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Abscisic acid (ABA) is a phytohormone that mediates stress responses and regulates plant development. Several ATP-binding cassette (ABC) transporters in the G subfamily of ABC (ABCG) proteins have been reported to transport ABA. We investigated whether there are any other ABCG proteins that mediate plant developmental processes regulated by ABA in Arabidopsis (Arabidopsis thaliana). The ABCG27 gene was upregulated in response to exogenous ABA treatment. The abcg27 knockout mutant exhibited two developmental defects: epinastic leaves and abnormally long pistils, which reduced fertility and silique length. ABCG27 expression was induced threefold when flower buds were exposed to exogenous ABA, and the promoter of ABCG27 had two ABA-responsive elements. ABA content in the pistil and true leaves were increased in the abcg27 knockout mutant. Detached abcg27 pistils exposed to exogenous ABA grew longer than those of the wild-type control. ABCG27 fused to GFP localized to the plasma membrane when expressed in Arabidopsis mesophyll protoplasts. A transcriptome analysis of the pistils and true leaves of the wild type and abcg27 knockout mutant revealed that the expression of organ development-related genes changed in the knockout mutant. In particular, the expression of trans-acting small interference (ta-si) RNA processing enzyme genes, which regulate flower and leaf development, was low in the knockout mutant. Together, these results suggest that ABCG27 most likely function as an ABA transporter at the plasma membrane, modulating ABA levels and thereby regulating the development of the pistils and leaves under normal, non-stressed conditions.

SUMMARYNitrogen (N) as well as Phosphorus (P) are key nutrients determining crop productivity. Legumes have developed strategies to overcome nutrient limitation by e.g., forming a symbiotic relationship with N-fixing rhizobia and the release of P-mobilizing exudates and are thus able to grow without supply of N or P fertilizers. The legume-rhizobial symbiosis starts with root release of isoflavonoids, that act as signaling molecules perceived by compatible bacteria. Subsequently, bacteria release nod factors, which induce signaling cascades allowing the formation of functional N-fixing nodules.We report here the identification and functional characterization of a plasma membrane-localized MATE-type transporter (LaMATE2) involved in the release of genistein from white lupin roots.The LaMATE2 expression in the root is upregulated under N deficiency as well as low phosphate availability, two nutritional deficiencies that induce the release of this isoflavonoid. LaMATE2 silencing reduced genistein efflux and even more the formation of symbiotic nodules, supporting the crucial role of LaMATE2 in isoflavonoid release and nodulation. Furthermore, silencing of LaMATE2 limited the P-solubilization activity of lupin root exudates. Transport assays in yeast vesicles demonstrated that LaMATE2 acts as a proton-driven isoflavonoid transporter.

Summary Abscisic acid (ABA) integrates internal and external signals to coordinate plant development, growth and architecture. It plays a central role in stomatal closure, and prevents germination of freshly produced seeds and germination of non‐dormant seeds under unfavorable circumstances. Here, we describe a Medicago truncatula ATP‐binding cassette (ABC) transporter, MtABCG20, as an ABA exporter present in roots and germinating seeds. In seeds, MtABCG20 was found in the hypocotyl–radicle transition zone of the embryonic axis. Seeds of mtabcg20 plants were more sensitive to ABA upon germination, due to the fact that ABA translocation within mtabcg20 embryos was impaired. Additionally, the mtabcg20 produced fewer lateral roots and formed more nodules compared with wild‐type plants in conditions mimicking drought stress. Heterologous expression in Arabidopsis thaliana provided evidence that MtABCG20 is a plasma membrane protein that is likely to form homodimers. Moreover, export of ABA from Nicotiana tabacum BY2 cells expressing MtABCG20 was faster than in the BY2 without MtABCG20. Our results have implications both in legume crop research and determination of the fundamental molecular processes involved in drought response and germination., Significance Statement In the presented manuscript we introduce the MtABCG20, a half‐size ABC transporter of the G subfamily, as a modulator of abscisic acid distribution in model legume Medicago truncatula, participating in seed germination as well as root morphology and nodulation. Our results have implications both in legume crop research and determination of the fundamental molecular processes involved in drought response and germination.

Synthetic herbicides have been used widely for more than 70 y and have substantially contributed to the efficiency of agriculture. Glyphosate [ N -(phosphonomethyl)glycine] was marketed in 1974 under the trade name Roundup and has become the most used herbicide worldwide. It is a postemergence, nonselective herbicide of low toxicity to animals and humans. While its metabolism in plants is limited, its breakdown in the soil is relatively fast (1). The slow action of glyphosate is related to the fact that it needs to be transported to meristematic areas to become effective. The observation that plants treated with glyphosate accumulate large amounts of shikimate led to the discovery that glyphosate inhibits the 5-enolpyruvyl-shikimate-3 phosphate synthase (EPSPS), an enzyme of the shikimate pathway, which in plants resides in the plastid (Fig. 1) (2). EPSP is converted to chorismate, which is a central metabolite in the synthesis of the three aromatic amino acids. For more than 20 y after its introduction no notable resistance to glyphosate was observed in weeds. In 1996, a transgenic glyphosate-tolerant soybean (Roundup Ready) was introduced, which carried a bacterial gene coding for a glyphosate-insensitive form of EPSPS. Other Roundup Ready major crop plants soon followed, leading to an enormous increase in the application of the herbicide (1, 3). This has raised increasing concern about ground and surface-water pollution and the appearance of residues in food products (4). As a consequence of the increased selective pressure, an increasing number of weeds in fields around the world developed glyphosate resistance (3, 4). Target (EPSPS)-related insensitivity to glyphosate was traced to either mutations in the enzyme or its overproduction by gene amplification or enhanced expression (Fig. 1). The stepwise evolution of mutations (up to three) increased the severity of weed resistance in the field. Other known mechanisms contributing … [↵][1]1To whom correspondence may be addressed. Email: enrico.martinoia{at}uzh.ch. [1]: #xref-corresp-1-1

Recent developments in the field of ABC proteins including newly identified functions and regulatory mechanisms expand the understanding of how they function in the development and physiology of plants.

Members of the ATP-binding cassette (ABC) transporter superfamily translocate a broad spectrum of chemically diverse substrates. While their eponymous ATP-binding cassette in the nucleotide-binding domains (NBDs) is highly conserved, their transmembrane domains (TMDs) forming the translocation pathway exhibit distinct folds and topologies, suggesting that during evolution, the ancient motor domains were combined with different transmembrane mechanical systems to orchestrate a variety of cellular processes. In recent years, it has become increasingly evident that the distinct TMD folds are best suited to categorize the multitude of ABC transporters. We therefore propose a new ABC transporter classification that currently comprises seven different types based on structural homology in the TMDs.

Abscisic acid (ABA) is generally known as the plant stress hormone. Functioning in a wide range of environmental responses, ABA plays a major role in drought tolerance. In addition to inducing stomatal closure during drought stress, ABA promotes suberization of the exodermis and endodermis, which reduces water loss from the root. Furthermore, ABA increases freezing tolerance and has a complex, but not completely understood, role in plant–pathogen interactions. ABA also functions in plant development; for example, ABA is a central player in maintaining seed dormancy. Whereas the enzymatic steps of ABA biosynthesis have been known for some time, our knowledge of ABA receptors and transporters is quite recent. This is due, at least partially, to redundancy among members of both the ABA receptor and transporter families. Many transporters from different transporter families cooperate to transport ABA. The weak but distinct phenotypes described for the different loss-of-function mutants indicate that each of these transporters plays a specific role and, at least under a given condition or in a specific tissue, they are not completely redundant. However, for each function described so far, delivery of ABA at the target site requires the activity of several different ABA transporters. This strategy may ensure that ABA is transported to the correct target even if one of the transporters is nonfunctional or that plants can transport ABA under a given condition via several routes.

Phospholipids (PLs) are emerging as important factors that initiate signal transduction cascades at the plasma membrane. Their distribution within biological membranes is tightly regulated, e.g. by ATP-binding cassette (ABC) transporters, which preferably translocate PLs from the cytoplasmic to the exoplasmic membrane leaflet and are therefore called PL-floppases. Here, we demonstrate that a plant ABC transporter, Lr34 from wheat (Triticum aestivum), is involved in plasma membrane remodeling characterized by an intracellular accumulation of phosphatidic acid and enhanced outward translocation of phosphatidylserine. In addition, the content of phosphatidylinositol 4,5-bisphosphate in the cytoplasmic leaflet of the plasma membrane was reduced in the presence of the ABC transporter. When heterologously expressed in Saccharomyces cerevisiae, Lr34 promoted oil body formation in a mutant defective in PL-transfer in the secretory pathway. Our results suggest that PL redistribution by Lr34 potentially affects the membrane-bound proteome and contributes to the previously reported stimuli-independent activation of biotic and abiotic stress responses and neutral lipid accumulation in transgenic Lr34-expressing barley plants.

Summary Arsenic (As) is a poisonous element that causes severe skin lesions and cancer in humans. Rice (Oryza sativa L.) is a major dietary source of As in humans who consume this cereal as a staple food. We hypothesized that increasing As vacuolar sequestration would inhibit its translocation into the grain and reduce the amount of As entering the food chain. We developed transgenic rice plants expressing two different vacuolar As sequestration genes, ScYCF1 and OsABCC1, under the control of the RCc3 promoter in the root cortical and internode phloem cells, along with a bacterial γ‐glutamylcysteine synthetase driven by the maize UBI promoter. The transgenic rice plants exhibited reduced root‐to‐shoot and internode‐to‐grain As translocation, resulting in a 70% reduction in As accumulation in the brown rice without jeopardizing agronomic traits. This technology could be used to reduce As intake, particularly in populations of South East Asia suffering from As toxicity and thereby improve human health.

Plant health in natural environments depends on interactions with complex and dynamic communities comprising macro- and microorganisms. While many studies have provided insights into the composition of rhizosphere microbiomes (rhizobiomes), little is known about whether plants shape their rhizobiomes. Here, we discuss physiological factors of plants that may govern plant-microbe interactions, focusing on root physiology and the role of root exudates. Given that only a few plant transport proteins are known to be involved in root metabolite export, we suggest novel families putatively involved in this process. Finally, building off of the features discussed in this review, and in analogy to well-known symbioses, we elaborate on a possible sequence of events governing rhizobiome assembly.

The Arabidopsis ATP-Binding Cassette (ABC) transporter ABCC1 sequesters arsenic (As)-phytochelatin conjugates into the vacuole, thereby conferring As resistance. Here, we report the results of a screen for phosphorylation-dependent regulation sites of AtABCC1. Variants of AtABCC1 harboring mutations that replaced amino acid residues Tyr682 , Tyr709 , Tyr822 , Ser846 , Ser1278 , or Thr1408 with alanine confer reduced resistance and decrease the intracellular As content relative to wild-type AtABCC1 when heterologously expressed in the SM7 yeast strain. This suggests that these mutations compromise the vacuolar sequestration of As by AtABCC1. Furthermore, through a phosphomimic mutant study, we found that phosphorylation of Ser846 is required for the As resistance function of AtABCC1. Our analysis provides a first clue as to the phosphorylation-mediated regulation of AtABCC1 activity.

Apocarotenoids are carotenoid derivatives produced by the nonenzymatic or enzymatic cleavage of carotenoids, followed by different enzymatic modifications. In plants, apocarotenoids play different roles, such as attraction of pollinators and seeds dispersal, defense against pathogens and herbivores, protection against photo-oxidative stresses, stimulation and inhibition of plant growth and regulation of biological processes in the case of phytohormones abscisic acid and strigolactones. While carotenoids are in general plastid-localized metabolites, apocarotenoids can reach different final destinations inside or outside the cell. The mechanisms of apocarotenoid transport through biological membranes have been poorly studied. This chapter describes a method to characterize transmembrane transporters involved in the transport of polar and amphipathic apocarotenoids. This protocol was successfully used to in vitro characterize the transport activity of ATP-binding cassette (ABC) and multidrug and toxic extrusion (MATE) in microsomes isolated from Saccharomyces cerevisiae expressing these plant transporters.

Apocarotenoids are carotenoid derivatives produced by the nonenzymatic or enzymatic cleavage of carotenoids, followed by different enzymatic modifications. In plants, apocarotenoids play different roles, such as attraction of pollinators and seeds dispersal, defense against pathogens and herbivores, protection against photo-oxidative stresses, stimulation and inhibition of plant growth and regulation of biological processes in the case of phytohormones abscisic acid and strigolactones. While carotenoids are in general plastid-localized metabolites, apocarotenoids can reach different final destinations inside or outside the cell. The mechanisms of apocarotenoid transport through biological membranes have been poorly studied. This chapter describes a method to characterize transmembrane transporters involved in the transport of polar and amphipathic apocarotenoids. This protocol was successfully used to in vitro characterize the transport activity of ATP-binding cassette (ABC) and multidrug and toxic extrusion (MATE) in microsomes isolated from Saccharomyces cerevisiae expressing these plant transporters.

Compartmentation is a key strategy enacted by plants for the storage of specialized metabolites. The saffron spice owes its red color to crocins, a complex mixture of apocarotenoid glycosides that accumulate in intracellular vacuoles and reach up to 10% of the spice dry weight. We developed a general approach, based on coexpression analysis, heterologous expression in yeast (Saccharomyces cerevisiae), and in vitro transportomic assays using yeast microsomes and total plant metabolite extracts, for the identification of putative vacuolar metabolite transporters, and we used it to identifyCrocus sativustransporters mediating vacuolar crocin accumulation in stigmas. Three transporters, belonging to both the multidrug and toxic compound extrusion and ATP binding cassette C (ABCC) families, were coexpressed with crocins and/or with the gene encoding the first dedicated enzyme in the crocin biosynthetic pathway, CsCCD2. Two of these, belonging to the ABCC family, were able to mediate transport of several crocins when expressed in yeast microsomes. CsABCC4a was selectively expressed inC. sativusstigmas, was predominantly tonoplast localized, transported crocins in vitro in a stereospecific and cooperative way, and was able to enhance crocin accumulation when expressed inNicotiana benthamianaleaves.

The PDR-type ABCG transporter, ABCG36/PDR8/PEN3, is thought to be implicated in the export of a few structurally unrelated substrates, including the auxin precursor, indole-3-butyric acid (IBA), although a clear-cut proof of transport is lacking. An outward facing, lateral root (LR) location for ABCG36 fuelled speculations that it might secrete IBA into the rhizosphere. Here, we provide strong evidence that ABCG36 catalyzes the export of IBA – but not of indole-3-acetic acid – through the plasma membrane. ABCG36 seems to function redundantly with the closely related isoform ABCG37/PDR9/PIS1 in a negative control of rootward IBA transport in roots, which might be dampened by concerted, lateral IBA export. Analyses of single and double mutant phenotypes suggest that both ABCG36 and ABCG37 function cooperatively in auxin-controlled plant development. Both seem to possess a dual function in the control of auxin homeostasis in the root tip and long-range transport in the mature root correlating with non-polar and polar expression profiles in the LR cap and epidermis, respectively.

Summary The wheat Lr34res allele, coding for an ATP‐binding cassette transporter, confers durable resistance against multiple fungal pathogens. The Lr34sus allele, differing from Lr34res by two critical nucleotide polymorphisms, is found in susceptible wheat cultivars. Lr34res is functionally transferrable as a transgene into all major cereals, including rice, barley, maize, and sorghum.Here, we used transcriptomics, physiology, genetics, and in vitro and in vivo transport assays to study the molecular function of Lr34.We report that Lr34res results in a constitutive induction of transcripts reminiscent of an abscisic acid (ABA)‐regulated response in transgenic rice. Lr34‐expressing rice was altered in biological processes that are controlled by this phytohormone, including dehydration tolerance, transpiration and seedling growth. In planta seedling and in vitro yeast accumulation assays revealed that both LR34res and LR34sus act as ABA transporters. However, whereas the LR34res protein was detected in planta the LR34sus version was not, suggesting a post‐transcriptional regulatory mechanism.Our results identify ABA as a substrate of the LR34 ABC transporter. We conclude that LR34res‐mediated ABA redistribution has a major effect on the transcriptional response and physiology of Lr34res‐expressing plants and that ABA is a candidate molecule that contributes to Lr34res‐mediated disease resistance.

Tip-focused accumulation of reactive oxygen species (ROS) is tightly associated with pollen tube growth and is thus critical for fertilization. However, it is unclear how tip-growing cells establish such specific ROS localization. Polyamines have been proposed to function in tip growth as precursors of the ROS, hydrogen peroxide. The ABC transporter AtABCG28 may regulate ROS status, as it contains multiple cysteine residues, a characteristic of proteins involved in ROS homeostasis. In this study, we found that AtABCG28 was specifically expressed in the mature pollen grains and pollen tubes. AtABCG28 was localized to secretory vesicles inside the pollen tube that moved toward and fused with the plasma membrane of the pollen tube tip. Knocking out AtABCG28 resulted in defective pollen tube growth, failure to localize polyamine and ROS to the growing pollen tube tip, and complete male sterility, whereas ectopic expression of this gene in root hair could recover ROS accumulation at the tip and improved the growth under high-pH conditions, which normally prevent ROS accumulation and tip growth. Together, these data suggest that AtABCG28 is critical for localizing polyamine and ROS at the growing tip. In addition, this function of AtABCG28 is likely to protect the pollen tube from the cytotoxicity of polyamine and contribute to the delivery of polyamine to the growing tip for incorporation into the expanding cell wall.

As a first line of defense against insect herbivores many plants store high concentrations of toxic and deterrent secondary metabolites in glandular trichomes. Plant Pleiotropic Drug Resistance (PDR)-type ABC transporters are known secondary metabolite transporters, and several have been implicated in pathogen or herbivore defense. Here, we report on Petunia hybrida PhPDR2 as a major contributor to trichome-related chemical defense. PhPDR2 was found to localize to the plasma membrane and be predominantly expressed in multicellular glandular trichomes of leaves and stems. Down-regulation of PhPDR2 via RNA interference (pdr2) resulted in a markedly higher susceptibility of the transgenic plants to the generalist foliage feeder Spodoptera littoralis. Untargeted screening of pdr2 trichome metabolite contents showed a significant decrease in petuniasterone and petuniolide content, compounds, which had previously been shown to act as potent toxins against various insects. Our findings suggest that PhPDR2 plays a leading role in controlling petuniasterone levels in leaves and trichomes of petunia, thus contributing to herbivory resistance.

Summary Plants reorganize their root architecture to avoid growth into unfavorable regions of the rhizosphere. In a screen based on chimeric repressor gene‐silencing technology, we identified the Arabidopsis thaliana GeBP‐LIKE 4 (GPL4) transcription factor as an inhibitor of root growth that is induced rapidly in root tips in response to cadmium (Cd).We tested the hypothesis that GPL4 functions in the root avoidance of Cd by analyzing root proliferation in split medium, in which only half of the medium contained toxic concentrations of Cd.The wild‐type (WT) plants exhibited root avoidance by inhibiting root growth in the Cd side but increasing root biomass in the control side. By contrast, GPL4‐suppression lines exhibited nearly comparable root growth in the Cd and control sides and accumulated more Cd in the shoots than did the WT. GPL4 suppression also altered the root avoidance of toxic concentrations of other essential metals, modulated the expression of many genes related to oxidative stress, and consistently decreased reactive oxygen species concentrations.We suggest that GPL4 inhibits the growth of roots exposed to toxic metals by modulating reactive oxygen species concentrations, thereby allowing roots to colonize noncontaminated regions of the rhizosphere.

Two Arabidopsis ABC transporters, ABCG1 and ABCG16, are expressed in the tapetal layer, specifically after postmeiotic microspore release, and play important roles in pollen surface development. The male gametophytic cells of terrestrial plants, the pollen grains, travel far before fertilization, and thus require strong protective layers, which take the form of a pollen coat and a pollen wall. The protective surface structures are generated by the tapetum, the tissue surrounding the developing gametophytes. Many ABC transporters, including Arabidopsis thaliana ABCG1 and ABCG16, have been shown to play essential roles in the development of such protective layers. However, the details of the mechanism of their function remain to be clarified. In this study, we show that ABCG1 and ABCG16 are localized at the plasma membrane of tapetal cells, specifically after postmeiotic microspore release, and play critical roles in the postmeiotic stages of male gametophyte development. Consistent with this stage-specific expression, the abcg1 abcg16 double knockout mutant exhibited defects in pollen development after postmeiotic microspore release; their microspores lacked intact nexine and intine layers, exhibited defects in pollen mitosis I, displayed ectopic deposits of arabinogalactan proteins, failed to complete cytokinesis, and lacked sperm cells. Interestingly, the double mutant exhibited abnormalities in the internal structures of tapetal cells, too; the storage organelles of tapetal cells, tapetosomes and elaioplasts, were morphologically altered. Thus, this work reveals that the lack of ABCG1 and ABCG16 at the tapetal cell membrane causes a broad range of defects in pollen, as well as in tapetal cells themselves. Furthermore, these results suggest that normal pollen surface development is necessary for normal development of the pollen cytoplasm.

Highlight Plants with altered intracellular Pi compartmentation surprisingly show accelerated dark-induced senescence. We propose that low cytosolic phosphate concentrations are of critical importance for proper plant development., The phosphate transporter PHT4;6 locates to the trans-Golgi compartment, and its impaired activity causes altered intracellular phosphate compartmentation, leading to low cytosolic Pi levels, a blockage of Golgi-related processes such as protein glycosylation and hemicellulose biosynthesis, and a dwarf phenotype. However, it was unclear whether altered Pi homeostasis in pht4;6 mutants causes further cellular problems, typically associated with limited phosphate availability. Here we report that pht4;6 mutants exhibit a markedly increased disposition to induce dark-induced senescence. In control experiments, in which pht4;6 mutants and wild-type plants developed similarly, we confirmed that accelerated dark-induced senescence in mutants is not a ‘pleiotropic’ process associated with the dwarf phenotype. In fact, accelerated dark-induced senescence in pht4;6 mutants correlates strongly with increased levels of toxic NH4 + and higher sensitivity to ammonium, which probably contribute to the inability of pht4;6 mutants to recover from dark treatment. Experiments with modified levels of either salicylic acid (SA) or trans-zeatin (tZ) demonstrate that altered concentrations of these compounds in pht4;6 plants act as major cellular mediators for dark-induced senescence. This conclusion gained further support from the notion that the expression of the pht4;6 gene is, in contrast to genes coding for major phosphate importers, substantially induced by tZ. Taken together, our findings point to a critical function of PHT4;6 to control cellular phosphate levels, in particular the cytosolic Pi availability, required to energize plant primary metabolism for proper plant development. Phosphate and its allocation mediated by PHT4;6 is critical to prevent onset of dark-induced senescence.

The plant vacuole is a cellular compartment that is essential to plant development and growth. Often plant vacuoles accumulate specialized metabolites, also called secondary metabolites, which constitute functionally and chemically diverse compounds that exert in planta many essential functions and improve the plant's fitness. These metabolites provide, for example, chemical defense against herbivorous and pathogens or chemical attractants (color and fragrance) to attract pollinators. The chemical composition of the vacuole is dynamic, and is altered during development and as a response to environmental changes. To some extent these alterations rely on vacuolar transporters, which import and export compounds into and out of the vacuole, respectively. During the past decade, significant progress was made in the identification and functional characterization of the transporters implicated in many aspects of plant specialized metabolism. Still, deciphering the molecular players underlying such processes remains a challenge for the future. In this review, we present a comprehensive summary of the most recent achievements in this field.

Soil contamination by heavy metals and metalloids such as cadmium (Cd) and arsenic (As) poses a major threat to the environment and to human health. Vacuolar sequestration is one of the main mechanisms by which plants control toxic materials including Cd and As. Understanding the mechanisms of heavy metal tolerance and accumulation can be useful for both phytoremediation and safe crop development. In this review, we summarize recent advances in deciphering the molecular mechanisms underlying vacuolar sequestration of Cd and As, and discuss potential biotechnological applications of this knowledge and efforts towards attaining these goals.

The exact transport characteristics of the vacuolar dicarboxylate transporter tDT from Arabidopsis are elusive. To overcome this limitation, we combined a range of experimental approaches comprising generation/analysis of tDT overexpressors, 13CO2 feeding and quantification of 13C enrichment, functional characterization of tDT in proteoliposomes, and electrophysiological studies on vacuoles. tdt knockout plants showed decreased malate and increased citrate concentrations in leaves during the diurnal light-dark rhythm and after onset of drought, when compared with wildtypes. Interestingly, under the latter two conditions, tDT overexpressors exhibited malate and citrate levels opposite to tdt knockout plants. Highly purified tDT protein transports malate and citrate in a 1:1 antiport mode. The apparent affinity for malate decreased with decreasing pH, whereas citrate affinity increased. This observation indicates that tDT exhibits a preference for dianion substrates, which is supported by electrophysiological analysis on intact vacuoles. tDT also accepts fumarate and succinate as substrates, but not α-ketoglutarate, gluconate, sulfate, or phosphate. Taking tDT as an example, we demonstrated that it is possible to reconstitute a vacuolar metabolite transporter functionally in proteoliposomes. The displayed, so far unknown counterexchange properties of tDT now explain the frequently observed reciprocal concentration changes of malate and citrate in leaves from various plant species. tDT from Arabidopsis is the first member of the well-known and widely present SLC13 group of carrier proteins, exhibiting an antiport mode of transport.

Strigolactones (SLs) are carotenoid‐derived phytohormones shaping plant architecture and inducing the symbiosis with endomycorrhizal fungi. In Petunia hybrida, SL transport within the plant and towards the rhizosphere is driven by the ABCG‐class protein PDR1. PDR1 expression is regulated by phytohormones and by the soil phosphate abundance, and thus SL transport integrates plant development with nutrient conditions. We overexpressed PDR1 (PDR1 OE) to investigate whether increased endogenous SL transport is sufficient to improve plant nutrition and productivity. Phosphorus quantification and nondestructive X‐ray computed tomography were applied. Morphological and gene expression changes were quantified at cellular and whole tissue levels via time‐lapse microscopy and quantitative PCR. PDR1 OE significantly enhanced phosphate uptake and plant biomass production on phosphate‐poor soils. PDR1 OE plants showed increased lateral root formation, extended root hair elongation, faster mycorrhization and reduced leaf senescence. PDR1 overexpression allowed considerable SL biosynthesis by releasing SL biosynthetic genes from an SL‐dependent negative feedback. The increased endogenous SL transport/biosynthesis in PDR1 OE plants is a powerful tool to improve plant growth on phosphate‐poor soils. We propose PDR1 as an as yet unexplored trait to be investigated for crop production. The overexpression of PDR1 is a valuable strategy to investigate SL functions and transport routes., New Phytologist, 217 (2), ISSN:0028-646X, ISSN:1469-8137

Plant hormones (phytohormones) integrate endogenous and exogenous signals thus synchronizing plant growth with environmental and developmental changes. Similar to animals, phytohormones have distinct source and target tissues, hence controlled transport and focused targeting are required for their functions. Many evidences accumulated in the last years about the regulation of long-distance and directional transport of phytohormones. ATP-binding cassette (ABC) transporters turned out to play major roles in routing phytohormones not only in the plant body but also towards the outer environment. The ABCG-type proteins ABCG25 and ABCG40 are high affinity abscisic acid (ABA) transporters. ABCG14 is highly co-expressed with cytokinin biosynthesis and is the major root-to-shoot cytokinin transporter. Pleiotropic drug resistance1 (PDR1) from Petunia hybrida transports strigolactones (SLs) from the root tip to the plant shoot but also outside to the rhizosphere, where SLs are the main attractants to mycorrhizal fungi. Last but not least, ABCG36 and ABCG37 possibly play a dual role in coumarine and IBA transport.

Mycorrhizas play a pivotal role in phosphorus (P) acquisition of plant roots, by enhancing the soil volume that can be explored. Non-mycorrhizal plant species typically occur either in relatively fertile soil or on soil with a very low P availability, where there is insufficient P in the soil solution for mycorrhizal hyphae to be effective. Soils with a very low P availability are either old and severely weathered or relatively young with high concentrations of oxides and hydroxides of aluminium and iron that sorb P. In such soils, cluster roots and other specialised roots that release P-mobilising carboxylates are more effective than mycorrhizas. Cluster roots are ephemeral structures that release carboxylates in an exudative burst. The carboxylates mobilise sparingly-available sources of soil P. The relative investment of biomass in cluster roots and the amount of carboxylates that are released during the exudative burst differ between species on severely weathered soils with a low total P concentration and species on young soils with high total P concentrations but low P availability. Taking a modelling approach, we explore how the optimal cluster-root strategy depends on soil characteristics, thus offering insights for plant breeders interested in developing crop plants with optimal cluster-root strategies.

Proteins containing a placenta-specific 8 domain (PLAC8) function as major organ size regulators in Solanum lycopersicum and Zea may, and putative metal ion transporters in Arabidopsis thaliana, Oryza sativa and Brassica juncea. However, it is unknown how PLAC8 domain-containing proteins fulfill such diverse roles. Here, we found that plant cadmium resistance 1 (PCR1) influences both zinc (Zn) accumulation and grain weight in rice. OsPCR1 knockout and knockdown lines produced lighter grains than the wild type, while OsPCR1 overexpression lines produced heavier grains. Furthermore, the grains of OsPCR1 knockdown lines exhibited substantially higher Zn and lower cadmium (Cd) concentrations than the control, as did yeast heterologously expressing OsPCR1. Through sequence analysis, we showed that the amino acid sequence of japonica-type PCR1 was distinct from that of indica-type and wild rice accessions. This difference was correlated with distinct Zn-related phenotypes. Japonica-type PCR1 had a shorter N-terminus than did PCR1 in the other rice types, and yeast heterologously expressing japonica-type PCR1 was more sensitive to Zn than was yeast expressing indica-type PCR1. Furthermore, japonica-type grains accumulated less Zn than did indica-type grains. Our study suggests that rice PCR1 maintains metal ion homeostasis and grain weight and might have been selected for during domestication.

Grape (Vitis vinifera) accumulates various polyphenolic compounds, which protect against environmental stresses, including ultraviolet-C (UV-C) light and pathogens. In this study, we looked at the transcriptome and metabolome in grape berry skin after UV-C irradiation, which demonstrated the effectiveness of omics approaches to clarify important traits of grape. We performed transcriptome analysis using a genome-wide microarray, which revealed 238 genes up-regulated more than 5-fold by UV-C light. Enrichment analysis of Gene Ontology terms showed that genes encoding stilbene synthase, a key enzyme for resveratrol synthesis, were enriched in the up-regulated genes. We performed metabolome analysis using liquid chromatography-quadrupole time-of-flight mass spectrometry, and 2,012 metabolite peaks, including unidentified peaks, were detected. Principal component analysis using the peaks showed that only one metabolite peak, identified as resveratrol, was highly induced by UV-C light. We updated the metabolic pathway map of grape in the Kyoto Encyclopedia of Genes and Genomes (KEGG) database and in the KaPPA-View 4 KEGG system, then projected the transcriptome and metabolome data on a metabolic pathway map. The map showed specific induction of the resveratrol synthetic pathway by UV-C light. Our results showed that multiomics is a powerful tool to elucidate the accumulation mechanisms of secondary metabolites, and updated systems, such as KEGG and KaPPA-View 4 KEGG for grape, can support such studies.

SummaryStrigolactones, first discovered as germination stimulants for parasitic weeds [1], are carotenoid-derived phytohormones that play major roles in inhibiting lateral bud outgrowth and promoting plant-mycorrhizal symbiosis [2–4]. Furthermore, strigolactones are involved in the regulation of lateral and adventitious root development, root cell division [5, 6], secondary growth [7], and leaf senescence [8]. Recently, we discovered the strigolactone transporter Petunia axillaris PLEIOTROPIC DRUG RESISTANCE 1 (PaPDR1), which is required for efficient mycorrhizal colonization and inhibition of lateral bud outgrowth [9]. However, how strigolactones are transported through the plant remained unknown. Here we show that PaPDR1 exhibits a cell-type-specific asymmetric localization in different root tissues. In root tips, PaPDR1 is co-expressed with the strigolactone biosynthetic gene DAD1 (CCD8), and it is localized at the apical membrane of root hypodermal cells, presumably mediating the shootward transport of strigolactone. Above the root tip, in the hypodermal passage cells that form gates for the entry of mycorrhizal fungi, PaPDR1 is present in the outer-lateral membrane, compatible with its postulated function as strigolactone exporter from root to soil. Transport studies are in line with our localization studies since (1) a papdr1 mutant displays impaired transport of strigolactones out of the root tip to the shoot as well as into the rhizosphere and (2) DAD1 expression and PIN1/PIN2 levels change in plants deregulated for PDR1 expression, suggestive of variations in endogenous strigolactone contents. In conclusion, our results indicate that the polar localizations of PaPDR1 mediate directional shootward strigolactone transport as well as localized exudation into the soil.

Decades of intensive research have led to the discovery of most plasma membrane ion channels and transporters and the characterization of their physiological functions. In contrast, although over 80% of transport processes occur inside the cells, the ion flux mechanisms across intracellular membranes (the endoplasmic reticulum, Golgi apparatus, endosomes, lysosomes, mitochondria, chloroplasts, and vacuoles) are difficult to investigate and remain poorly understood. Recent technical advances in super-resolution microscopy, organellar electrophysiology, organelle-targeted fluorescence imaging, and organelle proteomics have pushed a large step forward in the research of intracellular ion transport. Many new organellar channels are molecularly identified and electrophysiologically characterized. Additionally, molecular identification of many of these ion channels/transporters has made it possible to study their physiological functions by genetic and pharmacological means. For example, organellar channels have been shown to regulate important cellular processes such as programmed cell death and photosynthesis, and are involved in many different pathologies. This special issue (SI) on organellar channels and transporters aims to provide a forum to discuss the recent advances and to define the standard and open questions in this exciting and rapidly developing field. Along this line, a new Gordon Research Conference dedicated to the multidisciplinary study of intracellular membrane transport proteins will be launched this coming summer.

Biochemical and electrophysiological studies on plant vacuolar transporters became feasible in the late 1970s and early 1980s, when methods to isolate large quantities of intact vacuoles and purified vacuolar membrane vesicles were established. However, with the exception of the H+-ATPase and H+-PPase, which could be followed due to their hydrolytic activities, attempts to purify tonoplast transporters were for a long time not successful. Heterologous complementation, T-DNA insertion mutants, and later proteomic studies allowed the next steps, starting from the 1990s. Nowadays, our knowledge about vacuolar transporters has increased greatly. Nevertheless, there are several transporters of central importance that have still to be identified at the molecular level or have even not been characterized biochemically. Furthermore, our knowledge about regulation of the vacuolar transporters is very limited, and much work is needed to get a holistic view about the interplay of the vacuolar transportome. The huge amount of information generated during the last 35 years cannot be summarized in such a review. Therefore, I decided to concentrate on some aspects where we were involved during my research on vacuolar transporters, for some our laboratories contributed more, while others contributed less.

The exact transport characteristics of the vacuolar dicarboxylate transporter tDT from Arabidopsis are elusive. To overcome this limitation, we combined a range of experimental approaches comprising generation/analysis of tDT overexpressors, (13)CO(2) feeding and quantification of (13)C enrichment, functional characterization of tDT in proteoliposomes, and electrophysiological studies on vacuoles. tdt knockout plants showed decreased malate and increased citrate concentrations in leaves during the diurnal light-dark rhythm and after onset of drought, when compared with wildtypes. Interestingly, under the latter two conditions, tDT overexpressors exhibited malate and citrate levels opposite to tdt knockout plants. Highly purified tDT protein transports malate and citrate in a 1:1 antiport mode. The apparent affinity for malate decreased with decreasing pH, whereas citrate affinity increased. This observation indicates that tDT exhibits a preference for dianion substrates, which is supported by electrophysiological analysis on intact vacuoles. tDT also accepts fumarate and succinate as substrates, but not α-ketoglutarate, gluconate, sulfate, or phosphate. Taking tDT as an example, we demonstrated that it is possible to reconstitute a vacuolar metabolite transporter functionally in proteoliposomes. The displayed, so far unknown counterexchange properties of tDT now explain the frequently observed reciprocal concentration changes of malate and citrate in leaves from various plant species. tDT from Arabidopsis is the first member of the well-known and widely present SLC13 group of carrier proteins, exhibiting an antiport mode of transport.

Hormone transporters are crucial for plant hormone action, which is underlined by severe developmental and physiological impacts caused by their loss-of-function mutations. Here, we summarize recent knowledge on the individual roles of plant hormone transporters in local and long-distance transport. Our inventory reveals that many hormones are transported by members of distinct transporter classes, with an apparent dominance of the ATP-binding cassette (ABC) family and of the Nitrate transport1/Peptide transporter family (NPF). The current need to explore further hormone transporter regulation, their functional interaction, transport directionalities, and substrate specificities is briefly reviewed.

ABC transporters are essential for plant development, playing roles in processes such as gametogenesis, seed development, seed germination, organ formation, and secondary growth. ABC transporters are directly energized by ATP and can transport complex organic materials against concentration gradients; thus, they are uniquely suited to provide the complex building blocks required for the development of specialized plant cells. We review recent progress in our understanding of the contribution ABC transporters make to the growth and development of plants, including their roles in protective layer formation and in transporting phytohormones.

Plant pathogens cause huge yield losses. Plant defense often depends on toxic secondary metabolites that inhibit pathogen growth. Because most secondary metabolites are also toxic to the plant, specific transporters are needed to deliver them to the pathogens. To identify the transporters that function in plant defense, we screened Arabidopsis thaliana mutants of full-size ABCG transporters for hypersensitivity to sclareol, an antifungal compound. We found that atabcg34 mutants were hypersensitive to sclareol and to the necrotrophic fungi Alternaria brassicicola and Botrytis cinereaAtABCG34 expression was induced by Abrassicicola inoculation as well as by methyl-jasmonate, a defense-related phytohormone, and AtABCG34 was polarly localized at the external face of the plasma membrane of epidermal cells of leaves and roots. atabcg34 mutants secreted less camalexin, a major phytoalexin in Athaliana, whereas plants overexpressing AtABCG34 secreted more camalexin to the leaf surface and were more resistant to the pathogen. When treated with exogenous camalexin, atabcg34 mutants exhibited hypersensitivity, whereas BY2 cells expressing AtABCG34 exhibited improved resistance. Analyses of natural Arabidopsis accessions revealed that AtABCG34 contributes to the disease resistance in naturally occurring genetic variants, albeit to a small extent. Together, our data suggest that AtABCG34 mediates camalexin secretion to the leaf surface and thereby prevents Abrassicicola infection.

Stomatal pores are formed between a pair of guard cells and allow plant uptake of CO2 and water evaporation. Their aperture depends on changes in osmolyte concentration of guard cell vacuoles, specifically of K+ and Mal2−. Efflux of Mal2− from the vacuole is required for stomatal closure; however, it is not clear how the anion is released. Here, we report the identification of ALMT4 (ALUMINUM ACTIVATED MALATE TRANSPORTER4) as an Arabidopsis thaliana ion channel that can mediate Mal2− release from the vacuole and is required for stomatal closure in response to abscisic acid (ABA). Knockout mutants showed impaired stomatal closure in response to the drought stress hormone ABA and increased whole-plant wilting in response to drought and ABA. Electrophysiological data show that ALMT4 can mediate Mal2− efflux and that the channel activity is dependent on a phosphorylatable C-terminal serine. Dephosphomimetic mutants of ALMT4 S382 showed increased channel activity and Mal2− efflux. Reconstituting the active channel in almt4 mutants impaired growth and stomatal opening. Phosphomimetic mutants were electrically inactive and phenocopied the almt4 mutants. Surprisingly, S382 can be phosphorylated by mitogen-activated protein kinases in vitro. In brief, ALMT4 likely mediates Mal2− efflux during ABA-induced stomatal closure and its activity depends on phosphorylation.