NKS1/ELMO4 is an integral protein of a pectin synthesis protein complex and maintains Golgi morphology and cell adhesion in Arabidopsis.

Adjacent plant cells are connected by specialized cell wall regions, called middle lamellae, which influence critical agricultural characteristics, including fruit ripening and organ abscission. Middle lamellae are enriched in pectin polysaccharides, specifically homogalacturonan (HG). Here, we identify a plant-specific Arabidopsis DUF1068 protein, called NKS1/ELMO4, that is required for middle lamellae integrity and cell adhesion. NKS1 localizes to the Golgi apparatus and loss of NKS1 results in changes to Golgi structure and function. The nks1 mutants also display HG deficient phenotypes, including reduced seedling growth, changes to cell wall composition, and tissue integrity defects. These phenotypes are comparable to qua1 and qua2 mutants, which are defective in HG biosynthesis. Notably, genetic interactions indicate that NKS1 and the QUAs work in a common pathway. Protein interaction analyses and modeling corroborate that they work together in a stable protein complex with other pectin-related proteins. We propose that NKS1 is an integral part of a large pectin synthesis protein complex and that proper function of this complex is important to support Golgi structure and function.

Barley AGO4 proteins show overlapping functionality with distinct small RNA-binding properties in heterologous complementation.

KEY MESSAGE: Barley AGO4 proteins complement expressional changes of epigenetically regulated genes in Arabidopsis ago4-3 mutant and show a distinct affinity for the 5' terminal nucleotide of small RNAs, demonstrating functional conservation and divergence. The function of Argonaute 4 (AGO4) in Arabidopsis thaliana has been extensively characterized; however, its role in monocots, which have large genomes abundantly supplemented with transposable elements (TEs), remains elusive. The study of barley AGO4 proteins can provide insights into the conserved aspects of RNA-directed DNA methylation (RdDM) and could also have further applications in the field of epigenetics or crop improvement. Bioinformatic analysis of RNA sequencing data identified two active AGO4 genes in barley, HvAGO4a and HvAGO4b. These genes function similar to AtAGO4 in an Arabidopsis heterologous complementation system, primarily binding to 24-nucleotide long small RNAs (sRNAs) and triggering methylation at specific target loci. Like AtAGO4, HvAGO4B exhibits a preference for binding sRNAs with 5' adenine residue, while also accepting 5' guanine, uracil, and cytosine residues. In contrast, HvAGO4A selectively binds only sRNAs with a 5' adenine residue. The diverse binding capacity of barley AGO4 proteins is reflected in TE-derived sRNAs and in their varying abundance. Both barley AGO4 proteins effectively restore the levels of extrachromosomal DNA and transcript abundancy of the heat-activated ONSEN retrotransposon to those observed in wild-type Arabidopsis plants. Our study provides insight into the distinct binding specificities and involvement in TE regulation of barley AGO4 proteins in Arabidopsis by heterologous complementation.

Exo84c-regulated degradation is involved in the normal self-incompatible response in Brassicaceae.

The self-incompatibility system evolves in angiosperms to promote cross-pollination by rejecting self-pollination. Here, we show the involvement of Exo84c in the SI response of both Brassica napus and Arabidopsis. The expression of Exo84c is specifically elevated in stigma during the SI response. Knocking out Exo84c in B. napus and SI Arabidopsis partially breaks down the SI response. The SI response inhibits both the protein secretion in papillae and the recruitment of the exocyst complex to the pollen-pistil contact sites. Interestingly, these processes can be partially restored in exo84c SI Arabidopsis. After incompatible pollination, the turnover of the exocyst-labeled compartment is enhanced in papillae. However, this process is perturbed in exo84c SI Arabidopsis. Taken together, our results suggest that Exo84c regulates the exocyst complex vacuolar degradation during the SI response. This process is likely independent of the known SI pathway in Brassicaceae to secure the SI response.

Pseudomonas putida configures Arabidopsis root architecture through modulating the sensing systems for phosphate and iron acquisition.

Iron (Fe) and phosphate (Pi) are two essential nutrients that are poorly available in the soil and should be supplemented either as fertilizers or organic amendments to sustain crop production. Currently, determining how rhizosphere bacteria contribute to plant mineral nutrient acquisition is an area of growing interest regarding its potential application in agriculture. The aim of this study was to investigate the influence of root colonization by Pseudomonas putida for Arabidopsis growth through Fe and Pi nutritional signaling. We found that root colonization by the bacterium inhibits primary root elongation and promotes the formation of lateral roots. These effects could be related to higher expression of two Pi starvation-induced genes and AtPT1, the major Pi transporter in root tips. In addition, P. putida influenced the accumulation of Fe in the root and the expression of different elements of the Fe uptake pathway. The loss of function of the protein ligase BRUTUS (BTS), and the bHLH transcription factors POPEYE (PYE) and IAA-LEUCINE RESISTANT3 (ILR3) compromised the root branching stimulation triggered by bacterial inoculation while the leaf chlorosis in the fit1 and irt1-1 mutant plants grown under standard conditions could be bypassed by P. putida inoculation. The WT and both mutant lines showed similar Fe accumulation in roots. P. putida repressed the expression of the IRON-REGULATED TRANSPORTER 1 (IRT1) gene suggesting that the bacterium promotes an alternative Fe uptake mechanism. These results open the door for the use of P. putida to enhance nutrient uptake and optimize fertilizer usage by plants.

Exogenous application of pectin triggers stomatal closure and immunity in Arabidopsis.

Pectin has been extensively studied in animal immunity, and exogenous pectin as a food additive can provide protection against inflammatory bowel disease. However, the utility of pectin to improve immunity in plants is still unstudied. Here, we found exogenous application of pectin triggered stomatal closure in Arabidopsis in a dose- and time-dependent manner. Additionally, pectin activated peroxidase and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase to produce reactive oxygen species (ROS), which subsequently increased cytoplasmic Ca2+ concentration ([Ca2+ ]cyt ) and was followed by nitric oxide (NO) production, leading to stomatal closure in an abscisic acid (ABA) and salicylic acid (SA) signalling-dependent mechanism. Furthermore, pectin enhanced the disease resistance to Pseudomonas syringae pv. tomato DC3000 (Pst DC3000) with mitogen-activated protein kinases (MPKs) MPK3/6 activated and upregulated expression of defence-responsive genes in Arabidopsis. These results suggested that exogenous pectin-induced stomatal closure was associated with ROS and NO production regulated by ABA and SA signalling, contributing to defence against Pst DC3000 in Arabidopsis.

Pectic-AGP is a major form of Arabidopsis AGPs.

As a key component in cell walls of numerous organisms ranging from green algae to higher plants, AGPs play principal roles in many biological processes such as cell-cell adhesion and regulating Ca2+ signaling pathway as a Ca2+-capacitor. Consistently, AGP structures vary from species to species and from tissue to tissue. To understand the functions of AGPs, it is vital to know their structural differences relative to their location in the plant. Thus, AGPs were purified from different Arabidopsis tissues. Analyses of these AGPs demonstrated that the AGPs comprised covalently linked pectin and AGP, referred to as pectic-AGPs. Importantly, these pectic-AGPs were glycosylated with a remarkable variety of polysaccharides including homogalacturonan, rhamnogalacturonan-I, and type II arabinogalactan at different ratios and lengths. This result not only suggests that pectic-AGP is a major form of Arabidopsis AGPs, but also supports AGPs serve as crosslinkers covalently connecting pectins with structures tailored for tissue-specific functions.

Changes in Gene Expression After Exposing Arabidopsis thaliana Plants to Nanosecond High Amplitude Electromagnetic Field Pulses.

The biological effects of exposure to electromagnetic fields due to wireless technologies and connected devices are a subject of particular research interest. Ultrashort high-amplitude electromagnetic field pulses delivered to biological samples using immersed electrodes in a dedicated cuvette have widely demonstrated their effectiveness in triggering several cell responses including increased cytosolic calcium concentration and reactive oxygen species (ROS) production. In contrast, the effects of these pulses are poorly documented when electromagnetic pulses are delivered through an antenna. Here we exposed Arabidopsis thaliana plants to 30,000 pulses (237 kV m-1 , 280 ps rise-time, duration of 500 ps) emitted through a Koshelev antenna and monitored the consequences of electromagnetic fields exposure on the expression levels of several key genes involved in calcium metabolism, signal transduction, ROS, and energy status. We found that this treatment was mostly unable to trigger significant changes in the messenger RNA accumulation of calmodulin, Zinc-Finger protein ZAT12, NADPH oxidase/respiratory burst oxidase homolog (RBOH) isoforms D and F, Catalase (CAT2), glutamate-cystein ligase (GSH1), glutathione synthetase (GSH2), Sucrose non-fermenting-related Kinase 1 (SnRK1) and Target of rapamycin (TOR). In contrast, Ascorbate peroxidases APX-1 and APX-6 were significantly induced 3 h after the exposure. These results suggest that this treatment, although quite strong in amplitude, is mostly ineffective in inducing biological effects at the transcriptional level when delivered by an antenna. © 2023 The Authors. Bioelectromagnetics published by Wiley Periodicals LLC on behalf of Bioelectromagnetics Society.

Near-infrared light and PIF4 promote plant antiviral defense by enhancing RNA interference.

The molecular mechanism underlying phototherapy and light treatment, which utilize various wavelength spectra of light, including near-infrared (NIR), to cure human and plant diseases, is obscure. Here we revealed that NIR light confers antiviral immunity by positively regulating PHYTOCHROME-INTERACTING FACTOR 4 (PIF4)-activated RNA interference (RNAi) in plants. PIF4, a central transcription factor involved in light signaling, accumulates to high levels under NIR light in plants. PIF4 directly induces the transcription of two essential components of RNAi, RNA-DEPENDENT RNA POLYMERASE 6 (RDR6) and ARGONAUTE 1 (AGO1), which play important roles in resistance to both DNA and RNA viruses. Moreover, the pathogenic determinant ßC1 protein, which is evolutionarily conserved and encoded by betasatellites, interacts with PIF4 and inhibits its positive regulation of RNAi by disrupting PIF4 dimerization. These findings shed light on the molecular mechanism of PIF4-mediated plant defense and provide a new perspective for the exploration of NIR antiviral treatment.

Redundant function of the Arabidopsis phosphatidylinositol 4-phosphate 5-kinase genes PIP5K4-6 is essential for pollen germination.

Phosphatidylinositol 4-phosphate 5-kinase (PIP5K) is a key enzyme producing the signaling lipid phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2 ] in eukaryotes. Although PIP5K genes are reported to be involved in pollen tube germination and growth, the essential roles of PIP5K in these processes remain unclear. Here, we performed a comprehensive genetic analysis of the Arabidopsis thaliana PIP5K4, PIP5K5, and PIP5K6 genes and revealed that their redundant function is essential for pollen germination. Pollen with the pip5k4pip5k5pip5k6 triple mutation was sterile, while pollen germination efficiency and pollen tube growth were reduced in the pip5k6 single mutant and further reduced in the pip5k4pip5k6 and pip5k5pip5k6 double mutants. YFP-fusion proteins, PIP5K4-YFP, PIP5K5-YFP, and PIP5K6-YFP, which could rescue the sterility of the triple mutant pollen, preferentially localized to the tricolpate aperture area and the future germination site on the plasma membrane prior to germination. Triple mutant pollen grains under the germination condition, in which spatiotemporal localization of the PtdIns(4,5)P2 fluorescent marker protein 2xmCHERRY-2xPHPLC as seen in the wild type was abolished, exhibited swelling and rupture of the pollen wall, but neither the conspicuous protruding site nor site-specific deposition of cell wall materials for germination. These data indicate that PIP5K4-6 and their product PtdIns(4,5)P2 are essential for pollen germination, possibly through the establishment of the germination polarity in a pollen grain.

Acetylation of homogalacturonan and rhamnogalacturonan-I is catalyzed by a suite of trichome birefringence-like proteins.

Plant cell wall polysaccharides, including xylan, mannan, xyloglucan, and pectins, are often acetylated and members of the domain of unknown function 231 (DUF231)/trichome birefringence-like (TBL) family have been shown to be O-acetyltransferases mediating the acetylation of xylan, mannan, and xyloglucan. However, little is known about the O-acetyltransferases responsible for pectin acetylation. In this report, we biochemically characterized a suite of Arabidopsis DUF231/TBL proteins for their roles in pectin acetylation. We generated 24 TBL recombinant proteins in mammalian cells and demonstrated that 10 of them were able to transfer acetyl groups from acetyl-CoA onto the pectins homogalacturonan (HG) or rhamnogalacturonan-I (RG-I), and thus were named pectin O-acetyltransferase 1 to 10 (POAT1 to 10). It was found that POAT2,4,9,10 specifically acetylated HG and POAT5,6 acetylated RG-I, whereas POAT1,3,7,8 could act on both HG and RG-I. The acetylation of HG and RG-I by POATs was further corroborated by hydrolysis with pectin acetylesterases and by nuclear magnetic resonance spectroscopy. In addition, mutations of the conserved GDS and DXXH motifs in POAT3 and POAT8 were shown to lead to a loss of their ability to acetylate HG and RG-I. Furthermore, simultaneous RNA interference downregulation of POAT1,3,6,7,8 resulted in reduced cell expansion, impaired plant growth, and decreased pectin acetylation. Together, our findings indicate that these POATs are pectin O-acetyltransferases involved in acetylation of the pectin polysaccharides HG and RG-I.

Onion cryptochrome 1 (AcCRY1) regulates photomorphogenesis and photoperiod flowering in Arabidopsis and exploration of its functional mechanisms under blue light.

Cryptochromes (CRYs), as blue-light photoreceptors, play a crucial role in regulating flowering time and hypocotyl and cotyledon development. Their physiological functions have been extensively studied in various plant species. However, research on onions remains limited. In this study, we identified AcCRY1 and conducted preliminary investigations into its function. Our results demonstrate that AcCRY1 possesses a conserved domain typical of cryptochromes with high homology to those found in monocots. Furthermore, we examined the expression level of AcCRY1 in onion. The green tissues is significantly higher compared to non-green tissues, and it exhibits a significant response to blue-light induction. AcCRY1 demonstrates cytoplasmic localization under blue-light conditions, while it localizes in the nucleus during darkness, indicating a strong dependence on blue-light for its subcellular distribution. In comparison to cry1, overexpression of AcCRY1 leads to a significant shorten in seedling hypocotyl length, notable expansion of cotyledons, and acceleration of flowering time. The yeast two-hybrid experiment demonstrated the in vitro interaction between AcCRY1, AcCOP1, and AcSPA1. Additionally, BIFC analysis confirmed their interaction in Onion epidermis. Notably, under blue-light conditions, a significantly enhanced binding activity was observed compared to dark conditions. These findings establish a functional foundation for the regulatory role of AcCRY1 in important physiological processes of onion and provide initial insights into the underlying molecular mechanisms.

Mitigating cadmium accumulation in dicotyledonous vegetables by iron fertilizer through inhibiting Fe transporter IRT1-mediated Cd uptake.

The solubility of cadmium (Cd) in soil and its transfer to plants are influenced by soil pH. While increasing soil pH reduces Cd solubility and accumulation in rice plants grown in acidic soils, its effect on Cd accumulation in vegetables remains inconclusive. Here, we investigated the impact of soil pH on Cd accumulation in dicotyledonous vegetables and elucidated the underlying molecular mechanisms. Soils collected from various locations were supplemented with varying quantities of lime to achieve soil pH values of around 5.0, 6.0, 7.0, and 8.0. Raising soil pH from around 5.0 to 8.0 markedly decreased extractable Cd. However, increasing soil pH tended to promote shoot Cd accumulation in dicotyledonous vegetable species including lettuce, pakchoi, and Chinese cabbage, and the model dicotyledonous plant Arabidopsis thaliana. Conversely, soil pH increase resulted in a monotonic decrease in rice Cd accumulation. In our hydroponic experiments, we discovered that iron (Fe) deficiency substantially increased Cd uptake and accumulation in dicotyledonous plants but not in rice. Increasing soil pH reduced soil Fe availability and induced the Fe transporter gene IRT1 expression in dicotyledonous vegetables roots, which led to an increase in IRT1-mediated Cd uptake and subsequently increased Cd accumulation as soil pH increases. A comprehensive model incorporating extractable Cd and root IRT1 expression better explained Cd accumulation in vegetable shoots. The application of 50 mg/kg of Fe fertilizer in neutral or alkaline soils resulted in a significant reduction in Cd accumulation by 34-58% in dicotyledonous vegetables. These findings reveal that increasing soil pH has two opposite effects, decreasing soil Cd availability while promoting Cd uptake through IRT1 upregulation, reconciling the inconsistency in its effect on Cd accumulation in dicotyledonous plants. Our findings provide important insights for understanding the factors affecting Cd uptake in plants and offer a practical solution to mitigate Cd contamination in vegetables.

PHR1 involved in the regulation of low phosphate-induced leaf senescence by modulating phosphorus homeostasis in Arabidopsis.

Phosphorus (P) is a crucial macronutrient for plant growth, development, and reproduction. The effects of low P (LP) stress on leaf senescence and the role of PHR1 in LP-induced leaf senescence are still unknown. Here, we report that PHR1 plays a crucial role in LP-induced leaf senescence, showing delayed leaf senescence in phr1 mutant and accelerated leaf senescence in 35S:PHR1 transgenic Arabidopsis under LP stress. The transcriptional profiles indicate that 763 differentially expressed SAGs (DE-SAGs) were upregulated and 134 DE-SAGs were downregulated by LP stress. Of the 405 DE-SAGs regulated by PHR1, 27 DE-SAGs were involved in P metabolism and transport. PHR1 could bind to the promoters of six DE-SAGs (RNS1, PAP17, SAG113, NPC5, PLDζ2, and Pht1;5), and modulate them in LP-induced senescing leaves. The analysis of RNA content, phospholipase activity, acid phosphatase activity, total P and phosphate content also revealed that PHR1 promotes P liberation from senescing leaves and transport to young tissues under LP stress. Our results indicated that PHR1 is one of the crucial modulators for P recycling and redistribution under LP stress, and the drastic decline of P level is at least one of the causes of early senescence in P-deficient leaves.

Antagonistic MADS-box transcription factors SEEDSTICK and SEPALLATA3 form a transcriptional regulatory network that regulates seed oil accumulation.

Transcriptional regulation is essential for balancing multiple metabolic pathways that influence oil accumulation in seeds. Thus far, the transcriptional regulatory mechanisms that govern seed oil accumulation remain largely unknown. Here, we identified the transcriptional regulatory network composed of MADS-box transcription factors SEEDSTICK (STK) and SEPALLATA3 (SEP3), which bridges several key genes to regulate oil accumulation in seeds. We found that STK, highly expressed in the developing embryo, positively regulates seed oil accumulation in Arabidopsis (Arabidopsis thaliana). Furthermore, we discovered that SEP3 physically interacts with STK in vivo and in vitro. Seed oil content is increased by the SEP3 mutation, while it is decreased by SEP3 overexpression. The chromatin immunoprecipitation, electrophoretic mobility shift assay, and transient dual-luciferase reporter assays showed that STK positively regulates seed oil accumulation by directly repressing the expression of MYB5, SEP3, and SEED FATTY ACID REDUCER 4 (SFAR4). Moreover, genetic and molecular analyses demonstrated that STK and SEP3 antagonistically regulate seed oil production and that SEP3 weakens the binding ability of STK to MYB5, SEP3, and SFAR4. Additionally, we demonstrated that TRANSPARENT TESTA 8 (TT8) and ACYL-ACYL CARRIER PROTEIN DESATURASE 3 (AAD3) are direct targets of MYB5 during seed oil accumulation in Arabidopsis. Together, our findings provide the transcriptional regulatory network antagonistically orchestrated by STK and SEP3, which fine tunes oil accumulation in seeds.

Extracellular pectin-RALF phase separation mediates FERONIA global signaling function.

The FERONIA (FER)-LLG1 co-receptor and its peptide ligand RALF regulate myriad processes for plant growth and survival. Focusing on signal-induced cell surface responses, we discovered that intrinsically disordered RALF triggers clustering and endocytosis of its cognate receptors and FER- and LLG1-dependent endocytosis of non-cognate regulators of diverse processes, thus capable of broadly impacting downstream responses. RALF, however, remains extracellular. We demonstrate that RALF binds the cell wall polysaccharide pectin. They phase separate and recruit FER and LLG1 into pectin-RALF-FER-LLG1 condensates to initiate RALF-triggered cell surface responses. We show further that two frequently encountered environmental challenges, elevated salt and temperature, trigger RALF-pectin phase separation, promiscuous receptor clustering and massive endocytosis, and that this process is crucial for recovery from stress-induced growth attenuation. Our results support that RALF-pectin phase separation mediates an exoskeletal mechanism to broadly activate FER-LLG1-dependent cell surface responses to mediate the global role of FER in plant growth and survival.

Heterologous Expression of Two Brassica campestris CCCH Zinc-Finger Proteins in Arabidopsis Induces Cytoplasmic Foci and Causes Pollen Abortion.

The membrane-less organelles in cytoplasm that are presented as cytoplasmic foci were successively identified. Although multiple CCCH zinc-finger proteins have been found to be localized in cytoplasmic foci, the relationship between their specific localization and functions still needs further clarification. Here, we report that the heterologous expression of two Brassica campestris CCCH zinc-finger protein genes (BcMF30a and BcMF30c) in Arabidopsis thaliana can affect microgametogenesis by involving the formation of cytoplasmic foci. By monitoring the distribution of proteins and observing pollen phenotypes, we found that, when these two proteins were moderately expressed in pollen, they were mainly dispersed in the cytoplasm, and the pollen developed normally. However, high expression induced the assembly of cytoplasmic foci, leading to pollen abortion. These findings suggested that the continuous formation of BcMF30a/BcMF30c-associated cytoplasmic foci due to high expression was the inducement of male sterility. A co-localization analysis further showed that these two proteins can be recruited into two well-studied cytoplasmic foci, processing bodies (PBs), and stress granules (SGs), which were confirmed to function in mRNA metabolism. Together, our data suggested that BcMF30a and BcMF30c play component roles in the assembly of pollen cytoplasmic foci. Combined with our previous study on the homologous gene of BcMF30a/c in Arabidopsis, we concluded that the function of these homologous genes is conserved and that cytoplasmic foci containing BcMF30a/c may participate in the regulation of gene expression in pollen by regulating mRNA metabolism.

Plant cell wall patterning and expansion mediated by protein-peptide-polysaccharide interaction.

Assembly of cell wall polysaccharides into specific patterns is required for plant growth. A complex of RAPID ALKALINIZATION FACTOR 4 (RALF4) and its cell wall-anchored LEUCINE-RICH REPEAT EXTENSIN 8 (LRX8)-interacting protein is crucial for cell wall integrity during pollen tube growth, but its molecular connection with the cell wall is unknown. Here, we show that LRX8-RALF4 complexes adopt a heterotetrametric configuration in vivo, displaying a dendritic distribution. The LRX8-RALF4 complex specifically interacts with demethylesterified pectins in a charge-dependent manner through RALF4's polycationic surface. The LRX8-RALF4-pectin interaction exerts a condensing effect, patterning the cell wall's polymers into a reticulated network essential for wall integrity and expansion. Our work uncovers a dual structural and signaling role for RALF4 in pollen tube growth and in the assembly of complex extracellular polymers.

TFIIB-Related Protein BRP5/PTF2 Is Required for Both Male and Female Gametogenesis and for Grain Formation in Rice.

Transcription factor IIB (TFIIB) is a general transcription factor for RNA polymerase II, exerting its influence across various biological contexts. In the majority of eukaryotes, TFIIB typically has two homologs, serving as general transcription factors for RNA polymerase I and III. In plants, however, the TFIIB-related protein family has expanded greatly, with 14 and 9 members in Arabidopsis and rice, respectively. BRP5/pollen-expressed transcription factor 2 (PTF2) proteins belong to a subfamily of TFIIB-related proteins found only in plants and algae. The prior analysis of an Arabidopsis atbrp5 mutant, characterized by a T-DNA insertion at the 5' untranslated region, demonstrated the essential role of BRP5/PTF2 during the process of pollen germination and embryogenesis in Arabidopsis. Using a rice transformation system based on CRISPR/Cas9 technology, we have generated transgenic rice plants containing loss-of-function frameshift mutations in the BRP5/PTF2 gene. Unlike in the Arabidopsis atbrp5 mutant, the brp5/ptf2 frameshift mutations were not transmitted to progeny in rice, indicating an essential role of BRP5/PTF2 in both male and female gamete development or viability. The silencing of rice BRP5/PTF2 expression through RNA interference (RNAi) had little effect on vegetative growth and panicle formation but strongly affected pollen development and grain formation. Genetic analysis revealed that strong RNAi silencing of rice BRP5/PTF2 was still transmissible to progeny almost exclusively through female gametes, as found in the Arabidopsis atbrp5 knockdown mutant. Thus, reduced rice BRP5/PTF2 expression impacted pollen preferentially by interfering with male gamete development or viability. Drawing upon these findings, we posit that BRP5/PTF2 assumes a distinct and imperative function in the realm of plant sexual reproduction.

Xylobiose treatment triggers a defense-related response and alters cell wall composition.

Plant cell wall-derived oligosaccharides, i.e., damage-associated molecular patterns (DAMPs), could be generated after pathogen attack or during normal plant development, perceived by cell wall receptors, and can alter immunity and cell wall composition. Therefore, we hypothesised that xylo-oligosaccharides (XOS) could act as an elicitor and trigger immune responses. To test this, we treated Arabidopsis with xylobiose (XB) and investigated different parameters. XB-treatment significantly triggered the generation of reactive oxygen species (ROS), activated MAPK protein phosphorylation, and induced callose deposition. The combination of XB (DAMP) and flg22 a microbe-associated molecular pattern (MAMP) further enhanced ROS response and gene expression of PTI marker genes. RNA sequencing analysis revealed that more genes were differentially regulated after 30 min compared to 24 h XB-treated leaves, which correlated with ROS response. Increased xylosidase activity and soluble xylose level after 30 min and 3 h of XB-treatment were observed which might have weakened the DAMP response. However, an increase in total cell wall sugar and a decrease in uronic acid level was observed at both 30 min and 24 h. Additionally, arabinose, rhamnose, and xylose levels were increased in 30 min, and glucose was increased in 24 h compared to mock-treated leaves. The level of jasmonic acid, abscisic acid, auxin, and cytokinin were also affected after XB treatment. Overall, our data revealed that the shortest XOS can act as a DAMP, which triggers the PTI response and alters cell wall composition and hormone level.

PLC1 mediated Cycloastragenol-induced stomatal movement by regulating the production of NO in Arabidopsis thaliana.

BACKGROUND: Astragalus grows mainly in drought areas. Cycloastragenol (CAG) is a tetracyclic triterpenoid allelochemical extracted from traditional Chinese medicine Astragalus root. Phospholipase C (PLC) and Gα-submit of the heterotrimeric G-protein (GPA1) are involved in many biotic or abiotic stresses. Nitric oxide (NO) is a crucial gas signal molecule in plants. RESULTS: In this study, using the seedlings of Arabidopsis thaliana (A. thaliana), the results showed that low concentrations of CAG induced stomatal closure, and high concentrations inhibited stomatal closure. 30 µmol·L-1 CAG significantly increased the relative expression levels of PLC1 and GPA1 and the activities of PLC and GTP hydrolysis. The stomatal aperture of plc1, gpa1, and plc1/gpa1 was higher than that of WT under CAG treatment. CAG increased the fluorescence intensity of NO in guard cells. Exogenous application of c-PTIO to WT significantly induced stomatal aperture under CAG treatment. CAG significantly increased the relative expression levels of NIA1 and NOA1. Mutants of noa1, nia1, and nia2 showed that NO production was mainly from NOA1 and NIA1 by CAG treatment. The fluorescence intensity of NO in guard cells of plc1, gpa1, and plc1/gpa1 was lower than WT, indicating that PLC1 and GPA1 were involved in the NO production in guard cells. There was no significant difference in the gene expression of PLC1 in WT, nia1, and noa1 under CAG treatment. The gene expression levels of NIA1 and NOA1 in plc1, gpa1, and plc1/gpa1 were significantly lower than WT, indicating that PLC1 and GPA1 were positively regulating NO production by regulating the expression of NIA1 and NOA1 under CAG treatment. CONCLUSIONS: These results suggested that the NO accumulation was essential to induce stomatal closure under CAG treatment, and GPA1 and PLC1 acted upstream of NO.