Salicylic acid (SA) is central for the defense of plants to pathogens and abiotic stress. SA is synthesized in chloroplasts from chorismic acid by an isochorismate synthase (ICS1); SA biosynthesis is negatively regulated by autoinhibitory feedback at ICS1. Genetic studies indicated that the multidrug and toxin extrusion transporter ENHANCED DISEASE SUSCEPTIBILITY5 (EDS5) of Arabidopsis (Arabidopsis thaliana) is necessary for SA accumulation after biotic and abiotic stress, but so far it is not understood how EDS5 controls the biosynthesis of SA. Here, we show that EDS5 colocalizes with a marker of the chloroplast envelope and that EDS5 functions as a multidrug and toxin extrusion-like transporter in the export of SA from the chloroplast to the cytoplasm in Arabidopsis, where it controls the innate immune response. The location at the chloroplast envelope supports a model of the effect of EDS5 on SA biosynthesis: in the eds5 mutant, stress-induced SA is trapped in the chloroplast and inhibits its own accumulation by autoinhibitory feedback.

Plant architecture is influenced by the polar, cell-to-cell transport of auxin that is primarily provided and regulated by plasma membrane efflux catalysts of the PIN-FORMED and B family of ABC transporter (ABCB) classes. The latter were shown to require the functionality of the FK506 binding protein42 TWISTED DWARF1 (TWD1), although underlying mechanisms are unclear. By genetic manipulation of TWD1 expression, we show here that TWD1 affects shootward root auxin reflux and, thus, downstream developmental traits, such as epidermal twisting and gravitropism of the root. Using immunological assays, we demonstrate a predominant lateral, mainly outward-facing, plasma membrane location for TWD1 in the root epidermis characterized by the lateral marker ABC transporter G36/PLEIOTROPIC DRUG-RESISTANCE8/PENETRATION3. At these epidermal plasma membrane domains, TWD1 colocalizes with nonpolar ABCB1. In planta bioluminescence resonance energy transfer analysis was used to verify specific ABC transporter B1 (ABCB1)-TWD1 interaction. Our data support a model in which TWD1 promotes lateral ABCB-mediated auxin efflux via protein-protein interaction at the plasma membrane, minimizing reflux from the root apoplast into the cytoplasm.

The phytohormone auxin is critical for plant growth and many developmental processes. Members of the P-glycoprotein (PGP/ABCB) subfamily of ATP-binding cassette (ABC) transporters have been shown to function in the polar movement of auxin by transporting auxin over the plasma membrane in both monocots and dicots. Here, we characterize a new Arabidopsis member of the ABCB subfamily, ABCB21/PGP21, a close homolog of ABCB4, for which conflicting transport directionalities have been reported. ABCB21 is strongly expressed in the abaxial side of cotyledons and in junctions of lateral organs in the aerial part, whereas in roots it is specifically expressed in pericycle cells. Membrane fractionation by sucrose density gradient centrifugation followed by Western blot showed that ABCB21 is a plasma membrane-localized ABC transporter. A transport assay with Arabidopsis protoplasts suggested that ABCB21 was involved in IAA transport in an outward direction, while naphthalene acetic acid (NAA) was a less preferable substrate for ABCB21. Further functional analysis of ABCB21 using yeast import and export assays showed that ABCB21 mediates the 1-N-naphthylphthalamic acid (NPA)-sensitive translocation of auxin in an inward direction when the cytoplasmic IAA concentration is low, whereas this transporter mediates outward transport under high internal IAA. An increase in the cytoplasmic IAA concentration by pre-loading of IAA into yeast cells abolished the IAA uptake activity by ABCB21 as well as ABCB4. These findings suggest that ABCB21 functions as a facultative importer/exporter controlling auxin concentrations in plant cells.

Cyclophilin A is a conserved peptidyl-prolyl cis-trans isomerase (PPIase) best known as the cellular receptor of the immunosuppressant cyclosporine A. Despite significant effort, evidence of developmental functions of cyclophilin A in non-plant systems has remained obscure. Mutations in a tomato (Solanum lycopersicum) cyclophilin A ortholog, DIAGEOTROPICA (DGT), have been shown to abolish the organogenesis of lateral roots; however, a mechanistic explanation of the phenotype is lacking. Here, we show that the dgt mutant lacks auxin maxima relevant to priming and specification of lateral root founder cells. DGT is expressed in shoot and root, and localizes to both the nucleus and cytoplasm during lateral root organogenesis. Mutation of ENTIRE/ IAA9, a member of the auxin-responsive Aux/IAA protein family of transcriptional repressors, partially restores the inability of dgt to initiate lateral root primordia but not the primordia outgrowth. By comparison, grafting of a wild-type scion restores the process of lateral root formation, consistent with participation of a mobile signal. Antibodies do not detect movement of the DGT protein into the dgt rootstock; however, experiments with radiolabeled auxin and an auxin-specific microelectrode demonstrate abnormal auxin fluxes. Functional studies of DGT in heterologous yeast and tobacco-leaf auxin-transport systems demonstrate that DGT negatively regulates PIN-FORMED (PIN) auxin efflux transporters by affecting their plasma membrane localization. Studies in tomato support complex effects of the dgt mutation on PIN expression level, expression domain and plasma membrane localization. Our data demonstrate that DGT regulates auxin transport in lateral root formation.

We analyzed 314,254 soybean expressed sequence tags (ESTs), including 29,540 from our laboratory and 284,714 from GenBank. These ESTs were assembled into 56,147 unigenes. About 76.92% of the unigenes were homologous to genes from Arabidopsis thaliana ( Arabidopsis). The putative products of these unigenes were annotated according to their homology with the categorized proteins of Arabidopsis. Genes corresponding to cell growth and/or maintenance, enzymes and cell communication belonged to the slow-evolving class, whereas genes related to transcription regulation, cell, binding and death appeared to be fast-evolving. Soybean unigenes with no match to genes within the Arabidopsis genome were identified as soybean-specific genes. These genes were mainly involved in nodule development and the synthesis of seed storage proteins. In addition, we also identified 61 genes regulated by salicylic acid, 1,322 transcription factor genes and 326 disease resistance-like genes from soybean unigenes. SSR analysis showed that the soybean genome was more complex than the Arabidopsis and the Medicago truncatula genomes. GC content in soybean unigene sequences is similar to that in Arabidopsis and M. truncatula. Furthermore, the combined analysis of the EST database and the BAC-contig sequences revealed that the total gene number in the soybean genome is about 63,501.

Wood formation is a complex developmental process under multi-level transcriptional control executed by a large set of transcription factors. However, only limited members have been characterized to be key regulators of lignin biosynthesis in poplar. Here we report the conserved and unique functions of PtoMYB170, a transcription factor identified from Populus tomentosa (Chinese white poplar), in lignin deposition and drought tolerance in comparison with its duplicate paralog PtoMYB216. PtoMYB170 is preferentially expressed in young leaves and xylem tissues. Overexpression of PtoMYB170 in transgenic poplar plants resulted in stronger lignification and more thickened secondary wall in xylem compared with wild-type plants, whereas the CRISPR/Cas9-generated mutation of PtoMYB170 weakened lignin deposition, thereby leading to a more flexible and collapsed xylem phenotype. Transient expression experiments demonstrated that PtoMYB170 specifically activated the expression of lignin biosynthetic genes, consistent with the function of PtoMYB216. However, GUS staining assays revealed that PtoMYB170 was specifically expressed in guard cells of transgenic Arabidopsis while PtoMYB216 was not. Heterologous expression of PtoMYB170 in Arabidopsis enhanced stomatal closure in the dark and resulted in drought tolerance of the transgenic plants through reduced water loss, indicating a diversified role from PtoMYB216. These results revealed the PtoMYB170-dependent positive transcriptional regulation on lignin deposition in poplar and its coordinated function in enhancing drought tolerance by promoting dark-induced stomatal closure.

In the agriculture industry, adventitious root formation is a core issue of plants asexual propagation. However, the underlying molecular mechanism of adventitious root formation is far beyond understanding. In present study we found that target of rapamycin (TOR) signaling plays a key role in adventitious root formation in potato and Arabidopsis. The core components of TOR complex including TOR, RAPTOR, and LST8 are highly conserved in potato, but the seedlings of potato are insensitive to rapamycin, implying FK506 Binding Protein 12 KD (FKBP12) lost the function to bridge the interaction of rapamycin and TOR in potato. To dissect TOR signaling in potato, the rapamycin hypersensitive potato plants (BP12-OE) were engineered by introducing yeast FKBP12 (ScFKBP12) into potato. We found that rapamycin can significantly attenuate the capability of adventitious root formation in BP12-OE potatoes. KU63794 (KU, an active-site TOR inhibitor) combined with rapamycin can more significantly suppress adventitious root formation of BP12-OE potato than the single treatments, such as KU63794 or rapamycin, indicating its synergistic inhibitory effects on potato adventitious root formation. Furthermore, RNA-seq data showed that many genes associated with auxin signaling pathway were altered when BP12-OE potato seedlings were treated with rapamycin + KU, suggesting that TOR may play a major role in adventitious root formation via auxin signaling. The auxin receptor mutant tir1 was sensitive to TOR inhibitors and the double and quadruple mutants including tir1afb2, tir1afb3, and tir1afb1afb2afb3 displayed more sensitive to asTORis than single mutant tir1. Consistently, overexpression of AtTIR1 in Arabidopsis and potato can partially overcome the inhibitory effect of asTORis and promote adventitious root formation under asTORis treatments. These observations suggest that TOR signaling regulates adventitious root formation by mediating auxin signaling in Arabidopsis and potato.

SUMMARYPlant growth is achieved predominantly by cellular elongation, which is thought to be controlled on several levels by apoplastic auxin. Auxin export into the apoplast is achieved by plasma membrane efflux catalysts of the PIN-FORMED (PIN) and ATP-binding cassette protein subfamily B/phosphor-glycoprotein (ABCB/PGP) classes; the latter were shown to depend on interaction with the FKBP42, TWISTED DWARF1 (TWD1). Here by using a transgenic approach in combination with phenotypical, biochemical and cell biological analyses we demonstrate the importance of a putative C-terminal in-plane membrane anchor of TWD1 in the regulation of ABCB-mediated auxin transport. In contrast with dwarfed twd1 loss-of-function alleles, TWD1 gainof-function lines that lack a putative in-plane membrane anchor (HA-TWD1-C t ) show hypermorphic plant architecture, characterized by enhanced stem length and leaf surface but reduced shoot branching. Greater hypocotyl length is the result of enhanced cell elongation that correlates with reduced polar auxin transport capacity for HA-TWD1-C t . As a consequence, HA-TWD1-C t displays higher hypocotyl auxin accumulation, which is shown to result in elevated auxin-induced cell elongation rates. Our data highlight the importance of C-terminal membrane anchoring for TWD1 action, which is required for specific regulation of ABCB-mediated auxin transport. These data support a model in which TWD1 controls lateral ABCB1-mediated export into the apoplast, which is required for auxin-mediated cell elongation.