555 bp fragment of COI was obtained, confirming that it is not closely related to A. rotundella. Specimens from Austria, Italy, Hungary, Slovakia, Croatia and Greece have been checked, among them one reared from Trinia glauca, which had been misidentified as A. hippomarathri. A report of A. rotundella from Russia also belongs to this species.

Expression patterns of C- and N-metabolism related genes in wheat are changed during senescence under elevated CO2 in dry-land agriculture.

Projected climatic impacts on crop yield and quality, and increased demands for production, require targeted research to optimise nutrition of crop plants. For wheat, post-anthesis carbon and nitrogen remobilisation from vegetative plant parts and translocation to grains directly affects grain carbon (C), nitrogen (N) and protein levels. We analysed the influence of increased atmospheric CO2 on the expression of genes involved in senescence, leaf carbohydrate and nitrogen metabolism and assimilate transport in wheat under field conditions (Australian Grains Free Air CO2 Enrichment; AGFACE) over a time course from anthesis to maturity, the key period for grain filling. Wheat grown under CO2 enrichment had lower N concentrations and a tendency towards greater C/N ratios. A general acceleration of the senescence process by elevated CO2 was not confirmed. The expression patterns of genes involved in carbohydrate metabolism, nitrate reduction and metabolite transport differed between CO2 treatments, and this CO2 effect was different between pre-senescence and during senescence. The results suggest up-regulation of N remobilisation and down-regulation of C remobilisation during senescence under elevated CO2, which is consistent with greater grain N-sink strength of developing grains.

Genetic diversity for root plasticity and nitrogen uptake in wheat seedlings.

Enhancing nitrogen use efficiency (NUE) of wheat is a major focus for wheat breeding programs. NUE may be improved by identifying genotypes that are competitive for nitrogen (N) uptake in early vegetative stages of growth and are able to invest that N in grain. Breeders tend to select high yielding genotypes under conditions of medium to high N supply, but it is not known whether this influences the selection of root plasticity traits or whether, over time, breeders have selected genotypes with higher N uptake efficiency. To address this, genotypes were selected from CIMMYT (1966-1985) and Australian (1999-2007) breeding programs. Genotypes from both programs responded to low N supply by expanding their root surface area through increased total root number and/or length of lateral roots. Australian genotypes were N responsive (accumulated more N under high N than under low N) whereas CIMMYT genotypes were not very N responsive. This could not be explained by differences in N uptake capacity as shown by 15N flux analysis of two representative genotypes with contrasting N accumulation. Expression analysis of nitrate transporter genes revealed that the high-affinity transport system accounted for the majority of root nitrate uptake in wheat seedlings under both low and high N conditions.

Complex phylogeny and gene expression patterns of members of the NITRATE TRANSPORTER 1/PEPTIDE TRANSPORTER family (NPF) in wheat.

NPF (formerly referred to as low-affinity NRT1) and 'high-affinity' NRT2 nitrate transporter genes are involved in nitrate uptake by the root, and transport and distribution of nitrate within the plant. The NPF gene family consists of 53 members in Arabidopsis thaliana, however only 11 of these have been functionally characterized. Although homologous genes have been identified in genomes of different plant species including some cereals, there is little information available for wheat (Triticum aestivum). Sixteen genes were identified in wheat homologous to characterized Arabidopsis low-affinity nitrate transporter NPF genes, suggesting a complex wheat NPF gene family. The regulation of wheat NFP genes by plant N-status indicated involvement of these transporters in substrate transport in relation to N-metabolism. The complex expression pattern in relation to tissue specificity, nitrate availability and senescence may be associated with the complex growth patterns of wheat depending on sink/source demands, as well as remobilization during grain filling.

Mineral composition analysis: measuring anion uptake and anion concentrations in plant tissues.

This chapter describes two basic complementary methods relevant to at least three major macronutrients in plants: NO(3)(-), SO(4)(2-), and phosphate. The first method is the simultaneous determination of tissue content of the oxyanions, NO(3)(-), SO(4)(2-), and phosphate by HPLC, and the second is the determination of tissue uptake (transport) capacity for these same oxyanions. NO(3)(-), phosphate, and SO(4)(2-) , as well as other anions including chloride, malate, and nitrite are extracted from milligram quantities of plant tissue and are separated and quantified in a single chromatographic (HPLC) run. Information on uptake (flux) of these same anions through the roots may be obtained using isotopically labeled elements, enabling transport capacity of roots and subsequent translocation to shoot tissues to be determined.

A comparison of sulfate and selenium accumulation in relation to the expression of sulfate transporter genes in Astragalus species.

Sulfate and selenate uptake were investigated in both selenium (Se) hyperaccumulators (Astragalus racemosus and Astragalus bisulcatus) and closely related nonaccumulator species (Astragalus glycyphyllos and Astragalus drummondii). Sulfur (S) starvation increased Se accumulation, whereas increased selenate supply increased sulfate accumulation in both root and shoot tissues. cDNAs for homologs of groups 1 to 4 sulfate transporters were cloned from these Astragalus species to investigate patterns of expression and interactions with sulfate and selenate uptake. In contrast to all other previously analyzed plant species, abundant gene expression of putative sulfate transporters was observed for both Se-hyperaccumulating and nonaccumulating Astragalus, regardless of S and Se status. Furthermore, quantitative analysis of expression indicated a transcript level in Se-hyperaccumulating Astragalus comparable with other plant species under S deprivation. The high expression of sulfate transporters in certain Astragalus species may lead to enhanced Se uptake and translocation ability and therefore may contribute to the Se hyperaccumulation trait; however, it is not sufficient to explain S/Se discriminatory mechanisms.

Evolutionary relationships and functional diversity of plant sulfate transporters.

Sulfate is an essential nutrient cycled in nature. Ion transporters that specifically facilitate the transport of sulfate across the membranes are found ubiquitously in living organisms. The phylogenetic analysis of known sulfate transporters and their homologous proteins from eukaryotic organisms indicate two evolutionarily distinct groups of sulfate transport systems. One major group named Tribe 1 represents yeast and fungal SUL, plant SULTR, and animal SLC26 families. The evolutionary origin of SULTR family members in land plants and green algae is suggested to be common with yeast and fungal SUL and animal anion exchangers (SLC26). The lineage of plant SULTR family is expanded into four subfamilies (SULTR1-SULTR4) in land plant species. By contrast, the putative SULTR homologs from Chlorophyte green algae are in two separate lineages; one with the subfamily of plant tonoplast-localized sulfate transporters (SULTR4), and the other diverged before the appearance of lineages for SUL, SULTR, and SLC26. There also was a group of yet undefined members of putative sulfate transporters in yeast and fungi divergent from these major lineages in Tribe 1. The other distinct group is Tribe 2, primarily composed of animal sodium-dependent sulfate/carboxylate transporters (SLC13) and plant tonoplast-localized dicarboxylate transporters (TDT). The putative sulfur-sensing protein (SAC1) and SAC1-like transporters (SLT) of Chlorophyte green algae, bryophyte, and lycophyte show low degrees of sequence similarities with SLC13 and TDT. However, the phylogenetic relationship between SAC1/SLT and the other two families, SLC13 and TDT in Tribe 2, is not clearly supported. In addition, the SAC1/SLT family is absent in the angiosperm species analyzed. The present study suggests distinct evolutionary trajectories of sulfate transport systems for land plants and green algae.

Influence of sulfur deficiency on the expression of specific sulfate transporters and the distribution of sulfur, selenium, and molybdenum in wheat.

Interactions between sulfur (S) nutritional status and sulfate transporter expression in field-grown wheat (Triticum aestivum) were investigated using Broadbalk +S and -S treatments (S fertilizer withheld) at Rothamsted, United Kingdom. In 2008, S, sulfate, selenium (Se), and molybdenum (Mo) concentrations and sulfate transporter gene expression were analyzed throughout development. Total S concentrations were lower in all tissues of -S plants, principally as a result of decreased sulfate pools. S, Se, and Mo concentrations increased in vegetative tissues until anthesis, and thereafter, with the exception of Mo, decreased until maturity. At maturity, most of the S and Se were localized in the grain, indicating efficient remobilization from vegetative tissues, whereas less Mo was remobilized. At maturity, Se and Mo were enhanced 7- and 3.7-fold, respectively, in -S compared with +S grain, while grain total S was not significantly reduced. Enhanced expression of sulfate transporters, for example Sultr1;1 and Sultr4;1, in -S plants explains the much increased accumulation of Se and Mo (7- and 3.7-fold compared with +S in grain, respectively). Sultr5;2 (mot1), thought to be involved in Mo accumulation in Arabidopsis (Arabidopsis thaliana), did not fully explain patterns of Mo distribution; it was expressed in all tissues, decreasing in leaf and increasing in roots under -S conditions, and was expressed in florets at anthesis but not in grain at any other time. In conclusion, S fertilizer application has a marked impact on Mo and Se distribution and accumulation, which is at least partially a result of altered gene expression of the sulfate transporter family.

The sulfate transporter family in wheat: tissue-specific gene expression in relation to nutrition.

Sulfate uptake and distribution in plants are managed by the differential expression of a family of transporters, developmentally, spatially, and in response to sulfur nutrition. Elucidation of the signaling pathways involved requires a knowledge of the component parts and their interactions. Here, the expression patterns of the full complement of sulfate transporters in wheat, as influenced by development and sulfur nutrition, are described. The 10 wheat sulfate transporters characterized here are compared to the gene families for both rice and Brachypodium, for whom full genome information is available. Expression is reported in young seedlings with a focus on roles in uptake from nutrient solution and differential expression in relation to sulfate deprivation. In addition, patterns of expression in all organs at the grain filling stage are reported and indicate differential responses to nutritional signals of the individual transporters in specific tissues and an overall coordination of uptake, storage, and remobilization to deliver sulfur to the developing grain.

Expression and activity of sulfate transporters and APS reductase in curly kale in response to sulfate deprivation and re-supply.

Both activity and expression of sulfate transporters and APS reductase in plants are modulated by the sulfur status of the plant. To examine the regulatory mechanisms in curly kale (Brassica oleracea L.), the sulfate supply was manipulated by the transfer of seedlings to sulfate-deprived conditions, which resulted in an up to 3-fold increase in the sulfate uptake capacity by the root, accompanied by an induction of transcript abundances of the Group 1 and 4 sulfate transporters in root and shoot. Upon sulfate re-supply, there was no correlation between the activity and expression of the sulfate transporters. Despite the decrease in the abundance of the sulfate transporter transcripts, especially at the onset of the sulfate re-supply, the sulfate uptake capacity was affected very little for up to 96h. There was no relationship between changes in the sulfate or thiol content and activity and expression of the sulfate transporters. Thus, their significance as regulatory signal compounds remains unresolved. The activity and expression of APS reductase, which was enhanced strongly only in the shoots of sulfate-deprived plants, and rapidly decreased again upon sulfate re-supply, corresponded with changes in thiol content, consistent with this pool having a role as a regulatory signal.