Platelet Membrane-Coated Curcumin-PLGA Nanoparticles Promote Astrocyte-Neuron Transdifferentiation for Intracerebral Hemorrhage Treatment.

Intracerebral hemorrhage (ICH) is a hemorrhagic disease with high mortality and disability rates. Curcumin is a promising drug for ICH treatment due to its multiple biological activities, but its application is limited by its poor watersolubility and instability. Herein, platelet membrane-coated curcumin polylactic-co-glycolic acid (PLGA) nanoparticles (PCNPs) are prepared to achieve significantly improved solubility, stability, and sustained release of curcumin. Fourier transform infrared spectra and X-ray diffraction assays indicate good encapsulation of curcumin within nanoparticles. Moreover, it is revealed for the first time that curcumin-loaded nanoparticles can not only suppress hemin-induced astrocyte proliferation but also induce astrocytes into neuron-like cells in vitro. PCNPs are used to treat rat ICH by tail vein injection, using in situ administration as control. The results show that PCNPs are more effective than curcumin-PLGA nanoparticles in concentrating on hemorrhagic lesions, inhibiting inflammation, suppressing astrogliosis, promoting neurogenesis, and improving motor functions. The treatment efficacy of intravenously administered PCNPs is comparable to that of in situ administration, indicating a good targeting effect of PCNPs on the hemorrhage site. This study provides a potent treatment for hemorrhagic injuries and a promising solution for efficient delivery of water-insoluble drugs using composite materials of macromolecules and cell membranes.

The experience of shared decision-making for people with asthma: A systematic review and metasynthesis of qualitative studies.

OBJECTIVES: To identify, describe and synthesise the views and experiences of adults living with asthma regarding shared decision-making (SDM) in the existing qualitative literature METHODS: We conducted a comprehensive search of 10 databases (list databases) from inception until September 2023. Screening was performed according to inclusion criteria. Tools from the Joanna Briggs lnstitute were utilised for the purposes of data extraction and synthesis in this study. The data extraction process in this study employed the Capability, Opportunity and Motivation Model of Behaviour (COM-B model) as a framework, and a pragmatic meta-aggregative approach was employed to synthesise the collected results. RESULTS: Nineteen studies were included in the metasynthesis. Three synthesised themes were identified: the capability of people living with asthma, the opportunities of people living with asthma in SDM, and the motivation of the people living with asthma in SDM. CONCLUSIONS: We have identified specific factors influencing people living with asthma engaging in SDM. The findings of this study can serve as a basis for the implementation of SDM in people living with asthma and provide insights for the development of their SDM training programs. The ConQual score for the synthesised findings was rated as low. To enhance confidence, future studies should address dependability and credibility factors. PRACTICE IMPLICATIONS: This review contemplates the implementation of SDM from the perspective of people living with asthma, with the aim of providing patient-centred services for them. The results of this review can benefit the implementation of SDM and facilitate information sharing. It offers guidance for SDM skills training among adults living with asthma, fosters a better doctor-patient relationship and facilitates consensus in treatment decisions, thereby enabling personalised and tailored medical care. PATIENT OR PUBLIC CONTRIBUTION: Three nursing graduate students participated in the data extraction and integration process, with two students having extensive clinical experience that provided valuable insights for the integration.

Chondroitin Sulfate/Dermatan Sulfate Hybrid Chains from Swim Bladder: Isolation, Structural Analysis, and Anticoagulant Activity.

Glycosaminoglycans (GAGs) with unique structures from marine animals show intriguing pharmacological activities and negligible biological risks, providing more options for us to explore safer agents. The swim bladder is a tonic food and folk medicine, and its GAGs show good anticoagulant activity. In this study, two GAGs, CMG-1.0 and GMG-1.0, were extracted and isolated from the swim bladder of Cynoscion microlepidotus and Gadus morhua. The physicochemical properties, precise structural characteristics, and anticoagulant activities of these GAGs were determined for the first time. The analysis results of the CMG-1.0 and GMG-1.0 showed that they were chondroitin sulfate (CS)/dermatan sulfate (DS) hybrid chains with molecular weights of 109.3 kDa and 123.1 kDa, respectively. They were mainly composed of the repeating disaccharide unit of -{IdoA-α1,3-GalNAc4S-ß1,4-}- (DS-A). The DS-B disaccharide unit of -{IdoA2S-α1,3-GalNAc4S-ß1,4-}- also existed in both CMG-1.0 and GMG-1.0. CMG-1.0 had a higher proportion of CS-O disaccharide unit -{-GlcA-ß1,3-GalNAc-ß1,4-}- but a lower proportion of CS-E disaccharide unit -{-GlcA-ß1,3-GalNAc4S6S-ß1,4-}- than GMG-1.0. The disaccharide compositions of the GAGs varied in a species-specific manner. Anticoagulant activity assay revealed that both CMG-1.0 and GMG-1.0 had potent anticoagulant activity, which can significantly prolong activated partial thromboplastin time. GMG-1.0 also can prolong the thrombin time. CMG-1.0 showed no intrinsic tenase inhibition activity, while GMG-1.0 can obviously inhibit intrinsic tenase with EC50 of 58 nM. Their significantly different anticoagulant activities may be due to their different disaccharide structural units and proportions. These findings suggested that swim bladder by-products of fish processing of these two marine organisms may be used as a source of anticoagulants.

Nurse managers' perceptions and experiences of caring behavior for clinical nurses: a multicenter survey.

BACKGROUND: Humanistic care management is a necessary measure to improve the motivation and initiative of clinical nurses and is the foundation to improve the quality of nursing. Understanding the current status and identifying the influencing factors that promote or hinder humanistic care behaviors is essential. This study investigated the current status and experiences of nurse managers' caring behaviors toward clinical nurses. METHODS: We conducted a mixed-methods study with an explanatory sequential design. A survey on the nurse managers' caring behaviors in 101 hospitals from 23 provinces and four municipalities in China was investigated (n = 2022). Then, semi-structured interviews were conducted to obtain information about the participants' experiences associated with the performance of caring behaviors (n = 27). RESULTS: Survey data demonstrated that the nurse managers' overall caring behaviors were moderately good. The total scoring rate was 88.55%, and the overall score was 161.19 ± 20.68. Qualitative data revealed that the capacity of nurse managers and clinical nurses, opportunity, and motivation to implement humanistic care are key influencing factors of caring behaviors. CONCLUSIONS: The results suggested that intrinsic motivation, organizational support, and the humanistic care capabilities of clinical nurses and nurse managers are vital to implementing care behaviors. Thus, successful humanistic care management requires a concerted effort at the individual and organizational levels.

Pseudomonas syringae activates ZAT18 to inhibit salicylic acid accumulation by repressing EDS1 transcription for bacterial infection.

Phytopathogens can manipulate plant hormone signaling to counteract immune responses; however, the underlying mechanism is mostly unclear. Here, we report that Pseudomonas syringae pv tomato (Pst) DC3000 induces expression of C2H2 zinc finger transcription factor ZAT18 in a jasmonic acid (JA)-signaling-dependent manner. Biochemical assays further confirmed that ZAT18 is a direct target of MYC2, which is a very important regulator in JA signaling. CRISPR/Cas9-generated zat18-cr mutants exhibited enhanced resistance to Pst DC3000, while overexpression of ZAT18 resulted in impaired disease resistance. Genetic characterization of ZAT18 mutants demonstrated that ZAT18 represses defense responses by inhibiting the accumulation of the key plant immune signaling molecule salicylic acid (SA), which is dependent on its EAR motif. ZAT18 exerted this inhibitory effect by directly repressing the transcription of Enhanced Disease Susceptibility 1 (EDS1), which is the key signaling component of pathogen-induced SA accumulation. Overexpression of ZAT18 resulted in decreased SA content, while loss of function of ZAT18 showed enhanced SA accumulation upon pathogen infection. Furthermore, enhanced resistance and SA content in zat18-cr mutants was abolished by the mutation in EDS1. Our data indicate that pathogens induce ZAT18 expression to repress the transcription of EDS1, further antagonising SA accumulation for bacterial infection.

DEVELOPMENT AND VALIDATION OF AN EXPLAINABLE ARTIFICIAL INTELLIGENCE FRAMEWORK FOR MACULAR DISEASE DIAGNOSIS BASED ON OPTICAL COHERENCE TOMOGRAPHY IMAGES.

PURPOSE: To develop and validate an artificial intelligence framework for identifying multiple retinal lesions at image level and performing an explainable macular disease diagnosis at eye level in optical coherence tomography images. METHODS: A total of 26,815 optical coherence tomography images were collected from 865 eyes, and 9 retinal lesions and 3 macular diseases were labeled by ophthalmologists, including diabetic macular edema and dry/wet age-related macular degeneration. We applied deep learning to classify retinal lesions at image level and random forests to achieve an explainable disease diagnosis at eye level. The performance of the integrated two-stage framework was evaluated and compared with human experts. RESULTS: On testing data set of 2,480 optical coherence tomography images from 80 eyes, the deep learning model achieved an average area under curve of 0.978 (95% confidence interval, 0.971-0.983) for lesion classification. In addition, random forests performed accurate disease diagnosis with a 0% error rate, which achieved the same accuracy as one of the human experts and was better than the other three experts. It also revealed that the detection of specific lesions in the center of macular region had more contribution to macular disease diagnosis. CONCLUSION: The integrated method achieved high accuracy and interpretability in retinal lesion classification and macular disease diagnosis in optical coherence tomography images and could have the potential to facilitate the clinical diagnosis.

Diosgenin alleviates hypercholesterolemia via SRB1/CES-1/CYP7A1/FXR pathway in high-fat diet-fed rats.

Phytosterol diosgenin (DG) exhibits cholesterol-lowering properties. Few studies focused on the underlying mechanism of DG attenuation of hypercholesterolemia by promoting cholesterol metabolism. To investigate the roles of SRB1/CES-1/CYP7A1/FXR pathways in accelerating cholesterol elimination and alleviating hypercholesterolemia, a rat model of hypercholesterolemia was induced by providing a high-fat diet (HFD). Experimental rat models were randomly divided into a normal control (Con) group, HFD group, low-dose DG (LDG) group (150 mg/kg/d), high-dose DG (HDG) group (300 mg/kg) and Simvastatin (Sim) group (4 mg/kg/d). Body weights, serum and hepatic lipid parameters of rats were tested. The expression levels of scavenger receptor class B type I (SRB1), carboxylesterase-1 (CES-1), cholesterol7α- hydroxylase (CYP7A1), and farnesoid X receptor (FXR) were determined. The results showed that DG reduced weight and lowered lipid levels in HFD-fed rats. Pathological morphology analyses revealed that DG notably improved hepatic steatosis and intestinal structure. Further studies showed the increased hepatic SRB1, CES-1, CYP7A1 and inhibited FXR-mediated signaling in DG-fed rats, which contributing to the decrease of hepatic cholesterol. DG also increased intestinal SRB1 and CES-1, inhibiting cholesterol absorption and promoting RCT. The expression levels of these receptors in the HDG group were higher than LDG and Sim groups. These data suggested that DG accelerated reverse cholesterol transport (RCT) and enhanced cholesterol elimination via SRB1/CES-1/CYP7A1/FXR pathway, and DG might be a new candidate for the alleviation of hypercholesterolemia.

Nursing home staff perceptions of challenges and coping strategies during COVID-19 pandemic in China.

BACKGROUND: First-line nursing staff are responsible for protecting residents, the most vulnerable population, from COVID-19 infections. They are at a high risk of being infected with COVID-19 and experience high levels of psychological distress. AIMS: To explore the challenges and coping strategies perceived by nursing staff during the COVID-19 pandemic in China. METHODS: In April,2020, we conducted a qualitative study using in-depth semi-structured interviews with nursing staff. Participants were selected from seven nursing homes in three cities in Hunan Province, China. RESULTS: A total of 21 nursing staff participated in the study, including seven nurse managers, seven registered nurses and seven nursing assistants. Three main themes were identified. Different groups encountered different sources of stress and adopted various coping strategies to fulfil their responsibilities. CONCLUSIONS: Nursing home staff were underprepared for dealing with COVID-19-related challenges. Educational programs to improve the ability to deal with COVID-19 prevention and control are needed.

Evaluation of quality improvement intervention with nurse training in nursing homes: A systematic review.

INTRODUCTION: Quality improvement may be a promising approach to improve the quality of care in nursing homes, and nurse training is a key step in a successful quality improvement practice. The implementation of training measures may be related to the quality of quality improvement practice. Little is known about the quality of quality improvement practice or effective nurse training measures that affect the quality of quality improvement interventions in nursing homes. AIMS: The aim of this review was to assess the quality of available quality improvement intervention designs and present effective nurse training measures that contribute to a high-quality quality improvement intervention. METHODS: We searched the Cumulative Index to Nursing and Allied Health Literature (CINAHL), Medline, Embase and the Cochrane Library for articles published before March 2019. quality improvement intervention quality was evaluated using a standardised assessment tool. Descriptive synthesis was used for the analysis. The Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) Checklist was used for transparency. RESULTS: We included 12 articles, 1 was rated as perfect quality and 7 good quality. Out of these 8 studies, 3 features primarily reflected differences in quality: compliance, sustainability and replication ability of the interventions. They were affected by measures included provision of advanced training, available training resources, feedback process, building quality improvement teams, setting up mentors and nursing leadership training. Other recommended measures included external cooperation and leadership empowerment. CONCLUSION: A high-quality quality improvement intervention should consider how to improve compliance, sustainability and replication ability. Adapting measures that are compatible with nurse training may ensure a successful implementation of quality improvement programmes that are conducive to the effective improvement of service quality. RELEVANCE TO CLINICAL PRACTICE: Quality improvement programmes should take into account measures that are compatible with nursing staff training. These measures should help improve the quality of interventions and promote care service of nursing homes.

Nursing home staff experiences of implementing mentorship programmes: A systematic review and qualitative meta-synthesis.

AIM: To determine nursing home staff experiences in mentorship programmes, and staff perceptions of the enablers and barriers to implement mentorship programmes. BACKGROUND: Mentorship programmes are perceived as playing an important role in improving the quality of care in nursing homes. However, little is known about research evidence across the global about staff's experiences in the programmes. METHODS: A search for studies published from the earliest available date to April 2019 was undertaken. Two reviewers performed data extraction and an appraisal of eight studies using tools from the Joanna Briggs Institute. A pragmatic meta-aggregative approach was applied to synthesise the findings. The qualitative research that was included was analysed to identify 63 findings that were organised into 12 categories and combined into three syntheses. RESULTS: The implementation of effective mentorship programmes is influenced by three factors: mentor capability, opportunity in the mentorship programmes, and motivation in the mentorship programmes. CONCLUSIONS: There are a number of studies of nursing home staff experiences of mentorship programmes. However, systematic reviews that synthesise findings in this field are lacking. It is crucial to tailor the programme design to suit each unique nursing home care setting. We identified barriers and enablers, and learned that no barriers are insurmountable. IMPLICATIONS FOR NURSING MANAGEMENT: Findings will inform nurse managers of an ideal environment for the implementation of a successful mentorship programme. Nursing homes need to establish and sustain mentorship programmes to help improve workforce capacity in delivering high-quality care for residents.

[Research progress on anti-atherosclerosis effect and mechanism of flavonoids compounds mediated by macrophages].

Flavonoids are important active ingredients of traditional Chinese medicine, mainly with cardiovascular, anti-liver injury, antioxidant, antispasmodic, and estrogen-like effects. These compounds have obvious effects on the cardiovascular and cerebrovascular diseases. Macrophage-derived foam cells are the key medium in the process of atherosclerosis(AS). In plaque, allserum lipids, serum lipoproteins, and various pro-or anti-inflammatory stimulating factors, chemokines, and small bioactive molecules can significantly affect the macrophage phenotype and induce stronger pro-inflammatory or anti-inflammatory properties. Studies have shown that some flavonoids can be used for macrophages through different pathways and mechanisms, playing an anti-atherosclerosis effect to different degrees, including promotion of cholesterol efflux from macrophages, anti-foaming of macrophages, inhibition of secretion of inflammatory factors, and antioxidant modified low density lipoprotein(ox-LDL)-induced apoptosis of macrophages. Related gene regulation inclu-ded ATP-binding cassette transporter A1(ABCA1), ATP-binding cassette transporter G1(ABCG1), Toll-like receptor(TLR), and scavenger receptor(SR). In this article, we would review the recent research progress of flavonoids on anti-atherosclerosis effect me-diated by macrophage. It is expected to provide new treatment strategies for AS-related cardiovascular and cerebrovascular diseases, and provide research ideas and development directions for the use of related natural medicines and design of new products.

Estrogen receptor alpha activates MAPK signaling pathway to promote the development of endometrial cancer.

Endometrial cancer (EC) is a common malignant tumor of the female reproductive system in the world. For most of the treated patients, although the survival rate is improved, most patients still have a poor prognosis. The pathogenesis of EC has always been a strong scientific focus, but there is no clear conclusion. Therefore, in view of modularization, this study is to conduct an in-depth analysis on the effects of estrogen receptor alpha (ERα) regarding EC. The purpose is to identify the molecular course of EC. We obtained 10 co-expression modules, in which ANO2, EMP3, and other genes are significantly differentially expressed in patients with EC. Additionally, there are active regulatory effects in dysfunction modules, thus genes such as ANO2 and EMP3 would be identified as key genes, which are associated with the development of EC. Enrichment results showed that the module genes were significantly involved in RNA splicing, covalent chromatin modification, histone modification, and organelle fission, and other biological processes, as well as significantly regulated mitogen-activated protein kinases (MAPK) signaling pathway, Endocytosis, Rap1 signaling pathway, and viral carcinogenesis, and other signaling pathways. Finally, we identified noncoding RNA pivot including FENDRR, miR-520c-3p. Besides, transcription factors pivot including NFKB1, E2F1, and RELA which significantly regulate dysfunction module genes. Overall, our work deciphered a co-expression network involving differential gene regulation in ERα-associated EC. It helps reveal the core modules and potential regulatory factors of the diseases and enhances our understanding of the pathogenesis. More importantly, we revealed that ERα activates the MAPK signaling pathway to promote the development of EC. It helps to provide a new reference for later research.

Verticillium dahliae PevD1, an Alt a 1-like protein, targets cotton PR5-like protein and promotes fungal infection.

Alt a 1 family proteins (AA1s) have only been observed in the Dothideomycetes and Sordariomycetes classes of fungi, and their biological functions have remained poorly understood. Verticillium dahliae, a soil-borne pathogen that causes plant wilt disease, secretes hundreds of proteins during the process of pathogenic infection, including the AA1 member PevD1. In this study, we found that the pevd1 transcript was present in all of the hosts studied (cotton, Arabidopsis, tomato, and tobacco) and showed elevated expression throughout the infection process. Furthermore, pevd1 knockout mutants displayed attenuated pathogenicity compared with the wild-type (WT) strain and complemented strains in hosts. A partner protein of PevD1, pathogenesis-related protein 5 (PR5)-like protein GhPR5, was isolated from cotton (Gossypium hirsutum) plants by co-purification assays, and the PevD1-GhPR5 interaction was determined to be localized in the C-terminus (PevD1b, amino acids residues 113-155) by pull-down and yeast two-hybrid techniques. Re-introduction of the pevd1b gene into a pevd1 knockout mutant resulted in restoration of the virulence phenotype to WT levels. In addition, PevD1b, which is similar to PevD1, decreased the antifungal activity of GhPR5 in vitro. Our findings reveal an infection strategy in which V. dahliae secretes PevD1 to inhibit GhPR5 antifungal activity in order to overcome the host defence system.

Secreted protein MoHrip2 is required for full virulence of Magnaporthe oryzae and modulation of rice immunity.

MoHrip2, identified from Magnaporthe oryzae as an elicitor, can activate plant defense responses either in the form of recombinant protein in vitro or ectopic expressed protein in rice. However, its intrinsic function in the infective interaction of M. oryzae-rice is largely unknown. Here, we found that mohrip2 expression was significantly induced at stages of fungal penetration and colonization. Meanwhile, the induced MoHrip2 mainly accumulated in the rice apoplast by outlining the entire invasive hyphae during infection, and its secretion was via the conventional endoplasmic reticulum (ER)-to-Golgi pathway, demonstrating the nature of MoHrip2 as an apoplastic effector. What's more, the disease facilitating function of MoHrip2 was revealed by the significantly compromised virulence of Δmohrip2 mutants on rice seedlings and even on the wounded rice leaves. Inoculations of these mutant strains on rice leaf sheaths showed a reduction in penetration and subsequent expansion of fungal growth, which is probably due to activated host immunity including the expression of certain defense-related genes and the production of certain phytoalexins. Altogether, these results demonstrated the necessity of MoHrip2 in suppression of host immunity and the full virulence of M. oryzae.

The Secreted Protein MoHrip1 Is Necessary for the Virulence of Magnaporthe oryzae.

Secreted effectors from Magnaporthe oryzae play critical roles in the interaction with rice to facilitate fungal infection and disease development. M. oryzae-secreted protein MoHrip1 can improve plant defense as an elicitor in vitro, however, its biological function in fungal infection is not clear. In this study, we found that the expression of mohrip1 was significantly induced in the stages of fungal penetration and colonization. Although dispensable for the growth and conidiation, MoHrip1 was necessary for the full virulence of M. oryzae. Deletion of mohrip1 remarkably compromised fungal virulence on rice seedlings and even on rice leaves with wounds. Rice sheath inoculation assay further demonstrated the defects of mohrip1-deleted mutants on penetration and proliferation in rice cells. Additionally, compared with WT and complementation strain, the inoculation of mohrip1-deleted mutants induced a higher expression of specific defense related genes and a higher production of specific defensive compounds in rice leaves. These data collectively indicated that MoHrip1 is necessary for fungal penetration and invasive expansion, and further full virulence of rice blast fungus.

The Novel Cerato-Platanin-Like Protein FocCP1 from Fusarium oxysporum Triggers an Immune Response in Plants.

Panama disease, or Fusarium wilt, the most serious disease in banana cultivation, is caused by Fusarium oxysporum f. sp. cubense (FOC) and has led to great economic losses worldwide. One effective way to combat this disease is by enhancing host plant resistance. The cerato-platanin protein (CPP) family is a group of small secreted cysteine-rich proteins in filamentous fungi. CPPs as elicitors can trigger the immune system resulting in defense responses in plants. In this study, we characterized a novel cerato-platanin-like protein in the secretome of Fusarium oxysporum f. sp. cubense race 4 (FOC4), named FocCP1. In tobacco, the purified recombinant FocCP1 protein caused accumulation of reactive oxygen species (ROS), formation of necrotic reaction, deposition of callose, expression of defense-related genes, and accumulation of salicylic acid (SA) and jasmonic acid (JA) in tobacco. These results indicated that FocCP1 triggered a hypersensitive response (HR) and systemic acquired resistance (SAR) in tobacco. Furthermore, FocCP1 enhanced resistance tobacco mosaic virus (TMV) disease and Pseudomonas syringae pv. tabaci 6605 (Pst. 6605) infection in tobacco and improved banana seedling resistance to FOC4. All results provide the possibility of further research on immune mechanisms of plant and pathogen interactions, and lay a foundation for a new biological strategy of banana wilt control in the future.

Lignin metabolism involves Botrytis cinerea BcGs1- induced defense response in tomato.

BACKGROUND: BcGs1, a cell wall-degrading enzyme (CWDE), was originally derived from Botrytis cinerea. Our previous study revealed that BcGs1 could trigger defense responses and protect plants against various pathogens. We researched the defense response mechanism underlying this BcGs1 elicitation in tomato. RESULTS: We revealed that the two domains were required for BcGs1's full necrosis activity. According to analysis and quantitative real-time PCR of the up-regulated proteins and genes filtered by iTRAQ-based quantitative proteome approach, oxidative metabolism and phenylpropanoid metabolism were speculated to be involved in BcGs1-triggered defense response in tomato. Furthermore, experimental evidence showed that BcGs1 triggered reactive oxygen species (ROS) burst and increased the level of phenylalanine-ammonia lyase (PAL) and peroxidase (POD) enzyme activity, as well as lignin accumulation. Moreover, histochemical analysis revealed that infiltration of BcGs1 in tomato leaves exhibited cell wall thickening compared with untreated plants. CONCLUSIONS: The results suggested that BcGs1 activated the basal defense response included lignin metabolism contributed to BcGs1-induced resistance to Botrytis. cinerea infection in tomato.

A DREPP protein interacted with PeaT1 from Alternaria tenuissima and is involved in elicitor-induced disease resistance in Nicotiana plants.

PeaT1 is a proteinaceous elicitor from fungal pathogen Alternaria tenuissima. Our previous research revealed that this elicitor could induce defense response and enhance disease resistance in various plants including Nicotiana plants. However, immune activation mechanisms whereby PeaT1 elicits defense response remain unclear. In this study, the association between elicitor protein PeaT1 and the plasma membrane was assessed using the FITC (Fluorescein isothiocyanate) labeling method. A PeaT1-interacting protein was isolated via 125I-PeaT1 cross-linking and Far Western blot analyses, and designated PtBP1 (PeaT1 Binding Protein 1). From the data of Mass spectrometry (MS) and bioinformatics analysis, the 22 kDa plasma membrane protein PtBP1 was inferred to be a member of DREPP (developmentally regulated plasma membrane polypeptide) family that is induced in plants under stress conditions and might get involved in downstream signaling. For further verification of this association, Far Western blot, co-immunoprecipitation and bimolecular fluorescence complementation (BiFC) analyses were performed, showing PtBP1 could bind with PeaT1 in vitro and in vivo. Virus-induced gene silencing (VIGS) analysis exhibited that PtBP1 silencing in Nicotiana benthamiana attenuated tobacco mosaic virus (TMV) resistance compared to the tobacco rattle virus (TRV) control after PeaT1 treatment.

The Magnaporthe oryzae Alt A 1-like protein MoHrip1 binds to the plant plasma membrane.

MoHrip1, a protein isolated from Magnaporthe oryzae, belongs to the Alt A 1 (AA1) family. mohrip1 mRNA levels showed inducible expression throughout the infection process in rice. To determine the location of MoHrip1 in M. oryzae, a mohrip1-gfp mutant was generated. Fluorescence microscopy observations and western blotting analysis showed that MoHrip1 was both present in the secretome and abundant in the fungal cell wall. To obtain MoHrip1 protein, we carried out high-yield expression of MoHrip1 in Pichia pastoris. Treatment of tobacco plants with MoHrip1 induced the formation of necrosis, accumulation of reactive oxygen species and expression of several defense-related genes, as well as conferred disease resistance. By fusion to green fluorescent protein, we showed that MoHrip1 was able to bind to the tobacco and rice plant plasma membrane, causing rapid morphological changes at the cellular level, such as cell shrinkage and chloroplast disorganization. These findings indicate that MoHrip1 is a microbe-associated molecular pattern that is perceived by the plant immune system. This is the first study on an AA1 family protein that can bind to the plant plasma membrane.

Comparison of cerato-platanin family protein BcSpl1 produced in Pichia pastoris and Escherichia coli.

The Botrytis cinerea BcSpl1 protein is a member of the cerato-platanin family, and consists of 137 amino acids and two disulfide bridges. This protein induces the onset of necrosis in infiltrated plant hosts. Recombinant BcSpl1 proteins produced in Pichia pastoris (pBcSpl1) and Escherichia coli (eBcSpl1) were initially compared regarding their abilities to induce necrosis and systemic acquired resistance (SAR). The pBcSpl1 and eBcSpl1 treatments led to the development of necrotic lesions on tomato leaves, and provided tomato plants with SAR to B. cinerea. The lesion area of leaves infiltrated with the BcSpl1 proteins decreased by 22.7% (pBcSpl1) and 21.8% (eBcSpl1). Additionally, eBcSpl1 up-regulated the expression levels of some defense-related genes, including PR-1a, prosystemin, PI I, and PI II, as well as SIPK and TPK1b, which encode two protein kinases. Furthermore, eBcSpl1 exhibited chitin-binding properties. Our data revealed that the E. coli expression system produces higher BcSpl1 yields than the P. pastoris system. This high-yield expression of BcSpl1 may be relevant for future large-scale applications of this elicitor to improve crop production.