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Active DNA demethylation effectively modulates gene expression during plant development and in response to stress. However, little is known about the upstream regulatory factors that regulate DNA demethylation. We determined that the demethylation regulator 1 (demr1) mutant exhibits a distinct DNA methylation profile at selected loci queried by methylation-sensitive polymerase chain reaction and globally based on whole-genome bisulfite sequencing. Notably, the transcript levels of the DNA demethylase gene REPRESSOR OF SILENCING 1 (ROS1) were lower in the demr1 mutant. We established that DEMR1 directly binds to the ROS1 promoter in vivo and in vitro, and the methylation level in the DNA methylation monitoring sequence of ROS1 promoter decreased by 60% in the demr1 mutant. About 40% of the hyper-differentially methylated regions (DMRs) in the demr1 mutant were shared with the ros1-4 mutant. Genetic analysis indicated that DEMR1 acts upstream of ROS1 to positively regulate abscisic acid (ABA) signaling during seed germination and seedling establishment stages. Surprisingly, the loss of DEMR1 function also caused a rise in methylation levels of the mitochondrial genome, impaired mitochondrial structure and an early flowering phenotype. Together, our results show that DEMR1 is a novel regulator of DNA demethylation of both the nuclear and mitochondrial genomes in response to ABA and plant development in Arabidopsis.

Increasing temperature is one of the major threats to maize growth and yield globally. Under heat stress conditions, intracellular protein homeostasis is seriously disturbed, leading to accumulation of abnormally folded proteins, especially in the endoplasmic reticulum (ER). Molecular chaperones are vital players in the renaturation process and in preventing protein aggregation. However, heat stress tolerance-associated chaperones are not well documented in maize. Here, we characterized the biological roles of HEAT UP-REGULATED GENE 1 (ZmHUG1) in maize. ZmHUG1 encodes a heat-inducible holdase-type molecular chaperone localized in the ER. Knockout mutant of ZmHUG1 exhibited remarkably enhanced sensitivity to heat stress. Accordingly, the zmhug1 mutant showed severe ER stress under high temperature. MAIZE PRENYLATED RAB ACCEPTOR 1.C1 (ZmPRA1.C1) was identified as a client of ZmHUG1, and heat-induced aggregation of ZmPRA1.C1 was accelerated in the zmhug1 mutant. Furthermore, the expression of ZmHUG1 was rapidly transactivated by ER stress sensor BASIC LEUCINE ZIPPER DOMAIN 60 (bZIP60) when heat stress occurred. This study reveals a ZmHUG1-based thermo-protective mechanism in maize.

Fusarium graminearum is one of the most important causal agents of Fusarium head blight disease and is controlled mainly by chemicals such as demethylation inhibitor (DMI) fungicides. FgCYP51B is one of the DMI targets in F. graminearum, and Tyrosine123 (Y123) is an important amino acid in F. graminearum CYP51B, located in one of predicted substrate binding pockets based on the binding mode between DMIs and CYP51B. Previous studies suggest that resistance to DMI fungicides is attributed primarily to point mutations in the CYP51 gene and that the Y123H mutation in F. verticillioides CYP51 confers prochloraz resistance in the laboratory. To investigate the function of FgCYP51B Y123 residue in the growth and development, pathogenicity, and DMI resistance, we generated and analyzed the FgCYP51B Y123H mutant. Results revealed that the Y123H mutation led to reduced conidial sporulation and affected ascospore development; moreover, the mutation conferred reduced sensitivity to prochloraz. Quantitative PCR and molecular docking were performed to investigate the resistance mechanism. Results indicated that Y123H mutation changed the target gene expression and decreased the binding affinity of FgCYP51 to prochloraz. These results will attract more attention to the potential DMI-resistant mutation of F. graminearum and increase our understanding of the DMI resistance mechanism.

C-terminally encoded peptides (CEPs) are small peptides, typically post-translationally modified, and highly conserved in many species. CEPs are known to inhibit plant growth and development, but the mechanisms are not well understood. In this study, 14 CEPs were identified in Setaria italica and divided into two groups. The transcripts of most SiCEPs were more abundant in roots than in other detected tissues. SiCEP3, SiCEP4, and SiCEP5 were also highly expressed in panicles. Moreover, expression of all SiCEPs was induced by abiotic stresses and phytohormones. SiCEP3 overexpression and application of synthetic SiCEP3 both inhibited seedling growth. In the presence of abscisic acid (ABA), growth inhibition and ABA content in seedlings increased with the concentration of SiCEP3. Transcripts encoding eight ABA transporters and six ABA receptors were induced or repressed by synthetic SiCEP3, ABA, and their combination. Further analysis using loss-of-function mutants of Arabidopsis genes functioning as ABA transporters, receptors, and in the biosynthesis and degradation of ABA revealed that SiCEP3 promoted ABA import at least via NRT1.2 (NITRATE TRANSPORTER 1.2) and ABCG40 (ATP-BINDING CASSETTE G40). In addition, SiCEP3, ABA, or their combination inhibited the kinase activities of CEP receptors AtCEPR1/2. Taken together, our results indicated that the CEP-CEPR module mediates ABA signaling by regulating ABA transporters and ABA receptors in planta.

Grain yield and salt tolerance are critical for crop production. However, the genetic and biochemical basis underlying the trade-off of these characters remain poorly described in crops. We show here that SiPLATZ12 transcription factor positively regulates multiple elite yield traits at the expense of salt tolerance in foxtail millet. SiPLATZ12 overexpression increases seed size, panicle length, and stem diameter, while reduces plant height and salt tolerance of foxtail millet. A 9-bp insertion in the SiPLATZ12 promoter has significant effects on the different expression of SiPLATZ12, multiple yield traits, and salt tolerance between foxtail millet and its wild ancestor, green foxtail. Moreover, SiPLATZ12 upregulates the expression of genes involved in seed development, but repressing the transcription of most NHX, SOS, and CBL genes to regulate Na+, K+ and pH homeostasis. Therefore, our results uncover a domesticated site that could be used to improve grain yield and salt tolerance in foxtail millet.

Abscisic acid (ABA) transport plays an important role in systemic plant responses to environmental factors. However, it remains largely unclear about the precise regulation of ABA transporters in plants. In this study, we show that the C-terminally encoded peptide receptor 2 (CEPR2) directly interacts with the ABA transporter NRT1.2/NPF4.6. Genetic and phenotypic analyses revealed that NRT1.2/NPF4.6 positively regulates ABA response and that NRT1.2/NPF4.6 is epistatically and negatively regulated by CEPR2. Further biochemical assays demonstrated that CEPR2 phosphorylates NRT1.2/NPF4.6 at serine 292 to promote its degradation under normal conditions. However, ABA treatment and non-phosphorylation at serine 292 prevented the degradation of NRT1.2/NPF4.6, indicating that ABA inhibits the phosphorylation of this residue. Transport assays in yeast and Xenopus oocytes revealed that non-phosphorylated NRT1.2/NPF4.6 had high levels of ABA import activity, whereas phosphorylated NRT1.2/NPF4.6 did not import ABA. Analyses of complemented nrt1.2 mutants that mimicked non-phosphorylated and phosphorylated NRT1.2/NPF4.6 confirmed that non-phosphorylated NRT1.2S292A had high stability and ABA import activity in planta. Additional experiments showed that NRT1.2/NPF4.6 was degraded via the 26S proteasome and vacuolar degradation pathways. Furthermore, we found that three E2 ubiquitin-conjugating enzymes, UBC32, UBC33, and UBC34, interact with NRT1.2/NPF4.6 in the endoplasmic reticulum and mediate its ubiquitination. NRT1.2/NPF4.6 is epistatically and negatively regulated by UBC32, UBC33, and UBC34 in planta. Taken together, these results suggest that the stability and ABA import activity of NRT1.2/NPF4.6 are precisely regulated by its phosphorylation and degradation in response to environmental stress.

Normally, recycled fine aggregate (RFA) is an unwanted by-product of construction demolished waste crushing, with high contaminant content. The presence of RFA is highly restricted in ordinary Portland cement (OPC) concrete production because of the poor durability and mechanical performance. This paper presents a study of the effect of RFA on the mechanical and durability of alkali activated slag concrete (AASC). Results indicated that RFA can be used as fine aggregate in AASC with superior early age compressive strength compared to river sand in AASC. It is caused by the presence of non-hydrated cement paste attached to the RFA with alkali activate property, which is equivalent to increase the quantity of alkali activator, and accelerate the hydration of ground granulated blast furnace slag (GGBFS) especially in the early age. In later age, the mechanical properties of AASC incorporating RFA are slightly lower than conventional AASC due to the larger porosity in the concrete.

Alternative splicing (AS) is emerging as a critical co-transcriptional regulation for plants in response to environmental stresses. Although multiple splicing factors have been linked to the salt-sensitive signaling network, the molecular mechanism remains unclear. We discovered that a conserved serine/arginine-rich (SR)-like protein, SR45a, as a component of the spliceosome, was involved in post-transcriptional regulation of salinity tolerance in Arabidopsis thaliana. Furthermore, SR45a was required for the AS and messenger RNA (mRNA) maturation of several salt-tolerance genes. Two alternatively spliced variants of SR45a were induced by salt stress, full-length SR45a-1a and the truncated isoform SR45a-1b, respectively. Lines with overexpression of SR45a-1a and SR45a-1b exhibited hypersensitive to salt stress. Our data indicated that SR45a directly interacted with the cap-binding complex (CBC) subunit cap-binding protein 20 (CBP20) which mediated salt-stress responses. Instead of binding to other spliceosome components, SR45a-1b promoted the association of SR45a-1a with CBP20, therefore mediating salt-stress signal transduction pathways. Additionally, the mutations in SR45a and CBP20 led to different salt-stress phenotypes. Together, these results provide the evidence that SR45a-CBP20 acts as a regulatory complex to regulate the plant response to salt stress, through a regulatory mechanism to fine-tune the splicing factors, especially in stressful conditions.

Protein acetylation is one of the most common post-translational modifications. Many acetylated proteins in Magnaporthe oryzae play key roles in vegetative growth and pathogenicity. MoDabb1 from M. oryzae was also identified to be an acetylated protein, containing a Dabb domain with unknown function. To elucidate the function of this protein and the effect of acetylation on this protein, a native and selenomethionine-substituted MoDabb1 were expressed in Escherichia coli and purified to homogeneity. Crystals were obtained using sitting-drop vapour-diffusion method. Crystals of native and selenomethionine-substituted protein were diffracted to a resolution of 1.74 and 1.98 A and both belonged to the sp. gr. P42212. Matthews coefficient analysis indicated two molecules in an asymmetric unit with a Vm value of 2.41 and a corresponding solvent content of 49.03%.

PLATZ transcription factors play important roles in plant growth, development, and biotic and abiotic stress responses. However, how PLATZ regulates plant drought tolerance and ABA sensitivity remains largely unknown. Here, we show that PLATZ4 increases drought tolerance and ABA sensitivity in Arabidopsis thaliana by suppressing the expression of PIP2;8, DEG3, At1g33840 and At3g01345, while upregulating the expression of PR1, ABI3, ABI4 and ABI5. Moreover, PLATZ4 directly binds A/T-rich sequences within the PIP2;8 promoter but not ABI3, ABI4 and ABI5 promoters. Consistent with this, PIP2;8 acts epistatically to PLATZ4. Furthermore, the aquaporin activity of PIP2;8 was confirmed in Xenopus laevis oocytes and yeast cells in response to osmotic stress. Analysis of water loss of seedlings overexpressing PIP2;8 or lacking PIP2;8 function indicated that PIP2;8-mediated water flow is particularly active in response to drought stress in planta. Accordingly, stomatal closure was enhanced in pip2;8 mutants but reduced in PIP2;8-overexpressing plants. In platz4 mutant and PLATZ4-overexpressing plants, water loss and stomatal closure changed oppositely to those in pip2;8 mutants and PIP2;8-overexpressing plants, respectively. In addition, the interaction between PLATZ4 and AITR6 was confirmed by several assays, and the binding of PIP2;8 promoter by PLATZ4 was strengthened by an interaction with AITR6. Collectively, our findings reveal that PLATZ4 interacts with AITR6 to increase ABA sensitivity and drought tolerance in Arabidopsis by upregulating expression of PR1, ABI3, ABI4 and ABI5 while inhibiting the expression of PIP2;8 and associated genes.One sentence summaryPLATZ4 interacted with AITR6 to increase drought tolerance and ABA sensitivity in Arabidopsis thaliana by directly bind A/T-rich sequences within the plasma membrane aquaporin (PIP)2;8 promoter.

Backgrounds Lysine 2-hydroxyisobutyrylation (Khib) is a newly discovered posttranslational modification (PTM) and has been identified in several prokaryotic and eukaryotic organisms. Fusarium graminearum, a major pathogen of Fusarium head blight (FHB) in cereal crops, can cause considerable yield loss and produce various mycotoxins that threaten human health. The application of chemical fungicides such as tebuconazole (TEC) remains the major method to control this pathogen. However, the distribution of Khib in F. graminearum and whether Khib is remodified in response to fungicide stress remain unknown. Results Here, we carried out a proteome-wide analysis of Khib in F. graminearum, identifying the reshaping of the lysine 2-hydroxyisobutyrylome by tebuconazole, using the most recently developed high-resolution LC–MS/MS technique in combination with high-specific affinity enrichment. Specifically, 3501 Khib sites on 1049 proteins were identified, and 1083 Khib sites on 556 modified proteins normalized to the total protein content were changed significantly after TEC treatment. Bioinformatics analysis showed that Khib proteins are involved in a wide range of biological processes and may be involved in virulence and deoxynivalenol (DON) production, as well as sterol biosynthesis, in F. graminearum. Conclusions Here, we provided a wealth of resources for further study of the roles of Khib in the fungicide resistance of F. graminearum. The results enhanced our understanding of this PTM in filamentous ascomycete fungi and provided insight into the remodification of Khib sites during azole fungicide challenge in F. graminearum.

CYSTM3, a small mitochondrial protein, acts as a negative regulator in salt stress response by preventing Na+ efflux and disturbing reactive oxygen species (ROS) homeostasis in Arabidopsis. Cysteine-rich transmembrane module (CYSTM) is a not well characterized small peptide family in plants. In this study, we identified a novel mitochondrion-localized CYSTM member CYSTM3 from Arabidopsis, which was ubiquitously expressed in different tissues and dramatically induced by salt stress. Transgenic plants overexpressing CYSTM3 (OE) displayed hypersensitivity to salt stress compared with wild type (WT) plants, whereas a knockout mutant cystm3 was more tolerant to high salinity than WT. Moreover, OE lines accumulated higher contents of Na+ and ROS than WT and cystm3 upon exposure to high salinity. Further analysis revealed that CYSTM3 could deter root Na+ efflux and inhibit the activities of a range of ROS scavenging enzymes in Arabidopsis. In addition, the transcripts of nuclear salt stress-responsive genes were over-activated in cystm3 than those in WT and OE lines. Taken together, Arabidopsis CYSTM3 acts as a negative regulator in salt stress tolerance.

MoSub1 is an ortholog of yeast single stranded DNA binding protein Sub1 or human PC4 from rice blast fungus. All of them share a similar DNA binding region and may have similar biological roles. The well-studied Sub1/PC4 has been reported to play multiple roles in DNA metabolic processes, such as transcription and DNA repair and their DNA binding capacity is significantly affected by phosphorylation. Here, we determined the crystal structure of MoSub1 complexed with ssDNA in a phosphate solution. The crystal structure of the MoSub1-ssDNA complex was solved to a resolution of 2.04 Å. A phosphate ion at the interface of the protein-DNA interaction of the complex bridged the lys84 of the protein and two nucleotides. The DNA was bound in novel mode (L mode) in the MoSub1 complex in the presence of phosphate ions, while DNA bound in the straight mode in the absence of the phosphate ion and in U mode in the same binding motif of the PC4-ssDNA complex. The crystal structure of the complex and a small-angle X-ray scattering analysis revealed that the phosphate ion at the protein-DNA interface affected the DNA binding mode of MoSub1 to oligo-DNA and provided a new structural clue for studying its functions.

C-terminally encoded peptides (CEPs) are small peptides, typically post-translationally modified, and highly conserved in many species. CEPs are known to play roles in inhibition of plant growth and regulation of development, but the mechanisms are not well understood. In this study, we searched for CEP peptides in foxtail millet (Setaria italica). The 14 peptides we identified are divided into two subfamilies. The transcripts of most SiCEPs were more abundant in roots than in other tissues. SiCEP3, SiCEP4, and SiCEP5 were also expressed at high levels in panicles. Moreover, expression of all SiCEPs was induced by biotic stress and phytohormones. SiCEP3 overexpression and application of biosynthetic SiCEP3 both inhibited the growth of seedlings. In the presence of ABA, growth inhibition and ABA content of seedlings increased with the concentration of SiCEP3. Transcripts encoding two ABA transporters and one ABA receptor were induced by SiCEP3, ABA, and the two in combination. Further analysis revealed that SiCEP3 promoted ABA transport via NRT1.2 and ABCG40. In addition, SiCEP3, ABA, or the combination inhibited the kinase activities of CEP receptors CEPR1/2. Taken together, our results indicated that the CEP–CEPR module mediates ABA signaling by regulating ABCG40, NRT1.2, and PYL4 in planta.HighlightSiCEP3, a C-terminally encoded peptide, can promote ABA import and signaling pathway by inhibiting the kinase activity of its receptors under abiotic stress in Setaria italica.

Ethylenediaminetetraacetic acid (EDTA) is a well-established chelating agent used in industry, agriculture, food, and medicine. However, the analysis of an EDTA-sensitive Arabidopsis thaliana mutant revealed that EDTA can significantly promote nitric oxide (NO) accumulation, indicating that EDTA has unexpected biological functions beyond its chelating activity. This finding challenges our current understanding about the effects of EDTA on biological systems.One Sentence SummaryAn analysis of ethylenediaminetetraacetic acid (EDTA)-sensitive mutants suggests that EDTA can promote the accumulation of nitric oxide.

Abscisic acid (ABA) transport plays important role in systematic plant responses to environmental factors. Here, we showed that C-terminally encoded peptide receptor 2 (CEPR2) directly interacted with the ABA transporter NRT1.2. Using transgenic seedlings, we demonstrated that NRT1.2 positively regulated the ABA response, and that CEPR2 acted on NRT1.2 epistatically and negatively. Under normal conditions, CEPR2 phosphorylated NRT1.2 at least at serine 292 to promote the degradation of NRT1.2. However, ABA and serine 292 nonphospharylation strongly inhibited the degradation of NRT1.2, indicating that ABA-inhibited the phosphorylation of NRT1.2. Transport assays in yeast and Xenopus oocytes showed that nonphosphorylated NRT1.2 had high levels of ABA-import activity, but phosphorylated NRT1.2 did not import ABA. Analyses of complement nrt1.2 mutants by mimicking nonphospharylated and phospharylated NRT1.2 confirmed that nonphospharylated NRT1.2S292A had high stability and ABA import activity in planta. Further experiments indicated that NRT1.2 was degraded via the 26S proteasome and vacuolar degradation pathways. UBC32, UBC33, and UBC34 interacted with, and mediated the ubiquitination of NRT1.2. UBC32, UBC33, and UBC34 acted on NRT1.2 epistatically and negatively in planta. Thus, our results suggested the existence of novel plant mechanisms regulating NRT1.2 stability and ABA import activity in response to environmental conditions.

Although the salt overly sensitive (SOS) pathway plays essential roles in conferring salt tolerance in Arabidopsis thaliana, the regulatory mechanism underlying SOS gene expression remains largely unclear. In this study, AtPLATZ2 was found to function as a direct transcriptional suppressor of CBL4/SOS3 and CBL10/SCaBP8 in the Arabidopsis salt stress response. Compared with wild-type plants, transgenic plants constitutively overexpressing AtPLATZ2 exhibited increased sensitivity to salt stress. Loss of function of PLATZ2 had no observed salt stress phenotype in Arabidopsis, while the double mutant of PLATZ2 and PLATZ7 led to weaker salt stress tolerance than wild-type plants. Overexpression of AtPLATZ2 in transgenic plants decreased the expression of CBL4/SOS3 and CBL10/SCaBP8 under both normal and saline conditions. AtPLATZ2 directly bound to A/T-rich sequences in the CBL4/SOS3 and CBL10/SCaBP8 promoters in vitro and in vivo, and inhibited CBL4/SOS3 promoter activity in the plant leaves. The salt sensitivity of #11 plants constitutively overexpressing AtPLATZ2 was restored by the overexpression of CBL4/SOS3 and CBL10/SCaBP8. Salt stress-induced Na+ accumulation in both the shoots and roots was more exaggerated in AtPLATZ2-overexpressing plants than in the wild type. The salt stress-induced Na+ accumulation in #11 seedlings was also rescued by the overexpression of CBL4/SOS3 and CBL10/SCaBP8. Furthermore, the transcription of AtPLATZ2 was induced in response to salt stress. Collectively, these results suggest that AtPLATZ2 suppresses plant salt tolerance by directly inhibiting CBL4/SOS3 and CBL10/SCaBP8, and functions redundantly with PLATZ7.

Background Fusarium graminearum is the main pathogen of Fusarium head blight (FHB), a worldwide plant disease and one of the most significant wheat diseases in China. Demethylation inhibitor (DMI) fungicides, such as tebuconazole (TEC), are widely used to control FHB, but long-term use leads to low efficacy against FHB. Earlier studies showed that DMI resistance is associated with the fungal sterol 14α-demethylase (cytochrome P450 CYP51) gene, and that point mutations in the CYP51 gene are the primary mechanism of resistance to DMI fungicides. The aims of this study were to clarify the molecular mechanisms of resistance to TEC and identify the binding sites on the FgCYP51B protein. Results Site-directed mutagenesis was used to change the FgCYP51B gene of wild-type strain PH-1 from tyrosine to histidine at residue 137 (Y137H) to generate a mutant transformant, which was confirmed to be resistant to TEC compared with the parental strains. A three-dimensional FgCYP51B model was constructed, and molecular docking simulation studies were conducted to identify the optimum binding mode with TEC. The wild-type FgCYP51B protein displayed stronger affinity to TEC than that of the mutated FgCYP51B in the molecular docking analysis. Conclusion These results indicate that a Tyr137 amino acid mutation in the cytochrome P450 FgCYP51B could lead to resistance to TEC and that Y137 forms part of the tebuconazole-binding pocket. © 2017 Society of Chemical Industry.

Increasing evidence points to the tight relationship between alternative splicing (AS) and the salt stress response in plants. However, the mechanisms linking these two phenomena remain unclear. In this study, we have found that Salt-Responsive Alternatively Spliced gene 1 (SRAS1), encoding a RING-Type E3 ligase, generates two splicing variants: SRAS1.1 and SRAS1.2, which exhibit opposing responses to salt stress. The salt stress-responsive AS event resulted in greater accumulation of SRAS1.1 and a lower level of SRAS1.2. Comprehensive phenotype analysis showed that overexpression of SRAS1.1 made the plants more tolerant to salt stress, whereas overexpression of SRAS1.2 made them more sensitive. In addition, we successfully identified the COP9 signalosome 5A (CSN5A) as the target of SRAS1. CSN5A is an essential player in the regulation of plant development and stress. The full-length SRAS1.1 promoted degradation of CSN5A by the 26S proteasome. By contrast, SRAS1.2 protected CSN5A by competing with SRAS1.1 on the same binding site. Thus, the salt stress-triggered AS controls the ratio of SRAS1.1/SRAS1.2 and switches on and off the degradation of CSN5A to balance the plant development and salt tolerance. Together, these results provide insights that salt-responsive AS acts as post-transcriptional regulation in mediating the function of E3 ligase., Author summary High salinity severely affects plant growth and development, impairing crop production worldwide. E3 ligase is a stress-responsive regulator through ubiquitin-proteasome system for selective protein degradation. The E3s are regulated by transcriptional regulation and post-translational modifications. Here, we have discovered that stress-responsive AS acts as a post-transcriptional regulation modulating the function of E3 ligases. Intriguingly, the truncated proteins generated by salt-responsive AS play opposite roles compared with the full-length E3 ligase. The truncated isoform losing key domain could not degrade the target protein, instead, it interacts and competes with the E3 ligase through binding the same domain of the targets. This finding contributes significantly to a deeper mechanistic understanding of how AS regulates the function of E3 ligase in response to salt stress.

BACKGROUND Fusarium graminearum is the main pathogen of Fusarium Head Blight (FHB), a worldwide plant disease and one of the major wheat diseases in China. The control of the FHB is mainly dependent on the application of DMIs fungicides. Fungal sterol 14α-demethylase enzymes (CYP51) are the main target for DMIs fungicides. In order to investigate the binding mechanism between azoles and CYP51B in F.graminearum, the molecular modeling study and biological evaluation were performed. RESULTS Firstly, the homology model based on the crystal structure of Aspergillus fumigatus was built. Secondly, molecular docking and molecular dynamics (MD) simulation were used to identify the optimum binding mode between Propiconazole (PRP), Diniconazole (DIN), Triadimenol (TRL), Tebuconazole (TEC), and Triadimefon (TRN) with FgCYP51B. Furthermore, binding free energy of the five protein-inhibitor complexes was calculated by Molecular Mechanics Generalized Born Surface Area and Poisson–Boltzmann Surface Area (MM-GB/PBSA) methods. The key residues to the selective binding of azoles to FgCYP51B were recognized by per-residue free energy decomposition analysis. The five ligands have similar binding mode in the active pocket. Inhibitors PRP and TEC have more beneficial binding free energy to the enzyme than TRN, TRL and DIN. Furthermore, the Amino acid residues Phe511, Val136, Ile374, Ala308, Ser312 and Try137 of FgCYP51B are the key residues interacting with azoles fungicides. The EC50 value of PRP, TEC, DIN, TRL and TRN is 0.024, 0.047, 0.148, 0.154 and 0.474 mg/L according to the experimental evaluation, respectively. These five molecules exhibited potential inhibitory activity against CYP51B protein from the F. graminearum. CONCLUSION The results suggest that azoles fungicides for FgCYP51B should possess more hydrophobic groups interacting with residues Phe511, Val136, Ile374, Ala308, Ser312 and Tyr137. PRP and TEC should be preferred to control Fusarium Head Blight disease than DIN, TRL and TRN.

Numerous studies have demonstrated the function of salinity or jasmonic acid (JA) in plant growth and senescence. This study evaluated how the combination of salinity and methyl jasmonate (MeJA) (SaM) worked as a novel stress and then regulated plant growth in Arabidopsis. Firstly, we found that compared with MeJA or NaCl treatment alone, SaM would significantly intensified plant growth inhibition and senescence in wild-type (WT) seedlings, and these phenotypes could be partially compromised after SaM stress in JA-insensitive mutants. Meanwhile, genes involved in JA signaling and Senescence Associated Gene 13 (SAG13) were dramatically increased by SaM stress than that by MeJA or NaCl alone in WT. Moreover, a group of secondary metabolite - indolic glucosinolates (IGs) showed obvious over-accumulation after SaM treatment than that after each single one in WT, and the seedlings treated with IGs' metabolites performed similar inhibited growth and chlorotic leaves phenotypes compared with those caused by SaM stress. All these indicated the toxicity of IGs and their metabolites would prevent the growth progress of plants. Therefore, we concluded that SaM worked as a novel stress and intensified plant growth inhibition and senescence, which was dependent on JA-dependent and -independent signaling pathways.

Mitogen-activated protein kinase (MAPK) cascades are conserved and vital signaling components in the responses to various ambient stresses. Here, we report the identification of MAPKKK18, a drought resistance associated MAPK kinase kinase in Arabidopsis. The mapkkk18 knockout mutants displayed hypersensitivity to drought stress, whereas overaccumulation of MAPKKK18 in transgenic Arabidopsis plants significantly enhanced the resistance to drought. Expression pattern analysis revealed that the inducible expression of MAPKKK18 by osmotic stress was ABA and the canonical ABA signaling pathway dependent. Furthermore, MAPKKK18 mainly exerted its regulatory roles via downstream MAPKK3. These findings uncovered important roles for MAPKKK18 in drought resistance and expanded our understanding of the MAPK pathways in modulating abiotic stress responses.

CEPR2 interacts with some PYLs to promote their phosphorylation and degradation, whereas ABA inhibits this process. Thus, CEPR2 balances the growth regulation and stress response in Arabidopsis., Pyrabactin resistance 1 (PYR1)/PYR1-like (PYL) abscisic acid (ABA) receptors have been proved to be recruited in the plasma membrane (PM). In order to explain the roles of PYR/PYLs in the PM, PYL4 was used as bait to screen the PM-localized leucine-rich repeat receptor-like kinase family, and five members were found directly interacting with PYL4. Loss of function by T-DNA insertion in C-terminally encoded peptide receptor 2 (CEPR2) together with phloem intercalated with xylem (PXY) and PXY-Like 2 (PXL2) led to ABA hypersensitivity, while CEPR2 overexpression led to ABA insensitivity compared with the wild type, indicating the redundant and negative roles of CEPR2, PXY, and PXL2 in ABA signaling. The PYL4 proteins were strongly accumulated in cepr2/pxy/pxl2 compared with the wild type. Furthermore, higher phosphorylation levels accompanied by lower protein levels of PYL4 in CEPR2 overexpression lines were observed, indicating the requirement of phosphorylation of PYLs for degradation. Subsequently, MS and in vitro kinase assays demonstrated that CEPR2 phosphorylated PYL4 at Ser54, while this phosphorylation was diminished or even eliminated in the presence of ABA. Taken together, CEPR2 promotes the phosphorylation and degradation of PYLs in unstressed conditions, whereas ABA represses this process to initiate ABA response during times of stress.

Heat stress significantly disturbs the protein folding and processing capability in plants. Molecular chaperones are vital players in unfolded/misfolded protein assembly and abiotic stress tolerance. Here, we reported SES1, which encodes an endoplasmic reticulum (ER) localized molecular chaperone, is required for Arabidopsis heat tolerance. SES1 is obviously induced by heat treatment and ses1 mutants are hypersensitive to heat stress. The unfolded protein response genes were up-regulated, while cytosolic protein response genes were down-regulated in ses1 after heat stress. Furthermore, ER stress sensor basic leucine zipper 28 (bZIP28) acts as the upstream transcriptional activator of SES1 by binding to its promoter region. These results provide new insights into heat stress responses and ER stress, and shed lights on the mechanism of SES1 in modulating heat resistance.

Hibiscus rosa-sinensis Linn., family Malvaceae, is an attractive horticultural plant originating from China. Five viruses infecting H. rosa-sinensis that have been characterized previously are Hibiscus chlorotic ringspot virus (HCRSV, genus Carmovirus), Hibiscus latent ringspot virus (HLRSV, genus Nepovirus), Hibiscus yellow mosaic virus (genus Tobamovirus), Eggplant mottled dwarf virus (EMDV, genus Nucleorhabdovirus), and Okra mosaic virus (OkMV, genus Tymovirus) (2). Recently, two novel tobamoviruses infecting H. rosa-sinensis were characterized in Singapore and Florida (1). In this study, viral symptoms were observed on H. rosa-sinensis in Nanyang City in Henan Province, China. The systemic symptoms included dark and light green mosaic in young leaves, leaf puckering and malformation on older leaves, and significant stunting. Rod-shaped virus particles were isolated from H. rosa-sinensis expressing systemic symptoms. The virus was transmitted mechanically to 10 species from three families. Symptoms expressed on these plants included systemic leaf chlorosis and distortion on Lycopersicum esculentum, systemic mosaic on Capsicum annuum, Nicotiana tabacum, and Physalis floridana, and systemic chlorosis on Glycine max. N. tabacum-Xanthi nc and Datura stramonium were asymptomatic. The virus also produced chlorotic and necrotic local lesions on Chenopodium quinoa, C. amaranticolor, and C. murale. The virus was propagated in L. esculentum, N. tabacum, and P. floridana. Virions purified from systemically infected N. tabacum contained a single-stranded RNA of approximately 6.4 kb and a coat protein (CP) of approximately 17.6 kDa. The double-stranded RNA profile revealed a single band of approximately 6.4 kb. Sap extracted from virus-infected plants reacted positive with an antiserum prepared against Tobacco mosaic virus (TMV) using an antigen-coated plate enzyme-linked immunosorbent assay. The CP gene was amplified by reverse transcription-polymerase chain reaction with primers specific to Tomato mosaic virus (ToMV) and sequence data obtained from the resulting amplification product. The CP gene consisting of 159 amino acids (GenBank Accession No. AY313136) shared 99.37% identity with the ToMV Queensland isolate (GenBank Accession No. AF332868). On the basis of biology, serology, properties of virions, and the sequence of the CP gene, we conclude that the virus isolated from H. rosa-sinensis in China is Tomato mosaic virus(ToMV). References: (1) S. Adkins et al. Plant Dis. 87:1190, 2003. (2) M. H. V. van Regenmortel et al., eds. Virus Taxonomy. 7th Report of the ICTV, Academic Press, NY, 2000.