The large-scale untargeted proteomic and metabolomic studies were conducted in strawberry (Fragaria × ananassa) cv. Akihime fruit at five developmental stages. We found that some C6 volatiles highly contributed to the enrichment of volatiles at the red stage of strawberry fruit. We found that 12 genes involved in LOX pathway for volatile biosynthesis showed multiple patterns in protein abundance during fruit development and ripening, and 9 out of the 12 genes exhibited a significant increase in their relative expression levels at the red stage of fruit. We also found that the MYB9 gene (FaMYB9) expression level was positively correlated with the content of C6 volatiles (R = 0.989) and with the relative expression level and protein abundance of FaLOX5 at different strawberry fruit developmental stages (R = 0.954). The interaction between FaMYB9 and FaLOX5 was detected by yeast two-hybrid, co-immunoprecipitation (Co-IP), bimolecular fluorescence complementation (BiFC), and immunofluorescence (IF) analyses. Transient silencing of FaMYB9 delayed the fruit development and ripening, resulting in a significant decrease in the contents of C6 volatiles, while overexpression of FaMYB9 increased the fruit development and ripening and the contents of C6 volatiles in Akihime fruit. Therefore, FaMYB9 is positively involved in C6 volatile biosynthesis.

As the major nutritional component in soybean seeds storage proteins are initially synthesized on the endoplasmic reticulum as precursors and subsequently delivered to protein storage vacuoles (PSVs) via the Golgi-mediated pathway where they are converted into mature subunits and accumulated. However, the molecular machinery required for storage protein trafficking in soybean remains largely unknown. In this study, we cloned the sole soybean homolog of OsGPA3 that encodes a plant-unique kelch-repeat regulator of post-Golgi vesicular traffic for rice storage protein sorting. A complementation test showed that GmGPA3 could rescue the rice gpa3 mutant. Biochemical assays verified that GmGPA3 physically interacts with GmRab5 and its guanine exchange factor (GEF) GmVPS9. Expression of GmGPA3 had no obvious effect on the GEF activity of GmVPS9 toward GmRab5a. Notably, knock-down of GmGPA3 disrupted the trafficking of mmRFP-CT10 (an artificial cargo destined for PSVs) in developing soybean cotyledons. We identified two putative GmGPA3 interacting partners (GmGMG3 and GmGMG11) by screening a yeast cDNA library. Overexpression of GmGPA3 or GmGMG3 caused shrunken cotyledon cells. Our overall results suggested that GmGPA3 plays an important role in cell growth and development, in addition to its conserved role in mediating storage protein trafficking in soybean cotyledons.

When a plant is attacked by a pathogen, an immune response is activated to help protect it from harm. ERF transcription factors have been reported to regulate immune responses in plants. Here, three ERF transcription factors from Chinese wild Vitis quinquangularis, VqERF112, VqERF114 and VqERF072, are shown to respond to pathogen inoculation by powdery mildew, Pseudomonas syringae pv. tomato (Pst) DC3000 and Botrytis cinerea and to hormone treatments including with ET, SA, MeJA or ABA. Tissue specific expression analysis shows the highest expression levels of VqERF112 and VqERF114 were in mature berries and of VqERF072 was in tendrils. A GUS activity assay indicates that the promoters of VqERF112, VqERF114 and VqERF072 can be induced by powdery mildew inoculation and by hormone treatment, including with ET, SA and MeJA. Overexpression of VqERF112, VqERF114 and VqERF072 in transgenic Arabidopsis enhanced the resistance to Pseudomonas syringae pv. tomato DC3000 (Pst DC3000) and B. cinerea, and it increased the expression of the SA signaling-related genes AtNPR1 and AtPR1 and of the JA/ET signaling-related genes AtPDF1.2, AtLOX3, AtPR3 and AtPR4. Compared to Col-0 plants, the H2O2 accumulation in transgenic Arabidopsis increased after Pst DC3000 inoculation but decreased after B. cinerea inoculation. These results demonstrate that VqERF112, VqERF114 and VqERF072 positively regulate resistance to Pst DC3000 and B. cinerea.

Annexin, a multi-gene family in plants, is essential for plant growth and stress responses. Recent studies demonstrated a positive effect of annexin in abiotic stress responses. Interestingly, we found OsANN10, a putative annexin gene in rice, negatively regulated plant responses to osmotic stress. Knocking down OsANN10 significantly decreased the content of H2O2 by increasing Peroxidase (POD) and Catalase (CAT) activities, further reducing oxidative damage in rice leaves, suggesting a negative regulation of OsANN10 in protecting cell membrane against oxidative damage via scavenging ROS under osmotic stress.

Chrysanthemum is a typical short day (SD) flowering plant that requires a longer night period than a critical minimum duration to successfully flower. We identified FLOWERING LOCUS T-LIKE 3 (FTL3) and ANTI-FLORIGENIC FT/TFL1 FAMILY PROTEIN (AFT) as a florigen and antiflorigen, respectively, in a wild diploid chrysanthemum (Chrysanthemum seticuspe). Expression of the genes that produce these proteins, CsFTL3 and CsAFT, is induced in the leaves under SD or a noninductive photoperiod, respectively, and the balance between them determines the progression of floral transition and anthesis. However, how CsFTL3 and CsAFT are regulated to define the critical night length for flowering in chrysanthemum is unclear. In this study, we focused on the circadian clock-related gene GIGANTEA (GI) of C. seticuspe (CsGI) and generated transgenic C. seticuspe plants overexpressing CsGI (CsGI-OX). Under a strongly inductive SD (8 L/16D) photoperiod, floral transition occurred at almost the same time in both wild-type and CsGI-OX plants. However, under a moderately inductive (12 L/12D) photoperiod, the floral transition in CsGI-OX plants was strongly suppressed, suggesting that the critical night length for flowering was lengthened for CsGI-OX plants. Under the 12 L/12D photoperiod, CsAFT was upregulated in CsGI-OX plants. Giving a night break (NB) 10 h after dusk was the most effective time to inhibit flowering in wild-type plants, while the most effective time for NB was extended to dawn (12 and 14 h after dusk) in CsGI-OX plants. In wild-type plants, a red-light pulse delivered 8 or 10 h after dusk induced maximal CsAFT expression, but the length of the time period over which CsAFT could be induced by red light was extended until subjective dawn in CsGI-OX plants. Therefore, CsGI-OX plants required a longer dark period to maintain lower levels of CsAFT, and their critical night length for flowering was thus lengthened. These results suggested that CsGI has an important role in the control of photoperiodic flowering through shaping the gate for CsAFT induction by light in chrysanthemum.

Pear is one of the most commercially important fruit trees worldwide and is widely cultivated in temperate zones. Drought stress can greatly limit pear fruit yield and quality. Pyrus betulaefolia Bunge, a drought-resistant pear rootstock that is commonly used in northern China, confers favourable characteristics to pear scions, allowing them to respond rapidly to drought stress via the transport of macromolecules such as phloem-mobile mRNAs. How drought-responsive mRNAs function as phloem-mobile signals remains unknown, however. Here, we used RNA sequencing (RNA-seq) combined with SNP analysis to identify mobile mRNAs in P. betulaefolia. We focused on mobile mRNAs that respond to drought stress and found that the abundance of a novel mRNA named PbDRM (P. betulaefolia drought-responsive mobile gene) significantly increased in several different scion cultivars when they were grafted onto P. betulaefolia rootstock under drought conditions. In addition, downregulating PbDRM by virus-induced gene silencing (VIGS) increased the drought sensitivity of P. betulaefolia. CAPS RT-PCR analysis confirmed that PbDRM mRNA moves from rootstock to scion in micrografting systems. Therefore, PbDRM mRNA acts as a phloem-mobile signal in pear under drought stress.

Azospirillum brasilense colonizes plant roots and improves productivity, but the molecular mechanisms behind its phytostimulation properties remain mostly unknown. Here, we uncover an important role of TARGET OF RAPAMYCIN (TOR) signaling on the response of Arabidopsis thaliana to A. brasilense Sp245. The effect of the bacterium on TOR expression was analyzed in the transgenic line TOR/tor-1, which carries a translational fusion with the GUS reporter protein, and the activity of TOR was assayed thought the phosphorylation of its downstream signaling target S6K protein. Besides, the role of TOR on plant growth in inoculated plants was assessed using the ATP-competitive inhibitor AZD-8055. A decrease in growth of the primary root correlates with an improved branching and absorptive capacity via lateral root and root hair proliferation 6 days after transplant to different concentrations of the bacterium (103 or 105 CFU/mL). Bacterization increased the expression of TOR in shoot and root apexes and promoted phosphorylation of S6K 3 days after transplant. The TOR inhibitor AZD-8055 (1 µM) inhibited plant growth and cell division in root meristems and in lateral root primordia, interfering with the phytostimulation by A. brasilense. In addition, the role of auxin produced by the bacterium to stimulate TOR expression was explored. Noteworthy, the A. brasilense mutant FAJ0009, impaired in auxin production, was unable to elicit TOR signaling to the level observed for the wild-type strain, showing the importance of this phyhormone to stimulate TOR signaling. Together, our findings establish an important role of TOR signaling for the probiotic traits elicited by A. brasilense in A. thaliana.

Extreme heat events will challenge agricultural production and raise the risk of food insecurity. California is the largest agricultural producer in the United States, and climate change and extreme heat may significantly affect the state’s food production. This paper provides a summary of the current literature on crop responses to extreme heat, with a focus on perennial agriculture in California. We highlight contemporary trends and future projections in heat extremes, and the range of plant responses to extreme heat exposure, noting the variability in plant tolerance and response across season, crop, and cultivar. We also review practices employed to mitigate heat damage and the capacity for those practices to serve as adaptation options in a warmer and drier future. Finally, we discuss current and future research directions aimed at increasing the adaptive capacity of perennial agriculture to the increased heat exposure anticipated with climate change. Collectively, the literature reviewed makes clear the need to understand crop responses and tolerances to heat within the context of climate change and climate extremes in order to sustain crop production, preserve agricultural communities, and bolster food security at local, national, and global scales.

The word phenotyping can nowadays invoke visions of a drone or phenocart moving swiftly across research plots collecting high-resolution data sets on a wide array of traits. This has been made possible by recent advances in sensor technology and data processing. Nonetheless, more comprehensive often destructive phenotyping still has much to offer in breeding as well as research. This review considers the ‘breeder friendliness’ of phenotyping within three main domains: (i) the ‘minimum data set’, where being ‘handy’ or accessible and easy to collect and use is paramount, visual assessment often being preferred; (ii) the high throughput phenotyping (HTP), relatively new for most breeders, and requiring significantly greater investment with technical hurdles for implementation and a steeper learning curve than the minimum data set; (iii) detailed characterization or ‘precision’ phenotyping, typically customized for a set of traits associated with a target environment and requiring significant time and resources. While having been the subject of debate in the past, extra investment for phenotyping is becoming more accepted to capitalize on recent developments in crop genomics and prediction models, that can be built from the high-throughput and detailed precision phenotypes. This review considers different contexts for phenotyping, including breeding, exploration of genetic resources, parent building and translational research to deliver other new breeding resources, and how the different categories of phenotyping listed above apply to each. Some of the same tools and rules of thumb apply equally well to phenotyping for genetic analysis of complex traits and gene discovery.

Anthropogenic increases in fossil fuel emissions have been a primary driver of increased concentrations of atmospheric carbon dioxide ([CO2]) and other greenhouse gases resulting in warmer temperatures, alterations in precipitation patterns, and increased occurrence of extreme weather events in terrestrial areas across the globe. In agricultural growing regions, alterations in climate can challenge plant productivity in ways that impact the ability of the world to sustain adequate food production for a growing and increasingly affluent population with shifting access to affordable and nutritious food. While the knowledge gap that exists regarding potential climate change impacts is large across agriculture, it is especially large in specialty cropping systems. This includes fruit and vegetable crops, and perennial cropping systems which also contribute (along with row crops) to our global diet. In order to obtain a comprehensive view of the true impact of climate change on our global food supply, we must expand our narrow focus from improving yield and plant productivity to include the impact of climate change on the nutritional value of these crops. In order to address these questions, we need a multi-faceted approach that integrates physiology and genomics tools and conducts comprehensive experiments under realistic depictions of future projected climate. This review describes gaps in our knowledge in relation to these responses, and future questions and actions that are needed to develop a sustainable future food supply in light of global climate change.

The flexible development of plants is characterized by a high capacity for post-embryonic organ formation and tissue regeneration, processes, which require tightly regulated intercellular communication and coordinated tissue (re-)polarization. The phytohormone auxin, the main driver for these processes, is able to establish polarized auxin transport channels, which are characterized by the expression and polar, subcellular localization of the PIN1 auxin transport proteins. These channels are demarcating the position of future vascular strands necessary for organ formation and tissue regeneration. Major progress has been made in the last years to understand how PINs can change their polarity in different contexts and thus guide auxin flow through the plant. However, it still remains elusive how auxin mediates the establishment of auxin conducting channels and the formation of vascular tissue and which cellular processes are involved. By the means of sophisticated regeneration experiments combined with local auxin applications in Arabidopsis thaliana inflorescence stems we show that (i) PIN subcellular dynamics, (ii) PIN internalization by clathrin-mediated trafficking and (iii) an intact actin cytoskeleton required for post-endocytic trafficking are indispensable for auxin channel formation, de novo vascular formation and vascular regeneration after wounding. These observations provide novel insights into cellular mechanism of coordinated tissue polarization during auxin canalization.

In Brassica napus, pod number and pod density are critical factors to determine seed yield. Although the pod density is an essential yield trait, the regulation of yield formation in oil crops, as well as the genetic and molecular mechanisms, are poorly understood. In this study, we characterized a rapeseed high-density pod mutant (dpt247) from composite hybridization. To shed some light on the nature of this mutation, it was investigated morphologically, anatomically, physiologically, genetically and transcriptomically. The mutant plant showed noticeable phenotypic differences in comparison with the control plant, including reduced plant height and primary branch length, decreased number of primary branches, significantly increased number of pod on the main inflorescence, and more compact pod distribution. Besides, the mutant had higher levels of indole-3-acetic acid (IAA) and zeatin riboside (ZR) in the shoot apical meristem (SAM). The dense pod trait was controlled by two major recessive genes identified in the segregating genetic populations of GRE501 and dpt247. RNA sequencing indicated genes participated in auxin, cytokinin and WUS/CLV signalling pathway in dpt247 were more active in the mutant. These results provide important information for understanding the regulation of yield formation and high yield breeding in rapeseed.

Ultraviolet-B (UV-B) radiation promotes anthocyanin synthesis in many plants. Although several transcription factors promote anthocyanin synthesis in response to UV-B radiation, the underlying mechanism remains unclear. In this study, the MdWRKY72 transcription factor gene was isolated from the ‘Taishanzaoxia’ apple genome. Quantitative real-time PCR analyses revealed that the genes encoding enzymes and transcription factors involved in the anthocyanin synthesis pathway (MdANS, MdDFR, MdUFGT, and MdMYB1) were more highly expressed in MdWRKY72-overexpressing transgenic calli than in the wild-type ‘Orin’ apple calli. The results indicated that MdWRKY72 increases anthocyanin synthesis in transgenic calli exposed to UV-B radiation. The results of a gel shift assay and chromatin immunoprecipitation proved that MdWRKY72 promotes MdMYB1 expression indirectly by binding to a W-box element in the MdHY5 promoter and directly by binding to a W-box element in the MdMYB1 promoter. Thus, MdWRKY72 increases anthocyanin synthesis via direct and indirect mechanisms. These findings may be useful for elucidating the molecular mechanism underlying UV-B-induced anthocyanin synthesis mediated by MdWRKY72.

Protein N-glycosylation plays key roles in protein folding, stability, solubility, biogenesis, and enzyme activity. Tomato (Solanum lycopersicum L.) is an important vegetable crop with abundant nutritional value, and the formation of tomato fruit qualities primarily occurs in the fruit ripening process. However, a large number of N-glycosylation-mediated mechanisms in regulating tomato fruit ripening have not been elucidated to date. In this study, western blot assays showed that the extents of mature N-glycoproteins were differentially expressed in mature green fruits (fruit start ripening) and ripe fruits (fruit stop ripening). Next, through performing a comparative N-glycoproteome analysis strategy, a total of 553 N-glycosites from 363 N-glycoproteins were identified in mature green fruits compared with ripe fruits. Among them, 252 N-glycosites from 191 N-glycoproteins were differentially expressed in mature green fruits compared with ripe fruits. The differentially expressed N-glycoproteins were mainly located in the chloroplast (30%) and cytoplasm (16%). Gene Ontology (GO) analysis showed that these N-glycoproteins were involved in various biological processes, cellular components and molecular functions. These N-glycoproteins participate in biological processes, such as metabolic processes, cellular processes and single-organism processes. These N-glycoproteins are also cellular components in biological process cells, membranes and organelles and have different molecular functions, such as catalytic activity and binding. Notably, these N-glycoproteins were enriched in starch and sucrose metabolism and galactose metabolism by KEGG pathway analysis. This community resource regarding N-glycoproteins is the first large-scale N-glycoproteome during plant fruit ripening. This study will contribute to understanding the function of N-glycosylation in regulating plant fruit ripening.

Plants respond and adapt to changes in their environment by employing a wide variety of genetic, molecular, and biochemical mechanisms. When so doing, they trigger large-scale rearrangements at the metabolic and transcriptional levels. The dynamics and patterns of these rearrangements and how they govern a stress response is not clear. In this opinion, we discuss a plant’s response to stress from the perspective of the metabolic gene co-expression network and its rearrangement upon stress. As a case study, we use publicly available expression data of Arabidopsis thaliana plants exposed to heat and drought stress to evaluate and compare the co-expression networks of metabolic genes. The analysis highlights that stress conditions can lead to metabolic tightening and expansion of the co-expression network. We argue that this rearrangement could play a role in a plant’s response to stress and thus may be an additional tool to assess and understand stress tolerance/sensitivity. Additional studies are needed to evaluate the metabolic network in response to multiple stresses at various intensities and across different genetic backgrounds (e.g., intra- and inter-species, sensitive and tolerant eco/genotypes).

The Geminiviridae family is one of the most successful and largest families of plant viruses that infect a large variety of important dicotyledonous and monocotyledonous crops and cause significant yield losses worldwide. This broad spectrum of host range is only possible because geminiviruses have evolved sophisticated strategies to overcome the arsenal of antiviral defenses in such diverse plant species. In addition, geminiviruses evolve rapidly through recombination and pseudo-recombination to naturally create a great diversity of virus species with divergent genome sequences giving the virus an advantage over the host recognition system. Therefore, it is not surprising that efficient molecular strategies to combat geminivirus infection under open field conditions have not been fully addressed. In this review, we present the anti-geminiviral arsenal of plant defenses, the evolved virulence strategies of geminiviruses to overcome these plant defenses and the most recent strategies that have been engineered for transgenic resistance. Although, the in vitro reactivation of suppressed natural defenses as well as the use of RNAi and CRISPR/Cas systems hold the potential for achieving broad-range resistance and/or immunity, potential drawbacks have been associated with each case.

• Transcription factor (TF) gene clusters in plants, such as tomato, potato, petunia, tobacco, and almond, have been characterized for their roles in the biosynthesis of diverse array of specialized metabolites. In Catharanthus roseus, three AP2/ERF TFs, ORCA3, ORCA4, and ORCA5, have been shown to be present on the same genomic scaffold, forming a cluster that regulates the biosynthesis of pharmaceutically important terpenoid indole alkaloids (TIAs). Our analysis of the recently updated C. roseus genome sequence revealed that the ORCA cluster comprises two additional AP2/ERFs, the previously characterized ORCA2 and a newly identified member designated as ORCA6. Transcriptomic analysis revealed that the ORCAs are highly expressed in stems, followed by leaves, roots and flowers. Expression of ORCAs was differentially induced in response to methyl-jasmonate and ethylene treatment. In addition, ORCA6 activated the strictosidine synthase (STR) promoter in tobacco cells. Activation of the STR promoter was significantly higher when ORCA2 or ORCA6 was coexpressed with the mitogen-activated protein kinase kinase, CrMPKK1. Furthermore, transient overexpression of ORCA6 in C. roseus flower petals activated TIA pathway gene expression and TIA accumulation. The results described here advance our understanding of regulation of TIA pathway by the ORCA gene cluster and the evolution for plant ERF gene clusters.

Leaf color mutants are an ideal tool to study chlorophyll biosynthesis, chloroplast development and photosynthesis. In this study, we identified an EMS-induced yellow young leaf mutant C777. The mutant exhibited yellow cotyledons and emerging true leaves with stay-green dots that turn green gradually with leaf growth. Segregation analysis in several populations indicated that the mutant C777 was controlled by a recessive gene yyl-1. Fine mapping delimited the yyl-1 locus to a 45.3 kb region harboring 8 putative genes, but only one SNP (G to A) was identified between C777 and its wild-type parental line in this region which occurred in the 13th exon of CsHD that encodes a histidine and aspartic acid (HD) domain containing protein. This nonsense mutation introduced a stop codon and thus a premature protein. Uniqueness of this mutant allele was verified in 515 cucumber lines. Quantitative real-time PCR revealed significantly reduced expression of CsHD gene in the mutant. Further, silencing the NbHD gene by VIGS in tobacco resulted in virescent young leaves and significantly down-regulated expression of HD gene. These results strongly supported the association of the CsHD gene with the virescent young leaf phenotype in C777. This is the first report to clone and characterize the CsHD gene in the horticultural crops. The results may help understand the functions of the HD gene in chloroplast development and chlorophyll biosynthesis in plants.

Drought is an environmental stress that severely affects plant growth and crop production. Different studies have focused on drought responses but the molecular bases that regulate these mechanisms are still unclear. We report the participation of Aspartic Protease (APA1) in drought tolerance. Overexpressing APA1 Arabidopsis plants (OE-APA1), showed a phenotype more tolerant to drought compared with WT. On the contrary, apa1 insertional lines were more sensitive to this stress compared to WT plants. Morphological and physiological differences related with the water loss were observed between leaves of OE- APA1 and WT plants. OE-APA1 leaves showed lower stomata index and stomata density as well as a smaller of the stomatic aperture compared to WT plants. qPCR analysis in OE-APA1 leaves, showed higher expression levels of genes related to ABA signaling and synthesis. Analysis of plant lines expressing APA1 promoter fused to GUS showed that APA1 is expressed in epidermal and stomata cells. In summary, this work suggests that APA1 is involved in ABA-dependent response that its overexpression confers drought tolerance in Arabidopsis.

As a group of plant-specific proteins, OVATE family protein (OFP) members have been shown to function as transcriptional repressors and involve in plant growth regulation in Arabidopsis and rice. It has also been shown that OFPs can interact with TONNEAU1 Recruiting Motif (TRM) proteins to regulate tomato fruit shape. In this study, we show that mutant plants with knock-down expression of OFP1, OFP2, OFP3, and OFP5 exhibit longer hypocotyls and cotyledons due to enhanced cell elongation. Overexpression of OFPs disturb the arrangement of cortical microtubule arrays in pavement cells and promote abnormal pavement cell expansion perpendicular to the direction of petiole growth, resulting in the kidney-shaped cotyledons in transgenic plants. OFP2 and OFP5 interact directly with the microtubule regulating protein TONNEAU2 (TON2), and genetic analysis suggests TON2 is required for the function of OFPs. We also show that alter the expression of OFPs affects light and BR regulated microtubule re-orientation. BR treatment reduce the protein accumulation of OFP2, suggesting OFP2 mediates BR regulated microtubule reorientation. Taken together, our study provides evidences showing that OFP family proteins negatively regulate cell expansion by modulating microtubule reorganization, which requires the function of TON2.

Protease inhibitors (PIs) are regulatory proteins found in numerous animal tissues and fluids, plants, and microorganisms that reduce and inhibit the exacerbated and uncontrolled activity of the target proteases. Specific PIs are also effective tools for inactivating proteases involved in human diseases like arthritis, pancreatitis, hepatitis, cancer, AIDS, thrombosis, emphysema, hypertension, and muscular dystrophy among others. Plant PIs—small peptides with a high content of cystine residues in disulfide bridges—possess a remarkable resistance to heat treatment and a high stability against shifts in pH, denaturing agents, ionic strength, and proteolysis. In recent years, novel biologic activities have been reported for plant PIs, including antimicrobial, anticoagulant, antioxidant action plus inhibition of tumor-cell growth; thus pointing to possible applications in medicine, agriculture, and biotechnology. In this review, we provide a comparative overview of plant-PIs classifying them in four groups according of their thermal and pH stability (high stability and hyperstable -to temperature and to pHs-, respectively), then emphasizing the relevance of the physicochemical characteristics of these proteins for potential biotechnological and industrial applications. Finally, we analyze the biologic activities of the stable protease inhibitors previously characterized that are the most relevant to potential applications in biomedicine, the food industry, and agriculture.

Furanocoumarins are defense molecules mainly described in four plant families that are phylogenetically distant. Molecular characterization of the biosynthetic pathway has been started for many years in Apiaceae and Rutaceae. The results obtained thus far in Apiaceae indicated a major role of cytochromes P450 (P450 s) in the CYP71 family. In the present work, we describe the importance of another subfamily of P450 s, CYP82D, identified by using a deep analysis of the citrus (Rutaceae) genome and microarray database. CYP82D64 is able to hydroxylate xanthotoxin to generate 5-OH-xanthotoxin. Minor and limited amino acid changes in the CYP82D64 coding sequence between Citrus paradisi and Citrus hystrix provide the enzyme in the latter with the ability to hydroxylate herniarin, but with low efficiency. The kinetic constants of the enzyme are consistent with those of other enzymes of this type in plants and indicate that it may be the physiological substrate. The activity of the enzyme is identical to that of CYP71AZ6 identified in parsnip, showing possible evolutionary convergence between these two families of plants. It is highly possible that these molecules are derived from the synthesis of ubiquitous coumarins throughout the plant kingdom.

Pectate lyases (PELs) play important roles in plant growth and development, mainly by degrading the pectin in primary cell walls. However, the role of PELs in cotton fiber elongation, which also involves changes in cellular structure and components, is poorly understood. Therefore, we aimed to isolate and characterize GhPEL76, as we suspected it to contribute to the regulation of fiber elongation. Expression analysis (qRT-PCR) revealed that GhPEL76 is predominately expressed in cotton fiber, with significantly different expression levels in long- and short-fiber cultivars, and that GhPEL76 expression is responsive to gibberellic acid and indoleacetic acid treatment. Furthermore, GhPEL76 promoter-driven β-glucuronidase activity was detected in the roots, hypocotyls, and leaves of transgenic Arabidopsis plants, and the overexpression of GhPEL76 in transgenic Arabidopsis promoted the elongation of several organs, including petioles, hypocotyls, primary roots, and trichomes. Additionally, the virus-induced silencing of GhPEL76 in cotton reduced fiber length, and both yeast one-hybrid and transient dual-luciferase assays suggested that GhbHLH13, a bHLH transcription factor that is up-regulated during fiber elongation, activates GhPEL76 expression by binding to the G-box of the GhPEL76 promoter region. Therefore, these results suggest GhPEL76 positively regulates fiber elongation and provide a basis for future studies of cotton fiber development.

The nutritional value of a crop lies not only in its protein, lipid, and sugar content but also involves compounds such as the antioxidants lycopene, β-carotene and vitamin C. In the present study, wild tomato Solanum pimpinellifolium LA 1589 was assessed for its potential to improve antioxidant content. This wild species was found to be a good source of alleles for increasing β-carotene, lycopene, vitamin C and vitamin E contents in cultivated tomato. Characterization of an LA 1589 interspecific inbred backcross line (IBL) mapping population revealed many individuals with transgressive segregation for the antioxidants confirming the usefulness of this wild species for breeding of these traits. Molecular markers were used to identify QTLs for the metabolites in the IBL population. In total, 64 QTLs were identified for the antioxidants and their locations were compared to the map positions of previously identified QTLs for confirmation. Four (57%) of the carotenoid QTLs, four (36%) of the vitamin QTLs, and 11 (25%) of the phenolic acid QTLs were supported by previous studies. Furthermore, several potential candidate genes were identified for vitamins C and E and phenolic acids loci. These candidate genes might be used as markers in breeding programs to increase tomato’s antioxidant content.

MicroRNA (miRNA)-mediated post-transcriptional regulation plays a vital role in the response of plants to pathogens. Although the microRNA397 family has been implicated in physiological processes as an important regulator, little is known about its function in the resistance of plants to pathogens. Here, Malus hupehensis miR397, which was induced by Botryosphaeria dothidea infection, was identified to directly target M. hupehensis Laccase7 (MhLAC7). The expression analysis of mature Mh-miR397 and MhLAC7 revealed their partly opposite expression patterns. The coexpression of Mh-miR397b in MhLAC7 overexpressing Nicotiana benthamiana suppressed the accumulation of exogenous MhLAC7 and endogenous NbLAC7, which led to decreased lignin content and reduced plant resistance to Botrytis cinerea. As reflected by increasing disease severity and pathogen growth, overexpression of miR397b in both the resistant M. hupehensis and susceptible M. domestica ‘Gala’ resulted in an increased sensitivity to B. dothidea infection, owing to reduced LAC7 expression and lignin content; however, the inhibition of miR397 had opposite effects. MicroRNA397 functions as a negative regulator in the resistance of Malus to B. dothidea by modulating the LAC7 expression and lignin biosynthesis.

The plasma membrane (PM)-localized receptor-like kinases (RLKs) play important roles in pathogen defense. One of the first cloned RLKs is the Arabidopsis receptor kinase FLAGELLIN SENSING 2 (FLS2), which specifically recognizes a conserved 22 amino acid N-terminal sequence of Pseudomonas syringae pv．tomato DC3000 (Pst) flagellin protein (flg22). Although extensively studied in Arabidopsis, the functions of RLKs in crop plants remain largely uninvestigated. To understand the roles of RLKs in soybean (Glycine max), GmFLS2 was silenced via virus induced gene silencing (VIGS) mediated by Bean pod mottle virus (BPMV). No significant morphological differences were observed between GmFLS2-silenced plants and the vector control plants. However, silencing GmFLS2 significantly enhanced the susceptibility of the soybean plants to Pseudomonas syringae pv．glycinea (Psg). Kinase activity assay showed that silencing GmFLS2 significantly reduced the phosphorylation level of GmMPK6 in response to flg22 treatment. However, reduced phosphorylation level of both GmMPK3 and GmMPK6 in response to Psg infection was observed in GmFLS2-silenced plants, implying that defense response is likely transduced through activation of the downstream GmMAPK signaling pathway upon recognition of bacterial pathogen by GmFLS2. The core peptides of flg22 from Pst and Psg were highly conserved and only 4 amino acid differences were seen at their N-termini. Interestingly, it appeared that the Psg-flg22 was more effective in activating soybean MAPKs than activating Arabidopsis MAPKs, and conversely, Pst-flg22 was more effective in activating Arabidopsis MAPKs than activating soybean MAPKs, suggesting that the cognate recognition is more potent than heterologous recognition in activating downstream signaling. Taken together, our results suggest that the function of FLS2 is conserved in immunity against bacteria pathogens across different plant species.

Low freezing tolerance reduces the persistence of red clover under northern climate. The incidence of winter damages in perennial crops could increase in the future due to the adverse effects of the predicted warmer fall temperature on plant cold acclimation. To accelerate breeding progress, two cultivars of red clover Christie (C-TF0) and Endure (E-TF0) were exposed to a recurrent selection protocol for freezing tolerance performed indoor. New populations were obtained after five (C-TF5 and E-TF5), six (C-TF6 and E-TF6), and seven (C-TF7 and E-TF7) cycles of recurrent selection. These populations were overwintered under natural conditions and monitored for freezing tolerance and cold-induced molecular traits. Freezing tolerance was improved by up to 6 °C in recurrently selected populations when compared to initial cultivars confirming that further progress are achieved with advanced cycles of selection. Monthly analysis of biochemical changes shows that higher starch concentrations at the onset of the fall hardening period are contributing to the acquisition of superior freezing tolerance through its impact on sucrose accumulation. They also contribute to the vigor of spring regrowth by sustaining more pinitol and proline synthesis. Larger concentrations of these metabolites in populations with higher levels of freezing tolerance (TF7) hint at their involvement in winter survival of red clover. Among genes differentially expressed in response to both cold acclimation and recurrent selection, a concomitant cold induction of APPR9 and cold repression of 1-aminocyclopropane-carboxylate synthase suggests a link between the repression of a pathway regulated by ethylene and the improvement of freezing tolerance in red clover.

The cyclic electron flow (CEF) around photosystem I (PSI) plays a crucial role in photosynthesis and also functions in plant tolerance of abiotic environmental stress. However, the role of PGR5/PGRL1- and NDH-dependent CEF in tomato under hightemperature (HT) is poorly understood. Here, we assessed the photoprotective effect of these pathways in tomato leaves under HT by using antimycin A (AA) and rotenone (R), which are chemical inhibitors of PGR5/PGRL1- and NDH-dependent CEF, respectively. The results showed that AA treatment caused significantly greater inhibition of CEF under HT compared to R treatment. Moreover, AA treatment caused a greater decrease in maximal photochemistry efficiency (Fv/Fm) and increased damage to the donor and acceptor side of photosystem II (PSII); however, the limitation of the acceptor side in PSI [Y(NA)] was significantly increased. In addition, thylakoid membrane integrity was compromised and reactive oxygen species, proton gradient (ΔpH), antioxidant enzyme activity, and the expression of photosystem core subunit genes were significantly decreased under AA treatment. These findings indicate that PGR5/PGRL1-dependent CEF protects PSII and PSI from photooxidative damage through the formation of ΔpH while maintaining thylakoid membrane integrity and normal gene expression levels of core photosystem components. This study demonstrates that PGR5/PGRL1-dependent CEF plays a major role in HT response in tomato.

Root-derived abscisic acid (ABA) is known to regulate shoot physiology, such as stomata closure. Conversely, the basipetal regulatory effect of shoot-derived ABA is poorly understood. Herein, we report that simulation of shoot-ABA accumulation by exogenous application of ABA to shoots basipetally stimulates primary root (PR) growth. ABA applied to shoots accelerates root cell division, as evidenced by the increase in meristem size and cell number and the intensity of CYCB1;1::GFP (a mitosis marker). Root ABA content was not changed following shoot ABA application, although the ABA reporter line RAB18::GFP showed an increase in ABA in the cotyledons. Shoot-ABA application increases basipetal auxin transport by 114%. Shoot-ABA-promoted PR growth can be abolished by attenuating basipetal auxin flux using 2,3,5-triiodobenzoic acid (TIBA, an auxin transport inhibitor), demonstrating that ABA promotes PR growth by increasing basipetal auxin transport. Root cell elongation, evaluated by the total length of the first 7 cells in the elongation zone (EZ), was increased by 56% following shoot-ABA application. The cell walls of the root EZ were alkalinized by ABA, as exhibited by 8-hydroxypyrene-1,3,6-trisulfonic acid trisodium salt staining. Higher pH promotes both PR growth and cell elongation. Thus, shoot-ABA promotes cell elongation by alkalinizing the cell wall. In light of our results, we provide a representative detailed model of the basipetal regulatory effect of ABA on PR growth.

Water deficits are a major constraint on maize growth and yield, and deep roots are one of the major mechanisms of drought tolerance. In this study, four root and shoot traits were evaluated within an association panel consisting of 209 diverse maize accessions under well-watered (WW) and water-stressed (WS) conditions. A significant positive correlation was observed between seminal root length (SRL) under WS treatment and the drought tolerance index (DI) of maize seedlings. The transcriptome profiles of maize seminal roots were compared between four drought-tolerant lines and four drought-sensitive lines under both water conditions to identify genes associated with the drought stress response. After drought stress, 343 and 177 common differentially expressed genes (DEGs) were identified in the drought-tolerant group and drought-sensitive group, respectively. In parallel, a coexpression network underlying SRL was constructed on the basis of transcriptome data, and 10 hub genes involved in two significant associated modules were identified. Additionally, a genome-wide association study (GWAS) of the SRL revealed 62 loci for the two water treatments. By integrating the results of the GWAS, the common DEGs and the coexpression network analysis, 7 promising candidate genes were prioritized for further research. Together, our results provide a foundation for the enhanced understanding of seminal root changes in response to drought stress in maize.

Recent progress in "omics" methodologies allow us to gain insight into the complex molecular regulatory networks underlying plant responses to environmental stresses. Among the different genome-wide analysis, epigenomics is the most under-investigated “omic” approach requiring more critical and speculative discussion about approaches, methods and experimental designs. Epigenomics allows us to gain insight into the molecular adaptation of plants in response to environmental stresses. The identification of epigenetic marks transmitted during filial generations enables new theories to be developed on the evolution of living organisms in relation to environmental changes. The molecular mechanisms driving the capacity of plants to memorize a stress and to generate stress-resistant progenies are still unclear and scarcely investigated. The elucidation of these cryptic molecular switches will assist breeders in designing crops characterized by minimally compromised productivity in relation to stresses caused by climate change. The aim of this review is to briefly describe the most uptodate epigenomic approaches, update recent progresses in crop epigenomics in plant stress biology, and to stimulate the discussion of new epigenomic methods and approaches in the new era of “omic” sciences.

Salicylic acid (SA) plays an important role in the response of plants to abiotic stresses. Starvation stress affects plant cell metabolic activities, which further limits the normal growth and development of plants. It was reported that SA might play a regulatory role in the process of plant against starvation stress, but the mechanism involved in this process is still unclear. Thus, in this study, the transgenic plants overexpressing a SA binding protein 2 (SABP2) gene were exposed to starvation stress and the transgenic lines showed starvation-tolerant phenotype. Compared with wild-type (WT) plants, transgenic plants showed better growth status under poor-nutrition stress. Transgenic plants also showed more vigorous roots than WT plants. Physiological tests indicated that the transgenic plants showed higher relative water content (RWC), chlorophyll content, photosynthetic capacity, endogenous SA content, and lower ROS level compared to WT plants. Transcriptome analysis of tobacco plants identified 3, 748 differentially expressed genes (DEGs) between transgenic and WT plants under starvation stress. These DEGs are mainly involved in glycolysis/gluconeogenesis pathway group, MAPK signaling pathway group and plant hormone signal transduction pathway group. As determined by qPCR, up-regulated expression of fifteen genes such as abscisic acid receptor PYR1-like gene (NtPYR1-like), bidirectional sugar transporter N3-like gene (NtSWEETN3-like) and superoxide dismutase [Fe] chloroplastic-like gene (NtFeSOD-like), etc., was observed in transgenic plants under poor-nutrition stress which was in accordance with RNA-sequencing results. The modified pathways involved in plant hormone signaling are thought to be at least one of the main causes of the increased starvation tolerance of transgenic tobacco plants with altered SA homeostasis.

Symbiotic nitrogen fixation (SNF) has a high energetic cost for legume plants; legumes thus reduce SNF when soil N is available. The present study aimed to increase our understanding regarding the impacts of the two principal forms of available N in soils (ammonium and nitrate) on SNF. We continuously measured the SNF of Medicago truncatula under controlled conditions. This permitted nodule sampling for comparative transcriptome profiling at points connected to the nodules’ reaction following ammonium or nitrate applications. The N component of both ions systemically induced a rhythmic pattern of SNF, while the activity in control plants remained constant. This rhythmic activity reduced the per-day SNF. The nitrate ion had additional local effects; the more pronounced were a strong downregulation of leghaemoglobin, nodule cysteine-rich (NCR) peptides and nodule-enhanced nicotianamine synthase (neNAS). The neNAS has proven to be of importance for nodule functioning. Although other physiological impacts of nitrate on nodules were observed (e.g. nitrosylation of leghaemoglobin), the main effect was a rapid ion-specific and organ-specific change in gene expression levels. Contrastingly, during the first hours after ammonium applications, the transcriptome remained virtually unaffected. Therefore, nitrate-induced genes could be key for increasing the nitrate tolerance of SNF.

With the forecasted fast increase in world population and global climate change, providing sufficient amounts of quality food becomes a major challenge for human society. Seed and fruit crop yield is determined by developmental processes including flower initiation, pollen fertility and fruit set. Fruit set is defined as the transition from flower to young fruit, a key step in the development of sexually reproducing higher plants. Plant hormones have important roles during flower pollination and fertilization, leading to fruit set. Moreover, it is well established that fruit set can be triggered by phytohormones like auxin and gibberellins (GAs), in the absence of fertilization, both hormones being commonly used to produce parthenocarpic fruits and to increase fruit yield. Additionally, a number of studies highlighted the role of ethylene in plant reproductive organ development. The present review integrates current knowledge on the roles of auxin and ethylene in different steps of the fruit set process with a specific emphasis on the interactions between the two hormones. A deeper understanding of the interplay between auxin and ethylene may provide new leads towards designing strategies for a better control of fruit initiation and ultimately yield.

Polyamines (PAs) are small aliphatic amines with important regulatory activities in plants. Biotic stress results in changes in PA levels due to de novo synthesis and PA oxidation. In Arabidopsis thaliana five FAD-dependent polyamine oxidase enzymes (AtPAO1-5) participate in PA back-conversion and degradation. PAO activity generates H2O2, an important molecule involved in cell signaling, elongation, programmed cell death, and defense responses. In this work we analyzed the role of AtPAO genes in the Arabidopsis thaliana-Pseudomonas syringae pathosystem. AtPAO1 and AtPAO2 genes were transcriptionally up-regulated in infected plants. Atpao1-1 and Atpao2-1 single mutant lines displayed altered responses to Pseudomonas, and an increased susceptibility was found in the double mutant Atpao1-1 x Atpao2-1. These polyamine oxidases mutant lines showed disturbed contents of ROS (H2O2 and O2−) and altered activities of RBOH, CAT and SOD enzymes both in infected and control plants. In addition, changes in the expression levels of AtRBOHD, AtRBOHF, AtPRX33, and AtPRX34 genes were also noticed. Our data indicate an important role for polyamine oxidases in plant defense and ROS homeostasis.

Plant defensins have been implicated in the plant defense system, but their role in poplar immunity is still unclear. In the present study, we present evidence that PtDefensin, a putative plant defensin, participates in the defense of poplar plants against Septotis populiperda infection. After the construction of recombinant plasmid PET-32a-PtDefensin, PtDefensin protein was expressed in Escherichia coli strain BL21 (DE3) and purified through Ni-IDA resin affinity chromatography. The Trx-PtDefensin fusion protein displayed no cytotoxic activity against RAW264.7 cells but had cytotoxic activity against E. coli K12D31 cells. Analyses of PtDefensin transcript abundance showed that the expression levels of PtDefensin responded to abiotic and biotic stresses. Overexpression of PtDefensin in ‘Nanlin 895’ poplars (Populus × euramericana cv ‘Nanlin895’) increased resistance to Septotis populiperda, coupled with upregulation of MYC2 (basic helix-loop-helix (bHLH) transcription factor) related to jasmonic acid (JA) signal transduction pathways and downregulation of Jasmonate-zim domain (JAZ), an inhibitor in the JA signal transduction pathway. We speculate that systemic acquired resistance (SAR) was activated in non-transgenic poplars after S. populiperda incubation, and that induced systemic resistance (ISR) was activated more obviously in transgenic poplars after S. populiperda incubation. Hence, overexpression of PtDefensin may improve the resistance of poplar plants to pathogens.

In plants, the organ size is one of the most important features and regulated by an elaborate developmental program involving both internal and external signals. The steroidal hormone brassinosteroid (BR) plays an important role in regulating the organ size. BRASSINAZOLE RESISTANT 1 (BZR1) is one of important transcription factors that regulate organ size in BR signal pathway in Arabidopsis. The function of BZR1 on organ size is well characterized in Arabidopsis, but poorly understood in maize (Zea mays). To understand the mechanism of intrinsic organ size regulated by BZR1 during organogenesis, we identified the maize BZR1 and examined its function in Arabidopsis. Overexpression of ZmBZR1 displayed phenotypes of enlarged cotyledons, rosette leaves, floral organ and seed size in Arabidopsis. The cells in rosette leaves as well as other organs in transgenic ZmBZR1 lines were dramatically larger and longer than those in Col-0. ChIP and RNA-seq analysis showed ZmBZR1 can directly bind to the promoter region of organ size related genes, Germination Repression and Cell Expansion receptor-like kinase (GRACE) and KIP-RELATED PROTEIN6 (KRP6) to regulate their expression, suggesting ZmBZR1 is required for the progressive increase in cells during Arabidopsis development. Collectively, our findings provide significant insights into the mechanisms underlying regulation of organ size mediated by maize BZR1.

The soil-born vascular disease Verticillium wilt, which is caused by fungal pathogen Verticillium dahliae, is a devastating disease of cotton worldwide. In the last decade, a large number of genes have been found to participate in cotton–V. dahliae interactions, but the detailed mechanisms of cotton resistance to V. dahliae remain unclear. Here, we functionally characterized MPK3, a MAPK gene from cotton. MPK3 was induced in the roots of both resistant and susceptible cotton cultivars by V. dahliae inoculation. Transgenic cotton and tobacco with constitutively higher GbMPK3 expression conferred higher V. dahliae susceptibility, while MPK3 knockdown in cotton has limited effect on cotton resistance to V. dahliae. Expression profiling revealed that SA-mediated defense pathway genes (WRKY70, PR1, and PR5) accumulated after V. dahliae inoculation in roots of both wild-type and transgenic cotton, and the expression levels of these genes were higher in GbMPK3-overexpressing plants than in wild-type plants, indicating that GbMPK3 upregulation may reduce plant resistance to V. dahliae through regulating salicylic acid signaling transduction.

SnRK2 (sucrose non-fermenting 1-related protein kinases 2) protein kinase family involves in several abiotic stress response in plants. Although the regulatory mechanism of SnRK2 have been well demonstrated in Arabidopsis thaliana, their functions in rice are still largely unknown. Here, we report a SnRK2 family gene, OsSAPK8, can be strongly induced by abiotic stresses, including low-temperature, drought and high salt stress. The ossapk8 mutants showed lower tolerance to low-temperature, high salinity and drought stresses at the vegetative stages. Moreover, the expressions of marker genes for those abiotic stresses, e.g. OsDREB1, OsDREB2, OsNCED and OsRAB21, were downregulated in the ossapk8 mutants. We further confirmed that the yield was reduced in ossapk8 mutant lines compared with the wild type. Our results provide evidence for OsSAPK8 acting as a positive regulator in cold, drought, and salt stress responses.

The AP2/ERF (APETALA2/ethylene-responsive factor) family of transcription factors (TF) is involved in regulating biotic and abiotic stress responses in plants. To explore the role of AP2/ERFs in cold tolerance in woody plants, BpERF13 was cloned and characterized in Betula platyphylla (white birch), a species primarily found in Asia in temperate and boreal climates. Based on phylogenetic analysis, BpERF13 is a member of the IXb subfamily of ERFs. Using qRT-PCR, we found that BpERF13 was differentially expressed in different tissues, and its expression could be induced by cold treatment (4 °C). BpERF13 protein, fused with GFP, was exclusively localized to nuclei. To further assess the role of BpERF13 in cold tolerance, BpERF13 overexpression (OE) transgenic lines were generated in B. platyphylla and used for cold stress treatment and biochemical/physiological studies. BpERF13 overexpression lines had significantly increased tolerance to subfreezing treatment and reduced reactive oxygen species. Using a TF-centered yeast one-hybrid (Y1H) experimental system, we showed that BpERF13 could bind to LTRECOREATCOR15 and MYBCORE cis-elements to activate a reporter gene. ChIP-seq and ChIP-PCR experiments further demonstrated that BpERF13 bound to these cis-elements when present in the 5’ proximal regions of superoxide dismutase (SOD), peroxidase (POD), and C-repeat-binding factor (CBF) genes. qRT-PCR was employed to examine the expression levels of these genes in response to cold stress; SOD, POD, and CBF genes were significantly upregulated in BpERF13 transgenic lines compared to wild-type plants in response to cold stress. These results indicate that the transcription factor BpERF13 regulates physiological processes underlying cold tolerance in woody plants.

This study investigated the dynamic of zinc (Zn) uptake and the root-to-shoot Zn-transport when supplied as ZnSO4 (aq) or Zn-EDTA (aq) in soybean seedlings using in vivo X-ray fluorescence (XRF) and X-ray absorption spectroscopy (XANES). The time-resolved X-ray fluorescence showed that plants absorbed ca. 10-fold more Zn from ZnSO4 (aq) than from Zn-EDTA (aq). However, the uptake velocity did not influence the amount of Zn in the stem. It let furthermore appear that the plants were able to reduce the absorption of Zn from Zn-EDTA (aq) earlier than ZnSO4 (aq). Thus, the entrance of Zn2+ into the roots is not necessarily accompanied by SO42-(aq). Regardless the source, the Zn distribution and its transport in the stem were spatially correlated to the bundles and cortex nearby the epidermal cells. Its chemical speciation showed that Zn is neither transported as ZnSO4(aq) nor as Zn-EDTA(aq), indicating that these compounds are retained in the roots or biotransformed on in the root-solution interface. Zn2+ was long-distance transported complexed by organic molecules such as histidine, malate, and citrate, and the proportion of ligands was affected by the concentration of Zn2+ in the stem rather than by the type of Zn source.

Moderate heat stress is usually accompanied with fluctuating light in summer. Although either heat stress or fluctuating light can cause photoinhibition of photosystems I and II (PSI and PSII), it is unclear whether moderate heat stress accelerate photoinhibition under fluctuating light. Here, we measured chlorophyll fluorescence, P700 redox state and the electrochromic shift signal under fluctuating light at 25 °C and 42 °C for tobacco leaves. We found that (1) the thylakoid proton conductance was significantly enhanced at 42 °C, leading to a decline in trans-thylakoid proton gradient (ΔpH); (2) this low ΔpH at 42 °C did not decrease donor-side limitation of PSI and thermal energy dissipation in PSII; (3) the activation of cyclic electron flow (CEF) around PSI was elevated at 42 °C; and (4) the moderate heat stress did not accelerate photoinhibition of PSI and PSII under fluctuating light. These results strongly indicate that under moderate heat stress the stimulation of CEF protects PSI under fluctuating light in tobacco leaves.

The development of high-yielding wheat genotypes containing micronutrient-dense grains are the main priorities of biofortification programs. At the International Maize and Wheat Improvement Center, breeders have successfully crossed high zinc progenitors including synthetic hexaploid wheat, T. dicoccum, T. spelta and landraces to generate high-zinc varieties. In this study, we report a genome-wide association using a wheat diversity panel to dissect the genetics controlling zinc, iron, copper, manganese and phosphorus concentrations in the grain and rachis during grain development and at physiological maturity. Significant marker-trait associations (MTAs) were identified for each nutrient using multi-locus mixed model methodologies. For mature grain, markers that showed significant pleiotropic effects were found on chromosomes 1A, 3B and 5B, of which those on chromosome 5B at ∼95.5 cM were consistent over two growing seasons. Co-located MTAs were identified for the nutrient concentrations in developing grain, rachis and mature grain on multiple chromosomes. The identified genomic regions included putative candidate genes that involved in metal uptake and transport and storage protein processing. These findings add to our understanding of the genetics of the five important nutrients in wheat grain and provide information on genetic markers for selecting high micronutrient densities containing genotypes.

Ethanol is known to accumulate in various plant organs under various environmental conditions. However, there are very scarce data about ethanol sensing by plants. We observed that ethanol accumulates up to 3.5 mM during tomato seed imbibition, particularly when seeds were stacked. Stacked seeds germinated less than spread out seeds suggesting ethanol inhibits germination. In support of this, exogenous ethanol at physiological concentrations, ranging from 1 to 10 mM, inhibited germination of wild type tomato seeds. However, the germination pattern over the whole ethanol concentration range tested was modified in an ethylene insensitive mutant, never-ripe (nr). The effects of exogenous ethanol were not linked to differences in ethylene production by imbibed seeds. But, we observed that exogenous ethanol at a concentration as low as 0.01 mM down regulated the expression of some ethylene receptors. Moreover, the triple response induced by ethylene in tomato seedlings was partially alleviated by 1 mM ethanol. Similar observations were made on Arabidopsis seeds. These results show there are interactions between ethylene sensing and ethanol in plants.

Physiological effects of ammonium (NH4+) and nitrate (NO3-) on tea have confirmed that tea plants prefer NH4+ as the dominant nitrogen (N) source. To investigate the possible explanations for this preference, studies of 15NH4+ and 15NO3- assimilation using hydroponically grown tea plants were conducted. During the time course of 15NH4+ and 15NO3- assimilation, the absorption of 15N from 15NH4+ was more rapid than that from 15NO3-, as there was a more efficient expression pattern of NH4+ transporters compared with that of NO3- transporters. 15NH4+-fed tea plants accumulated more 15N than 15NO3--fed plants, which was demonstrated by that genes related to primary N assimilation, like CsNR, CsNiR, CsGDH and CsGOGAT, were more affected by 15NH4+ than 15NO3-. Markedly higher NH4+ concentrations were observed in 15NH4+-fed tea roots in comparison with NO3- treatment, whereas tea plants maintained a balanced concentration of NH4+ in tea leaves under both these two N forms. This maintenance was achieved through the increased expression of genes involved in theanine biosynthesis and the inhibition of genes related to catechins derived from phenylpropanoid pathway. The current results suggest that efficient NH4+ transportation, assimilation, and reutilization enables tea plant as an ammonium preferring plant species.

Salt stress is one of the major environmental factors limiting crop productivity. Although physiological and molecular characterization of salt stress response in plants has been the focus for many years, research on transporters for sodium ion (Na+) uptake, translocation and accumulation in plants, particularly in food crops like rice is limited. In this study, we functionally identified an uncharacterized sodium ion transporter named OsNHAD which encodes a putative Na+ ⁄ H+ antiporter in rice. Homology search shows its close relation to the Arabidopsis Na+/H+ antiporter AtNHD1 with 72.74% identity of amino acids. OsNHAD transcripts mainly express in leaves and are induced by Na+ stress. Confocal laser scanning microscopy analysis of OsNHAD::GFP fusion in tobacco leaves shows that OsNHAD resides in the chloroplast envelop. Knock-down of OsNHAD by RNA interference led to increased rice sensitivity to Na+, manifested by stunted plant growth, enhanced cellular damage, reduced PSII activity and changed chloroplast morphology. Mutation of OsNHAD also resulted in accumulation of more Na+ in chloroplasts as well as shoots, suggesting that OsNHAD is involved in mediating efflux and detoxification of Na+ but does not affect K+ accumulation in plant cells and in tissues. Complementation test reveals that OsNHAD was able to functionally restore the Arabidopsis mutant atnhd1-1 growth phenotype. These results suggest that OsNHAD possibly mediates homeostasis of sodium ions in the subcellular compartments and tissues of the plants when challenged to salt stress.

Soybean mosaic virus (SMV) is a major pathogen causing yield loss. Developing soybean plants tolerant or resistant to SMV is important for mitigating the adverse effects of the viral infection. However, most studies have focused on the resistance to normal SMV strains. Thus, investigations of the resistance or tolerance to the novel recombinant SMV strain have been limited. To address the threat of the recombinant SMV, two soybean parent genotypes with contrasting reactions to the recombinant SMV and 211 F9:11 recombinant inbred lines were evaluated under artificial inoculation conditions. The JD12 plants are resistant to the recombinant SMV, whereas HT is highly tolerant, but still susceptible. Genetic analyses suggested that the resistance of JD12 is controlled by a single dominant gene and the tolerance is a quantitative trait. The QTL mapping results revealed one QTL (qTsmv-13) for resistance and two QTLs (qTsmv-2 and qTsmv-3) for tolerance. A comparison between known resistance genes and the QTLs identified in this study suggested that qTsmv-13 and qTsmv-2 may correspond to Rsv1 and Rsv4, respectively, whereas qTsmv-3 represents a newly identified QTL for SMV tolerance. We further delimited qTsmv-3 to an interval of approximately 86 kb with a map-based cloning strategy. Only two of five candidate genes, Glyma.03G00550 and Glyma.03G00570, varied between the parents. Additionally, Glyma.03G00550, which is a multidrug and toxic compound extrusion transporter gene, is the likely candidate gene for qTsmv-3. In summary, our research opens a new avenue for formulating strategies to breed soybean varieties tolerant to SMV.

Phenolic compounds are among the most diverse and widespread of specialized plant compounds and underly many important agronomic traits. Our comprehensive analysis of the maize genome unraveled new aspects of the genes involved in phenylpropanoid, monolignol, and flavonoid production in this important crop. Remarkably, just 19 genes accounted for 70% of the overall mRNA accumulation of these genes across 95 tissues, indicating that these are the main contributors to the flux of phenolic metabolites. Eighty genes with intermediate to low expression play minor and more specialized roles. Remaining genes are likely undergoing loss of function or are expressed in limited cell types. Phylogenetic and expression analyses revealed which members of gene families governing metabolic entry and branch points exhibit duplication, subfunctionalization, or loss of function. Co-expression analysis applied to genes in sequential biosynthetic steps revealed that certain isoforms are highly co-expressed and are candidates for metabolic complexes that ensure metabolite delivery to correct cellular compartments. Co-expression of biosynthesis genes with transcription factors discovered connections that provided candidate components for regulatory modules governing this pathway. Our study provides a comprehensive analysis of maize phenylpropanoid related genes, identifies major pathway contributors, and novel candidate enzymatic and regulatory modules of the metabolic network.

A RARSeq based Association mapping study was performed in a population of 104 Elaeis oleifera x E. guineensis hybrids of five origins with the aim of finding functional markers associated to six productive and 19 oil quality traits. For this purpose mRNA of each genotype was isolated and double stranded cDNA was synthesized. Following digestion with two restriction enzymes and adapter ligation, a size selected pool of barcoded amplicons was produced and sequenced using Illumina MiSeq. The obtained sequences were processed with a “snakemake” pipeline, filtered and missing values were imputed. For all traits except two significant effects of the origin was observed. Genetic diversity analyses revealed high variability within origins and an excess of heterozygosity in the population. Two GLM models with Q matrix or PCA matrix as covariates and two MLM models incorporating in addition a Kinship matrix were tested for genotype-phenotype associations using GAPIT software. Using unadjusted p values (< 0.01) 78 potential associations were detected involving 25 SNP and 20 traits. When applying FDR multiple testing with p < 0.05, 25 significant associations remained involving eight SNP and six quality traits. Four SNP were located in genes with a potential relevant biological meaning.

Recent reports have uncovered the multifunctional role of H2S in the physiological response of plants to biotic and abiotic stress. Here, we studied whether NaHS (H2S donor) pretreatment could provoke the tolerance of cucumber (Cucumis sativus L.) seedlings subsequently exposed to chilling stress and whether glutathione was involved in this process. Results showed that cucumber seedlings sprayed with NaHS exhibited remarkably increased chilling tolerance, as evidenced by the observed plant tolerant phenotype, as well as the lower levels of electrolyte leakage (EL), malondialdehyde (MDA) content, hydrogen peroxide (H2O2) content and RBOH mRNA abundance, compared with the control plants. In addition, NaHS treatment increased the endogenous content of the reduced glutathione (GSH) and the ratio of reduced / oxidized glutathione (GSH/GSSG), meanwhile, the higher net photosynthetic rate (Anet), the light-saturated CO2 assimilation rate (Asat), the photochemical efficiency (Fv/Fm) and the maximum photochemical efficiency of PSII in darkness (φPSII) as well as the mRNA levels and activities of the key photosynthetic enzymes (Rubisco, TK, SBPase and FBAase) were observed in NaHS-treated seedlings under chilling stress, whereas this effect of NaHS was weakened by buthionine sulfoximine (BSO, an inhibitor of glutathione ) or 6-Aminonicotinamide (6-AN, a specific pentose inhibitor and thus inhibits the NADPH production), which preliminarily proved the interaction between H2S and GSH. Moreover, transcription profiling analysis revealed that the GSH-associated genes (GST Tau, MAAI, APX, GR, GS and MDHAR) were significantly up-regulated in NaHS-treated cucumber seedlings, compared to the H2O-treated seedlings under chilling stress. Thus, novel results highlight the importance of glutathione as a downstream signal of H2S-induced plant tolerance to chilling stress.

Leaf protein synthesis is an essential process at the heart of plant nitrogen (N) homeostasis and turnover that preferentially takes place in the light, that is, when N and CO2 fixation occur. The carbon allocation to protein synthesis in illuminated leaves generally accounts for ca. 1% of net photosynthesis. It is likely that protein synthesis activity varies with photosynthetic conditions (CO2/O2 atmosphere composition) since changes in photorespiration and carbon provision should in principle impact on amino acid supply as well as metabolic regulation via leaf sugar content. However, possible changes in protein synthesis and translation activity when gaseous conditions vary are virtually unknown. Here, we address this question using metabolomics, isotopic techniques, phosphoproteomics and polysome quantitation, under different photosynthetic conditions that were varied with atmospheric CO2 and O2 mole fraction, using illuminated Arabidopsis rosettes under controlled gas exchange conditions. We show that carbon allocation to proteins is within 1–2.5% of net photosynthesis, increases with photosynthesis rate and is unrelated to total amino acid content. In addition, photosynthesis correlates to polysome abundance and phosphorylation of ribosomal proteins and translation initiation factors. Our results demonstrate that translation activity follows photosynthetic activity, showing the considerable impact of metabolism (carboxylation–oxygenation balance) on protein synthesis.

Sclerotinia sclerotiorum causes a devastating disease in oilseed rape (Brassica napus), resulting in major economic losses. Resistance response of B. napus against S. sclerotiorum exhibits a typical quantitative disease resistance (QDR) characteristic, but the molecular determinants of this QDR are largely unknown. In this study, we isolated a B. napus mitogen-activated protein kinase gene, BnaMPK6, and found that BnaMPK6 expression is highly responsive to infection by S. sclerotiorum and treatment with salicylic acid (SA) or jasmonic acid (JA). Moreover, overexpression (OE) of BnaMPK6 significantly enhances resistance to S. sclerotiorum, whereas RNAi in BnaMPK6 significantly reduces this resistance. These results showed that BnaMPK6 plays an important role in defense to S. sclerotiorum. Furthermore, expression of defense genes associated with SA-, JA- and ethylene (ET)-mediated signaling was investigated in BnaMPK6-RNAi, WT and BnaMPK6-OE plants after S. sclerotiorum infection, and consequently, it was indicated that the activation of ET signaling by BnaMPK6 may play a role in the defense. Further, four BnaMPK6-encoding homologous loci were mapped in the B. napus genome. Using the allele analysis and expression analysis on the four loci, we demonstrated that the locus BnaA03.MPK6 makes an important contribution to QDR against S. sclerotiorum. Our data indicated that BnaMPK6 is a previously unknown determinant of QDR against S. sclerotiorum in B. napus.

Light modulates almost every aspect of plant physiology, including plant-pathogen interactions. Among these, the hypersensitive response (HR) of plants to pathogens is characterized by a rapid and localized programmed cell death (PCD), which is critical to restrict the spread of pathogens from the infection site. The aim of this work was to study the role of light in the interaction between Pseudomonas syringae pv. tomato DC3000 (Pto DC3000) and non-host tobacco plants. To this end, we examined the HR under different light treatments (white and red light) by using a range of well-established markers of PCD. The alterations found at the cellular level included: i) loss of membrane integrity and nuclei, ii) RuBisCo and DNA degradation, and iii) changes in nuclease profiles and accumulation of cysteine proteinases. Our results suggest that red light plays a role during the HR of tobacco plants to Pto DC3000 infection, delaying the PCD process.

In higher plants, phytohormone indole-3-acetic acid is characteristically transported from the apex towards the base of the plant, termed as polar auxin transport (PAT). Among the inhibitors blocking PAT, N-1-naphthylphthalamic acid (NPA) that targets ABCB transporters is most commonly used. NPA-treated light-grown Arabidopsis seedlings show severe inhibition of hypocotyl and root elongation. In light-grown tomato seedlings, NPA inhibited root growth, but contrary to Arabidopsis stimulated hypocotyl elongation. The NPA-stimulation of hypocotyl elongation was milder in blue, red, and far-red light-grown seedlings. The NPA-treatment stimulated emission of ethylene from the seedlings. The scrubbing of ethylene by mercuric perchlorate reduced NPA-stimulated hypocotyl elongation. NPA action on hypocotyl elongation was antagonized by 1-methylcyclopropene, an inhibitor of ethylene action. NPA-treated seedlings had reduced levels of indole-3-butyric acid and higher levels of zeatin in the shoots. NPA did not alter indole-3-acetic levels in shoots. The analysis of metabolic networks indicated that NPA-treatment induced moderate shifts in the networks compared to exogenous ethylene that induced a drastic shift in metabolic networks. Our results indicate that in addition to ethylene, NPA-stimulated hypocotyl elongation in tomato may also involve zeatin and indole-3- butyric acid. Our results indicate that NPA-mediated physiological responses may vary in a species-specific fashion.

Daucus carota is a biennale crop that develops an edible storage root. Orange carrots, the most consumed cultivar worldwide, accumulate high levels of β-carotene and α-carotene in the storage root during secondary growth. Genes involved in β-carotene synthesis have been identified in carrots and unlike most species, D. carota has two ζ-carotene desaturase genes, named ZDS1 and ZDS2, that share 91.3% identity in their coding regions. ZDS1 expression falls during leaf, but not root development, while ZDS2 is induced in leaves and storage roots of a mature plant. In this work, by means of post-transcriptional gene silencing, we determined that ZDS1 is essential for initial carrot development. The suppression of the expression of this gene by RNAi triggered a reduction in the transcript levels of ZDS2 and PSY2 genes, with a concomitant decrease in the carotenoid content in both, leaves and storage roots. On the contrary, transgenic lines with reduced ZDS2 transcript abundance maintain the same levels of expression of endogenous ZDS1 and PSY2 and carotenoid profile as wild-type plants. The simultaneous silencing of ZDS1 and ZDS2 resulted in lines with a negligible leaf and root development, as well as significantly lower endogenous PSY2 expression. Further functional analyses, such as a plastidial subcellular localization of ZDS1:GFP and the increment in carotenoid content in transgenic tobacco plants overexpressing the carrot ZDS1, confirmed that ZDS1 codifies for a functional enzyme. Overall, these results lead us to propose that the main ζ-carotene desaturase activity in carrot is encoded by the ZDS1 gene and ZDS2 gene has a complementary and non essential role.