STRESS INDUCED FACTOR 2 Regulates Arabidopsis Stomatal Immunity through Phosphorylation of the Anion Channel SLAC1.

Upon recognition of microbes, pattern recognition receptors (PRRs) activate pattern-triggered immunity. FLAGELLIN SENSING2 (FLS2) and BRASSINOSTEROID INSENSITIVE1-ASSOCIATED KINASE1 (BAK1) form a typical PRR complex that senses bacteria. Here, we report that the kinase activity of the malectin-like receptor-like kinase STRESS INDUCED FACTOR 2 (SIF2) is critical for Arabidopsis (Arabidopsis thaliana) resistance to bacteria by regulating stomatal immunity. SIF2 physically associates with the FLS2-BAK1 PRR complex and interacts with and phosphorylates the guard cell SLOW ANION CHANNEL1 (SLAC1), which is necessary for abscisic acid (ABA)-mediated stomatal closure. SIF2 is also required for the activation of ABA-induced S-type anion currents in Arabidopsis protoplasts, and SIF2 is sufficient to activate SLAC1 anion channels in Xenopus oocytes. SIF2-mediated activation of SLAC1 depends on specific phosphorylation of Ser 65. This work reveals that SIF2 functions between the FLS2-BAK1 initial immunity receptor complex and the final actuator SLAC1 in stomatal immunity.

The Arabidopsis Malectin-Like/LRR-RLK IOS1 Is Critical for BAK1-Dependent and BAK1-Independent Pattern-Triggered Immunity.

Plasma membrane-localized pattern recognition receptors (PRRs) such as FLAGELLIN SENSING2 (FLS2), EF-TU RECEPTOR (EFR), and CHITIN ELICITOR RECEPTOR KINASE1 (CERK1) recognize microbe-associated molecular patterns (MAMPs) to activate pattern-triggered immunity (PTI). A reverse genetics approach on genes responsive to the priming agent ß-aminobutyric acid (BABA) revealed IMPAIRED OOMYCETE SUSCEPTIBILITY1 (IOS1) as a critical PTI player. Arabidopsis thaliana ios1 mutants were hypersusceptible to Pseudomonas syringae bacteria. Accordingly, ios1 mutants showed defective PTI responses, notably delayed upregulation of the PTI marker gene FLG22-INDUCED RECEPTOR-LIKE KINASE1, reduced callose deposition, and mitogen-activated protein kinase activation upon MAMP treatment. Moreover, Arabidopsis lines overexpressing IOS1 were more resistant to bacteria and showed a primed PTI response. In vitro pull-down, bimolecular fluorescence complementation, coimmunoprecipitation, and mass spectrometry analyses supported the existence of complexes between the membrane-localized IOS1 and BRASSINOSTEROID INSENSITIVE1-ASSOCIATED KINASE1 (BAK1)-dependent PRRs FLS2 and EFR, as well as with the BAK1-independent PRR CERK1. IOS1 also associated with BAK1 in a ligand-independent manner and positively regulated FLS2-BAK1 complex formation upon MAMP treatment. In addition, IOS1 was critical for chitin-mediated PTI. Finally, ios1 mutants were defective in BABA-induced resistance and priming. This work reveals IOS1 as a novel regulatory protein of FLS2-, EFR-, and CERK1-mediated signaling pathways that primes PTI activation.

The Role of ENHANCED RESPONSES TO ABA1 (ERA1) in Arabidopsis Stomatal Responses Is Beyond ABA Signaling.

Proper stomatal responses are essential for plant function in an altered environment. The core signaling pathway for abscisic acid (ABA)-induced stomatal closure involves perception of the hormone that leads to the activation of guard cell anion channels by the protein kinase OPEN STOMATA1. Several other regulators are suggested to modulate the ABA signaling pathway, including the protein ENHANCED RESPONSE TO ABA1 (ERA1), that encodes the farnesyl transferase ß-subunit. The era1 mutant is hypersensitive to ABA during seed germination and shows a more closed stomata phenotype. Using a genetics approach with the double mutants era1 abi1-1 and era1 ost1, we show that while era1 suppressed the high stomatal conductance of abi1-1 and ost1, the ERA1 function was not required for stomatal closure in response to ABA and environmental factors. Further experiments indicated a role for ERA1 in blue light-induced stomatal opening. In addition, we show that ERA1 function in disease resistance was independent of its role in stomatal regulation. Our results indicate a function for ERA1 in stomatal opening and pathogen immunity.

The Arabidopsis malectin-like leucine-rich repeat receptor-like kinase IOS1 associates with the pattern recognition receptors FLS2 and EFR and is critical for priming of pattern-triggered immunity.

Plasma membrane-localized pattern recognition receptors such as FLAGELLIN SENSING2 (FLS2) and EF-TU RECEPTOR (EFR) recognize microbe-associated molecular patterns (MAMPs) to activate the first layer of plant immunity termed pattern-triggered immunity (PTI). A reverse genetics approach with genes responsive to the priming agent ß-aminobutyric acid (BABA) revealed IMPAIRED OOMYCETE SUSCEPTIBILITY1 (IOS1) as a critical PTI player. Arabidopsis thaliana ios1 mutants were hypersusceptible to Pseudomonas syringae bacteria. Accordingly, ios1 mutants demonstrated defective PTI responses, notably delayed upregulation of PTI marker genes, lower callose deposition, and mitogen-activated protein kinase activities upon bacterial infection or MAMP treatment. Moreover, Arabidopsis lines overexpressing IOS1 were more resistant to P. syringae and demonstrated a primed PTI response. In vitro pull-down, bimolecular fluorescence complementation, coimmunoprecipitation, and mass spectrometry analyses supported the existence of complexes between the membrane-localized IOS1 and FLS2 and EFR. IOS1 also associated with BRASSINOSTEROID INSENSITIVE1-ASSOCIATED KINASE1 (BAK1) in a ligand-independent manner and positively regulated FLS2/BAK1 complex formation upon MAMP treatment. Finally, ios1 mutants were defective in BABA-induced resistance and priming. This work reveals IOS1 as a regulatory protein of FLS2- and EFR-mediated signaling that primes PTI activation upon bacterial elicitation.

The Arabidopsis LecRK-VI.2 associates with the pattern-recognition receptor FLS2 and primes Nicotiana benthamiana pattern-triggered immunity.

Pattern-triggered immunity (PTI) is broad spectrum and manipulation of PTI is believed to represent an attractive way to engineer plants with broad-spectrum disease resistance. PTI is activated upon perception of microbe-associated molecular patterns (MAMPs) by pattern-recognition receptors (PRRs). We have recently demonstrated that the L-type lectin receptor kinase-VI.2 (LecRK-VI.2) positively regulates Arabidopsis thaliana PTI. Here we show through in vitro pull-down, bimolecular fluorescence complementation and co-immunoprecipitation analyses that LecRK-VI.2 associates with the PRR FLS2. We also demonstrated that LecRK-VI.2 from the cruciferous plant Arabidopsis remains functional after interfamily transfer to the Solanaceous plant Nicotiana benthamiana. Wild tobacco plants ectopically expressing LecRK-VI.2 were indeed more resistant to virulent hemi-biotrophic and necrotrophic bacteria, but not to the fungal pathogen Botrytis cinerea suggesting that, as with Arabidopsis, the LecRK-VI.2 protective effect in N. benthamiana is bacteria specific. Ectopic expression of LecRK-VI.2 in N. benthamiana primed PTI-mediated reactive oxygen species production, mitogen-activated protein kinase (MAPK) activity, callose deposition and gene expression upon treatment with the MAMP flagellin. Our findings identified LecRK-VI.2 as a member of the FLS2 receptor complex and suggest that heterologous expression of components of PRR complexes can be used as tools to engineer plant disease resistance to bacteria.

A Ratiometric Fluorescent Sensor for Penicillin G Based on Color-Tunable Gold-Silver Nanoclusters.

Excessive administration of penicillin G and improper disposal of its residues pose a serious risk to human health; therefore, the development of convenient methods for monitoring penicillin G levels in products is essential. Herein, novel gold-silver nanoclusters (AuAgNCs) were synthesized using chicken egg white and 6-aza-2-thiothymine as dual ligands with strong yellow fluorescence at 509 and 689 nm for the highly selective detection of penicillin G. The AuAgNCs were characterized using transmission electron microscopy, X-ray photoelectron spectroscopy, ultraviolet-visible absorption spectrophotometry, and fluorescence spectrophotometry. Under optimum conditions, the fluorescence intensity decreased linearly with the concentration of penicillin G from 0.2 to 6 µM, with a low detection limit of 18 nM. Real sample analyses indicated that a sensor developed using the AuAgNCs could detect penicillin G in urine and water samples within 10 min, with the recoveries ranging from 99.7 to 104.0%. The particle size of the AuAgNCs increased from 1.80 to 9.06 nm in the presence of penicillin G. We believe the aggregation-induced quenching of the fluorescence of the AuAgNCs was the main mechanism for the detection of penicillin G. These results demonstrate the ability of our sensor for monitoring penicillin G levels in environmental and clinic samples.

The diversity of cultivable endophytic fungi of the sand coast plant Ipomoeapes-caprae in Taiwan.

Background: Ipomoeapes-caprae is a plant of sand coasts and it can tolerate stresses, such as high salinity, strong wind and sand movements and lack of nutrients. It plays an important role in coast protection and preventing erosion. Fungal endophytes show high biodiversity and have a strong influence on the survival of plants under different stress factors. Although this plant is important for sand coast ecosystems, little is known about the associated fungi. In this study, we isolated and identified endophytic fungi of Ipomoeapes-caprae, a dominant plant along the shore of Taiwan. The dataset contains 896 records, which correspond to 177 species. The geographical scope of the dataset covers the northern subtropical area of the main island of Taiwan, with its sand coasts in New Taipei, Taoyuan, Hsinchu and Taichung and two botanical gardens in Taipei and Taichung. The detailed original data of fungal diversity are rarely publicly shared under strictly formalised and, thus, reusable standards. As an example for such an approach, the complete occurrence dataset was made available in the Darwin Core Archive format via the Global Biodiversity Information Facility (GBIF) under Version 1.13, Taiwan Biodiversity Information Facility (TaiBIF) https://doi.org/10.15468/9h9rcg. In this first data paper on endophytic fungi, the scientific name and associated DNA sequence in the dataset were directly linked to other free online resource (Index Fungorum, GenBank), which shows the potential of GBIF for linking together different online data repositories. New information: We describe a dataset, in which the diversity of endophytic fungi of the sand coast plant Ipomoeapes-caprae in Taiwan was investigated.

Frequency-Domain Analysis for Accurate and Robust Gait Cycle Time Detection with Clinical Data.

Gait tasks have become a topic of increasing inter-est in biological engineering research in recent years. One way to obtain the gait cycle time (GCT) is to analyze a subject's gait acceleration signal as recorded by an inertial measurement unit (IMU) [1]. An accurate peak detection of the IMU acceleration has thus become a requirement for GCT analysis. This study proposes a detection procedure for accurately detecting the peaks in a noisy IMU acceleration signal based on a frequency-domain analysis of the acceleration.

Diversity of Endophytic Fungi of the Coastal Plant Vitex rotundifolia in Taiwan.

Vitex rotundifolia L. f. (Lamiaceae), which commonly grows at sand coasts, is important for coast protection and the prevention of erosion. However, the diversity and roles of fungi associated with this plant remain unclear. A total of 1,052 endophytic isolates from 1,782 plants tissues from two sand beaches in northern Taiwan were classified into 76 morphospecies based on culture morphology and ITS or LSU rRNA gene sequence comparisons. Critical species were further identified using protein gene sequences and microscopy. Most of the isolates at both sites belonged to the phylum Ascomycota, with Pleosporales having the most species (15 species). The largest number of isolates (47.7%) was from the stems, followed by the roots (22.5%), leaves (16.6%), and branches (13.1%). The three species with the highest isolation frequencies at both sites were Alternaria alternata, Aspergillus terreus, and an undescribed species of Alpestrisphaeria. A. terreus was found in all organs. A. alternata was detected in all organs, except the roots. Alpestrisphaeria sp. was only found in the roots and stems. In the stems and roots, strain numbers from cortical tissues were approximately two-fold higher than those from the corresponding woody tissue. The overall colonization rate in the stems was significantly higher than those that in the roots and leaves. The majority of fungi appeared to be saprobes, which may play important roles in nutrient recycling during sand burial and mediate further stress factors in the coastal habitat.

Enhanced Arabidopsis pattern-triggered immunity by overexpression of cysteine-rich receptor-like kinases.

Upon recognition of microbe-associated molecular patterns (MAMPs) such as the bacterial flagellin (or the derived peptide flg22) by pattern-recognition receptors (PRRs) such as the FLAGELLIN SENSING2 (FLS2), plants activate the pattern-triggered immunity (PTI) response. The L-type lectin receptor kinase-VI.2 (LecRK-VI.2) is a positive regulator of Arabidopsis thaliana PTI. Cysteine-rich receptor-like kinases (CRKs) possess two copies of the C-X8-C-X2-C (DUF26) motif in their extracellular domains and are thought to be involved in plant stress resistance, but data about CRK functions are scarce. Here, we show that Arabidopsis overexpressing the LecRK-VI.2-responsive CRK4, CRK6, and CRK36 demonstrated an enhanced PTI response and were resistant to virulent bacteria Pseudomonas syringae pv. tomato DC3000. Notably, the flg22-triggered oxidative burst was primed in CRK4, CRK6, and CRK36 transgenics and up-regulation of the PTI-responsive gene FLG22-INDUCED RECEPTOR-LIKE 1 (FRK1) was potentiated upon flg22 treatment in CRK4 and CRK6 overexpression lines or constitutively increased by CRK36 overexpression. PTI-mediated callose deposition was not affected by overexpression of CRK4 and CRK6, while CRK36 overexpression lines demonstrated constitutive accumulation of callose. In addition, Pst DC3000-mediated stomatal reopening was blocked in CRK4 and CRK36 overexpression lines, while overexpression of CRK6 induced constitutive stomatal closure suggesting a strengthening of stomatal immunity. Finally, bimolecular fluorescence complementation and co-immunoprecipitation analyses in Arabidopsis protoplasts suggested that the plasma membrane localized CRK4, CRK6, and CRK36 associate with the PRR FLS2. Association with FLS2 and the observation that overexpression of CRK4, CRK6, and CRK36 boosts specific PTI outputs and resistance to bacteria suggest a role for these CRKs in Arabidopsis innate immunity.

Sarocladium spinificis, a new endophytic species from the coastal grass Spinifex littoreus in Taiwan.

BACKGROUND: Sarocladium species are frequently associated with grasses as saprobes, parasites, and mutualistic endophytes. A species of Sarocladium (anamorphic Hypocreales) was isolated as endophytic fungus from the coastal grass Spinifex littoreus (Poaceae). RESULTS: According to characterization by LSU and ITS rDNA sequences and culture morphology and micromorphology, the species differed from the species hitherto described in Sarocladium. A key to the known species of Sarocladium is given. CONCLUSIONS: Sarocladium spinificis is proposed as a new species. LSU rDNA sequences and conidiophore branching and conidium size are useful characters for distinguishing between species of Sarocladium.