Regulatory effects of Trichinella spiralis serpin-type serine protease inhibitor on endoplasmic reticulum stress and oxidative stress in host intestinal epithelial cells.

Endoplasmic reticulum stress (ERS) and oxidative stress (OS) are adaptive responses of the body to stressor stimulation. Although it has been verified that Trichinella spiralis (T. spiralis) can induce ERS and OS in the host, their association is still unclear. Therefore, this study explored whether T. spiralis-secreted serpin-type serine protease inhibitor (TsAdSPI) is involved in regulating the relationship between ERS and OS in the host intestine. In this study, mice jejunum and porcine small intestinal epithelial cells (IECs) were detected using qPCR, western blotting, immunohistochemistry (IHC), immunofluorescence (IF), and detection kits. The results showed that ERS- and OS-related indexes changed significantly after TsAdSPI stimulation, and Bip was located in IECs, indicating that TsAdSPI could induce ERS and OS in IECs. After the use of an ERS inhibitor, OS-related indexes were inhibited, suggesting that TsAdSPI-induced OS depends on ERS. When the three ERS signalling pathways, ATF6, IRE1, and PERK, were sequentially suppressed, OS was only regulated by the PERK pathway, and the PERK-eif2α-CHOP-ERO1α axis played a key role. Similarly, the expression of ERS-related indexes and the level of intracellular Ca2+ were inhibited after adding the OS inhibitor, and the expression of ERS-related indexes decreased significantly after inhibiting calcium transfer. This finding indicated that TsAdSPI-induced OS could affect ERS by promoting Ca2+ efflux from the endoplasmic reticulum. The detection of the ERS and OS sequences revealed that OS occurred before ERS. Finally, changes in apoptosis-related indexes were detected, and the results indicated that TsAdSPI-induced ERS and OS could regulate IEC apoptosis. In conclusion, TsAdSPI induced OS after entering IECs, OS promoted ERS by enhancing Ca2+ efflux, and ERS subsequently strengthened OS by activating the PERK-eif2α-CHOP-ERO1α axis. ERS and OS induced by TsAdSPI synergistically promoted IEC apoptosis. This study provides a foundation for exploring the invasion mechanism of T. spiralis and the pathogenesis of host intestinal dysfunction after invasion.

Effectiveness of Continuous Care Interventions in Elderly Patients with High-Risk Pressure Ulcers and Impact on Patients' Activities of Daily Living.

Objective: To explore the influence of continuous care interventions on the incidence of pressure ulcers in elderly patients with high-risk pressure ulcers and their activities of daily living (ADL). Methods: The clinical records of 114 elderly patients at high risk of pressure ulcers (Patients had a Braden pressure ulcer risk score less than or equal to 18 and suffered no pressure pressure ulcers) who attended our hospital from February 2020 to February 2022 were collected for retrospective analysis. Among them, 54 patients treated with conventional nursing interventions were assigned to the control group (CG) (64-91 years old), and the remaining 60 treated with continuous care interventions were assigned to the observation group (OG) (61-91years old). The detection indices and clinical related data were collected from LIS system of our hospital. The self-rating depression scale (SDS) and self-rating anxiety scale (SAS) scores of the two groups were compared before and after the intervention, and the improvement in quality of life and ADL scores were counted. The post-care satisfaction and incidence of pressure ulcers were analyzed, and the caregivers' pressure ulcer care behaviors and pressure ulcer management abilities before and after treatment were compared between both groups of patients. Results: The SAS and SDS scores were lower in the OG (38.33±3.21 and 39.07±4.15) than in the CG (44.74±4.13 and 45.80±3.92) after the intervention (P < .05); quality of life and ADL were higher in the OG (44.07±7.15 and 58.33±6.21) than in the CG (36.20±6.92 and 45.84±6.12) after the intervention (P < .05). The incidence of pressure ulcers in the OG (18.33%) was lower than that in the CG (37.04%) after the intervention (P < .05); total nursing satisfaction in the OG (91.67%) was higher than that in the CG (74.07%) after the intervention (P < .05), and caregivers' pressure ulcer care behaviors and pressure ulcer management abilities of patients in the OG (36.42±2.41 and 44.07±7.15) were higher than those in the CG (29.23±2.50 and 36.20±6.92) after the intervention (P < .05). Conclusion: Continuous care interventions can be promoted for elderly patients with high-risk pressure ulcers outside the hospital, which can reduce the incidence of pressure ulcers, negative emotions, related complications and treatment costs, and improve the effective utilization of medical resources. In the future, continuous care guidelines and training plans can be developed for elderly patients with high-risk pressure ulcers, so as to improve the understanding and application ability of health care providers and nurses on intervention measures.

Chemical Profiling and Antifungal Activity of Ziziphora clinopodioides Leaf Essential Oil against the Pathogen Verticillium dahliae.

Essential oils (EOs) are often used as natural antifungal agents to control the growth of phytopathogenic fungi. The aim of this study was to determine the effect of Ziziphora clinopodioides leaf EO against Verticillium dahliae, a pathogenic fungus of cotton. Gas chromatography-mass spectrometry (GC/MS) analysis revealed the presence of 15 compounds of the total of extracted oil, which was consisted of 98.79 % monoterpenes and 0.61 % sesquiterpenes. The major constituents were pulegone (62.17 %), isomenthone (18.42 %), l-menthone (5.55 %) and piperitenone (3.99 %). The mycelial growth of Verticillium dahliae was completely inhibited at 0.24 µL/mL air under vapor phase condition. Considerable morphological variations were also observed in the fungal sclerotia at the contact phase at 3 µL/mL. This study demonstrated for the first time that Z. clinopodioides EO can effectively inhibit the growth of V. dahliae, implying that it has the potential to be explored as an antifungal agent against Verticillium Wilt of cotton.

Postmyomectomy uterocutaneous fistula: a case report and literature review.

BACKGROUND: Uterocutaneous fistula (UCF) is an uncommon clinical entity. The aetiology and optimal treatment strategy of UCF are still not clear. GnRHa (gonadotropin-releasing hormone agonist) treatment as an option for conservative hormone therapy for UCF has diverse clinical outcomes. The true status of hormone receptors in UCF lesions and its relationship with the therapeutic effect of GnRHa treatment have not yet been studied. CASE PRESENTATION AND REVIEW OF THE LITERATURE: A 39-year-old woman underwent an uneventful myomectomy during her initial visit to our centre. The patient returned to our centre with complaints of fluid oozing from her abdominal wound. Misdiagnosed with poor wound healing, the patient received a secondary closure. Consequent blood-stained drainage from the wound during menstruation and imaging tests confirmed the diagnosis of UCF. Surgical repair and excision of the fistula tract were performed. Histopathological analysis revealed fistulization and a negative status of hormone receptors in the fistula. Considering the controversial role of GnRHa treatment in UCF and the negative oestrogen receptor (ER) and progesterone receptor (PR) status in the current case, we further systematically reviewed the literature for GnRHa treatment for UCF, with the results showing that GnRHa manipulation as a conservative hormone therapeutic schedule is effective in 2 (50%) of the 4 patients in which it was attempted. CONCLUSION: UCF is a rare entity and tends to be misdiagnosed. GnRHa as a conservative therapy for UCF has different clinical outcomes. The current finding of a negative hormone receptor status in the fistula may provide new clues for further exploration of the mechanisms of the diverse therapeutic effects of GnRHa on UCF.

The role of ARHGAP9: clinical implication and potential function in acute myeloid leukemia.

BACKGROUND: Rho GTPase activating protein 9 (ARHGAP9) is expressed in various types of cancers and can inactivate Rho GTPases that mainly regulate cytoskeletal dynamics. However, the exact role of ARHGAP9 in acute myeloid leukemia (AML) has yet to be clarified. METHODS: We compared the transcriptional expression, prognosis, differentially expressed genes, functional enrichment, and hub genes in AML patients on the basis of the data published in the following databases: UALCAN, GEPIA, Gene Expression Omnibus, the Human Protein Atlas, Cancer Cell Line Encyclopedia, LinkedOmics, Metascape, and String. Data from the Cancer Genome Atlas database was used to evaluate the correlations between ARHGAP9 expression and various clinicopathological parameters, as well as the significantly different genes associated with ARHGAP9 expression. RESULTS: We found that ARHGAP9 expression was higher in the tissues and cell lines extracted from patients with AML than corresponding control tissues and other cancer types. ARHGAP9 overexpression was associated with decreased overall survival (OS) in AML. Compared with the ARHGAP9low group, the ARHGAP9high group, which received only chemotherapy, showed significantly worse OS and event-free survival (EFS); however, no significant difference was observed after treatment with autologous or allogeneic hematopoietic stem cell transplantation (auto/allo-HSCT). The ARHGAP9high patients undergoing auto/allo-HSCT also had a significantly better prognosis with respect to OS and EFS than those receiving only chemotherapy. Most overlapping genes of the significantly different genes and co-expression genes exhibited enriched immune functions, suggesting the immune regulation potential of ARHGAP9 in AML. A total of 32 hub genes were identified from the differentially expressed genes, within which the KIF20A had a significant prognostic value for AML. CONCLUSIONS: ARHGAP9 overexpression was associated with poor OS in AML patients and can be used as a prognostic biomarker. AML patients with ARHGAP9 overexpression can benefit from auto/allo-HSCT rather than chemotherapy.

RNAi-mediated silencing of Trichinella spiralis glutaminase results in reduced muscle larval infectivity.

Trichinella spiralis is an important foodborne parasitic nematode distributed worldwide that infects humans and animals. Glutaminase (GLS) is an important gene in the glutamine-dependent acid resistance (AR) system; however, its role in T. spiralis muscle larvae (ML) remains unclear. The present study aimed to characterize T. spiralis GLS (TsGLS) and assess its function in T. spiralis ML AR both in vitro and in vivo using RNA interference. The results indicated that native TsGLS (72 kDa) was recognized by anti-rTsGLS serum at the muscle larvae stage; moreover, an immunofluorescence assay confirmed that TsGLS was located in the epidermis of ML. After silencing the TsGLS gene, the relative expression of TsGLS mRNA and the survival rate of T. spiralis ML were reduced by 60.11% and 16.55%, respectively, compared to those in the PBS and control groups. In vivo AR assays revealed that the worm numbers at 7 and 35 days post-infection (dpi) decreased by 61.64% and 66.71%, respectively, compared to those in the PBS group. The relative expression of TsGLS mRNA in F1 generation T. spiralis ML was reduced by 42.52%, compared to that in the PBS group. To the best of our knowledge, this is the first study to report the presence of the glutamine-dependent AR system in T. spiralis. Our results indicate that TsGLS plays a crucial role in the T. spiralis AR system; thus, it could be used as a potential candidate target molecule for producing vaccines against T. spiralis infection.

Chemical composition and phytotoxicity of essential oil from invasive plant, Ambrosia artemisiifolia L.

Essential oils have been evaluated as appropriate phytotoxins with mechanisms of action that are different from those of synthetic herbicides applied in weed management activities, but little is known about the effect of Ambrosia artemisiifolia essential oil (EO) on weeds. Here, the chemical composition of A. artemisiifolia EO was analyzed using a Gas Chromatography-Mass Spectrometry system. and the phytotoxic activities of the EO against monocot (Poa annua, Setaria viridis) and dicot (Amaranthus retroflexus, Medicago sativa) species are evaluated under laboratory and green-house conditions for the first time. The EO was rich in sesquiterpenes (62.51%), with germacrene D (32.92%), ß-pinene (15.14%), limonene (9.90%), and caryophyllene (4.49%) being the major compounds based on Gas Chromatography-Mass Spectrometry analysis results. A. artemisiifolia EO inhibited seed germination and seedling development significantly in the tested species even at low concentrations (0.25 mg mL-1). In addition, bioassay results for the activities of superoxide dismutase (SOD) and peroxidase (POD) increased and then decreased with an increase in EO concentration. Unlike the enzymatic activity, root cell viability declined significantly in the tested weeds in all EO treatments. Besides, a foliar spray experiment resulted in visible injury in leaves and a decrease in chlorophyll content and eventually led to wilting of all tested weeds. The EO (0.25-5.00 mg mL-1) altered Allium cepa root tip cells with a decline in mitotic index and an increase in chromosomal aberrations after 24 h treatment. The cytotoxic evaluation confirmed the mitotic inhibitory effect of EO, although the intensity varied under different concentrations. According to the results, A. artemisiifolia EO has the potential applications as a natural herbicide owing to its phytotoxic activity; which also helps to explain their potential involvement in allelopathic interaction of volatile compounds present in the EO that facilitate the invasion success of the exotic species.

Chemical Profile and Phytotoxic Action of Hibiscus trionum Essential Oil.

The chemical profile and phytotoxic action of Hibiscus trionum essential oil (EO) was studied. In total 17 compounds were identified via GC/MS, representing 94.18 % of the entire oil, with phytol (40.37 %) being the dominant constituent. Bioassay revealed that the EO inhibited root elongation of Medicago sativa and Amaranthus retroflexus by 32.66 % and 61.86 % at 5 mg/mL, respectively; meanwhile, the major component phytol also exhibited significant phytotoxic activity, suppressing radical elongation of Pennisetum alopecuroides, M. sativa and A. retroflexus by 26.08 %, 27.55 % and 43.96 % at 1 mg/mL, respectively. The fact that the EO showed weaker activity than phytol implied that some constituents might trigger antagonistic action to decrease the oil's activity. Our study is the first on the chemical profile and phytotoxic effect of H. trionum EO.

Epigenetic silencing of miR564 contributes to the leukemogenesis of t(8;21) acute myeloid leukemia.

The AML1-ETO oncoprotein, which results from t(8;21) translocation, is considered an initial event of t(8;21) acute myeloid leukemia (AML). However, the precise mechanisms of the oncogenic activity of AML1-ETO is yet to be fully determined. The present study demonstrates that AML1-ETO triggers the heterochromatic silencing of microRNA-564 (miR564) by binding at the AML1 binding site along the miR564 promoter region and recruiting chromatin-remodeling enzymes. Suppression of miR564 enhances the oncogenic activity of the AML1-ETO oncoprotein by directly inhibiting the expression of CCND1 and the DNMT3A genes. Ectopic expression of miR564 can induce retardation of G1/S transition, reperform differentiation, promote apoptosis, as well as inhibit the proliferation and colony formation of AML1-ETO+ leukemia cells in vitro. Enhanced miR564 levels can significantly inhibit the tumor proliferation of t(8;21)AML in vivo. We first identify an unexpected and important epigenetic circuitry of AML1-ETO/miR564/CCND1/DNMT3A that contributes to the leukemogenesis in vitro/vivo of AML1-ETO+ leukemia, indicating that miR564 enhancement could provide a potential therapeutic method for AML1-ETO+ leukemia.

The regulatory mechanism of chilling-induced dormancy transition from endo-dormancy to non-dormancy in Polygonatum kingianum Coll.et Hemsl rhizome bud.

KEY MESSAGE: We identified three dormant stages of Polygonatum kingianum and changes that occurred during dormancy transition in the following aspects including cell wall and hormones, as well as interaction among them. Polygonatum kingianum Coll.et Hemsl (P. kingianum) is an important traditional Chinese medicine, but the mechanism of its rhizome bud dormancy has not yet been studied systematically. In this study, three dormancy phases were induced under controlled conditions, and changes occurring during the transition were examined, focusing on phytohormones and the cell wall. As revealed by HPLC-MS (High Performance Liquid Chromatography-Mass Spectrometry) analysis, the endo- to non-dormancy transition was association with a reduced abscisic acid (ABA)/gibberellin (GA3) ratio, a decreased level of auxin (IAA) and an increased level of trans-zeatin (tZR). Transmission electron microscopy showed that plasmodesmata (PDs) and the cell wall of the bud underwent significant changes between endo- and eco-dormancy. A total of 95,462 differentially expressed genes (DEGs) were identified based on transcriptomics, and clustering and principal component analysis confirmed the different physiological statuses of the three types of bud samples. Changes in the abundance of transcripts associated with IAA, cytokinins (CTKs), GA, ABA, brassinolide (BR), jasmonic acid (JA), ethylene, salicylic acid (SA), PDs and cell wall-loosening factors were analysed during the bud dormancy transition in P. kingianum. Furthermore, nitrilase 4 (NIT4) and tryptophan synthase alpha chain (TSA1), which are related to IAA synthesis, were identified as hub genes of the co-expression network, and strong interactions between hormones and cell wall-related factors were observed. This research will provide a good model for chilling-treated rhizome bud dormancy in P. kingianum and cultivation of this plant.

Retraction Note to: Integrated physiological and proteomic analysis of embryo and endosperm reveals central salt stress response proteins during seed germination of winter wheat cultivar Zhengmai 366.

The editor has retracted this article [1] because parts of Figs. 1 and 4 were duplicated from a previously published paper by the same authors [2] without appropriate disclosure. None of the authors have responded to any correspondence from the editor about publication of this retraction notice.

Integrated physiological and proteomic analysis of embryo and endosperm reveals central salt stress response proteins during seed germination of winter wheat cultivar Zhengmai 366.

BACKGROUND: Salinity is a major abiotic stressor that affects seed germination, plant growth, and crop production. Seed germination represents the beginning of plant growth and is closely linked with subsequent crop development and ultimate yield formation. This study attempted to extend findings regarding the potential proteomic dynamics during wheat seed germination under salt stress and to explore the mechanism of crop salt response. RESULTS: Salt stress significantly affected seed physiological activities during the germination process, resulting in significant decreases in phytohormone and α-amylase activity and significant increases in soluble sugar, starch, and ADP glucose pyrophosphorylase activity. A comparative proteomics approach was applied to analyze the dynamic proteome changes of embryo and endosperm during seed germination in Chinese winter wheat cultivar Zhengmai 366 under salt stress. Two-dimensional electrophoresis identified 92 and 61 differentially accumulated proteins (DAPs) in response to salt stress in embryo and endosperm, respectively. Both organs contained a high proportion of DAPs involved in stress defense, energy metabolism, and protein/amino acid metabolism. The endosperm had more DAPs related to storage proteins and starch metabolism than the embryo, and 2% of DAPs participating in lipid and sterol metabolism were specifically detected in the embryo. CONCLUSIONS: Seed physiological activities were significantly affected during the germination process when subjected to salt stress. The DAPs involved in stress defense and energy metabolism were upregulated whereas those related to reserve substance degradation and protein/amino acid metabolism were significantly downregulated, leading to delayed seed germination under salt stress. Our proteomic results revealed synergistic regulation of the response to salt stress during seed germination.

Immunomodulatory effects of Trichinella spiralis excretory-secretory antigens on macrophages.

Helminths and their products can suppress the host immune response and other immunopathologies which may benefit parasite survival. Parasitic nematode Trichinella spiralis (T. spiralis) exert immunomodulatory effect on the host immune response through excretory-secretory (ES) products. However, the immunomodulatory mechanism is not yet completely understood or defined. Macrophages play a key role in modulating the host immune response to helminth parasite infection. Here, we focus on the effect of T. spiralis ES antigens on the immune response by studying the effect of ES antigens on RAW264.7 macrophages in vitro. RAW264.7 macrophages were incubated with ES antigens either alone or in combination with LPS. The cytokine production (TNF-α, IL-10 and IL-12 p70) and the expression of TLR were measured. Moreover, the nuclear translocation of MyD88 and NF-кB was assessed by Western blot analysis. Results indicate that ES products had impacts on reducing the expression of TLRs in LPS-induced macrophages. In addition, ES products inhibited the cytokine production of IL-12 p70 and TNF-α and alone boosted the expression of cytokine IL-10 in RAW264.7 macrophages. In conclusion, our results implied that T. spiralis ES antigens may regulate host immune response at the macrophages level in vitro.

Chemical Composition and Allelopathic, Phytotoxic and Pesticidal Activities of Atriplex cana Ledeb. (Amaranthaceae) Essential Oil.

The chemical composition and allelopathic, phytotoxic and pesticidal activities of Atriplex cana Ledeb. (Amaranthaceae) essential oil were investigated. Nineteen compounds were identified via GC/MS, representing 82.3 % of the total oil, and the most abundant constituents were dibutyl phthalate (21.79 %), eucalyptol (20.14 %) and myrtenyl acetate (15.56 %). The results showed that volatile organic compounds (VOCs) released by A. cana significantly inhibited seedling growth of Amaranthus retroflexus L. and Poa annua L., and 80 g of fresh stems and leaves of A. cana in a 1.5 L airtight container almost completely suppressed the seed germination of both plants. Meanwhile, 5 µg/mL essential oil completely inhibited the seed germination of A. retroflexus, Medicago sativa L., P. annua and Echinochloa crusgalli L. Pesticidal testing revealed that the essential oil had strong behavioral avoidance and lethal effects on Aphis pomi DeGeer. Five microliters of essential oil/Petri dish treatment resulted in an 84.5 % mortality rate after 12 h, and the mortality rate reached nearly 100 % after 48 h. This report is the first one on the chemical composition as well as the biological activity of the essential oil of A. cana, and our results indicate that the oil is valuable in terms of being further exploited as a bioherbicide/insecticide.

Chemical Composition and Phytotoxic Activity of Seriphidium terrae-albae (Krasch.) Poljakov (Compositae) Essential Oil.

To evaluate the potential value of Seriphidium terrae-albae (Krasch.) Poljakov essential oil as bioherbicide, its chemical composition as well as phytotoxic activity was investigated. Seventeen compounds were identified via GC/MS, representing 98.1 % of the total oil, and the most abundant constituents were α-thujone (43.18 %), ß-thujone (16.92 %), eucalyptol (17.55 %), and camphor (13.88 %). Phytotoxic assay revealed that the essential oil as well as its major constituents exhibited inhibitory activity on root and shoot growth of receiver plants in a dose-dependent manner. When the concentration reached 20 µg/mL, root length of Amaranthus retroflexus was reduced to 31.3 %, 70.6 %, 36.9 %, and 66.6 % of the control, respectively, when treated with α-thujone, eucalyptol, camphor, and the mixture of these compounds; meanwhile, root length of Poa annua was 3.0 %, 24.2 %, 0 %, and 4.4 % of the control when the same chemicals were applied. On the other hand, the essential oil showed a much stronger activity. At 1.5 µL/mL, root and shoot length of A. retroflexus and P. annua were reduced to 0.65 %, 0.5 %, and 1.53 %, 1.51 % of the control, respectively, and seed germination of A. retroflexus and P. annua was completely inhibited when the oil concentration reached 3 µg/mL and 5 µg/mL, respectively. This is the first report on the chemical composition of the essential oil of S. terrae-albae, and our results indicated that it has the potential to be further exploited as a bioherbicide.

Dynamic metabolome profiling reveals significant metabolic changes during grain development of bread wheat (Triticum aestivum L.).

BACKGROUND: Metabolites in wheat grains greatly influence nutritional values. Wheat provides proteins, minerals, B-group vitamins and dietary fiber to humans. These metabolites are important to human health. However, the metabolome of the grain during the development of bread wheat has not been studied so far. In this work the first dynamic metabolome of the developing grain of the elite Chinese bread wheat cultivar Zhongmai 175 was analyzed, using non-targeted gas chromatography/mass spectrometry (GC/MS) for metabolite profiling. RESULTS: In total, 74 metabolites were identified over the grain developmental stages. Metabolite-metabolite correlation analysis revealed that the metabolism of amino acids, carbohydrates, organic acids, amines and lipids was interrelated. An integrated metabolic map revealed a distinct regulatory profile. The results provide information that can be used by metabolic engineers and molecular breeders to improve wheat grain quality. CONCLUSION: The present metabolome approach identified dynamic changes in metabolite levels, and correlations among such levels, in developing seeds. The comprehensive metabolic map may be useful when breeding programs seek to improve grain quality. The work highlights the utility of GC/MS-based metabolomics, in conjunction with univariate and multivariate data analysis, when it is sought to understand metabolic changes in developing seeds. © 2015 Society of Chemical Industry.

Rapid separation of seed gliadins by reversed-phase ultra performance liquid chromatography (RP-UPLC) and its application in wheat cultivar and germplasm identification.

To separate gliadin from wheat flour, a novel and stability-indicating reversed-phase ultra performance liquid chromatography (RP-UPLC) method is established and optimized. A comparative analysis of routine capillary electrophoresis (CE), reversed-phase high-performance liquid chromatography (RP-HPLC), and RP-UPLC was performed and the results showed that the resolution and efficiency of RP-UPLC were significantly higher than those of CE and RP-HPLC. Characteristic RP-UPLC patterns of different bread wheat variety and related species were readily identified. These results demonstrated that our RP-UPLC procedure resulted in significant improvements in sensitivity, speed, and resolution, and thus is highly useful in wheat cultivar and germplasm identification.

Deletion of the low-molecular-weight glutenin subunit allele Glu-A3a of wheat (Triticum aestivum L.) significantly reduces dough strength and breadmaking quality.

BACKGROUND: Low-molecular-weight glutenin subunits (LMW-GS), encoded by Glu-3 complex loci in hexaploid wheat, play important roles in the processing quality of wheat flour. To date, the molecular characteristics and effects on dough quality of individual Glu-3 alleles and their encoding proteins have been poorly studied. We used a Glu-A3 deletion line of the Chinese Spring (CS-n) wheat variety to conduct the first comprehensive study on the molecular characteristics and functional properties of the LMW-GS allele Glu-A3a. RESULTS: The Glu-A3a allele at the Glu-A3 locus in CS and its deletion in CS-n were identified and characterized by proteome and molecular marker methods. The deletion of Glu-A3a had no significant influence on plant morphological and yield traits, but significantly reduced the dough strength and breadmaking quality compared to CS. The complete sequence of the Glu-A3a allele was cloned and characterized, which was found to encode a B-subunit with longer repetitive domains and an increased number of α-helices. The Glu-A3a-encoded B-subunit showed a higher expression level and accumulation rate during grain development. These characteristics of the Glu-A3a allele could contribute to achieving superior gluten quality and demonstrate its potential application to wheat quality improvement. Furthermore, an allele-specific polymerase chain reaction (AS-PCR) marker for the Glu-A3a allele was developed and validated using different bread wheat cultivars, including near-isogenic lines (NILs) and recombinant inbred lines (RILs), which could be used as an effective molecular marker for gluten quality improvement through marker-assisted selection. CONCLUSIONS: This work demonstrated that the LMW-GS allele Glu-A3a encodes a specific LMW-i type B-subunit that significantly affects wheat dough strength and breadmaking quality. The Glu-A3a-encoded B-subunit has a long repetitive domain and more α-helix structures as well as a higher expression level and accumulation rate during grain development, which could facilitate the formation of wheat with a stronger dough structure and superior breadmaking quality.

Dynamic development of starch granules and the regulation of starch biosynthesis in Brachypodium distachyon: comparison with common wheat and Aegilops peregrina.

BACKGROUND: Thorough understanding of seed starch biosynthesis and accumulation mechanisms is of great importance for agriculture and crop improvement strategies. We conducted the first comprehensive study of the dynamic development of starch granules and the regulation of starch biosynthesis in Brachypodium distachyon and compared the findings with those reported for common wheat (Chinese Spring, CS) and Aegilops peregrina. RESULTS: Only B-granules were identified in Brachypodium Bd21, and the shape variation and development of starch granules were similar in the B-granules of CS and Bd21. Phylogenetic analysis showed that most of the Bd21 starch synthesis-related genes were more similar to those in wheat than in rice. Early expression of key genes in Bd21 starch biosynthesis mediate starch synthesis in the pericarp; intermediate-stage expression increases the number and size of starch granules. In contrast, these enzymes in CS and Ae. peregrina were mostly expressed at intermediate stages, driving production of new B-granules and increasing the granule size, respectively. Immunogold labeling showed that granule-bound starch synthase (GBSSI; related to amylose synthesis) was mainly present in starch granules: at lower levels in the B-granules of Bd21 than in CS. Furthermore, GBSSI was phosphorylated at threonine 183 and tyrosine 185 in the starch synthase catalytic domain in CS and Ae. peregrina, but neither site was phosphorylated in Bd21, suggesting GBSSI phosphorylation could improve amylose biosynthesis. CONCLUSIONS: Bd21 contains only B-granules, and the expression of key genes in the three studied genera is consistent with the dynamic development of starch granules. GBSSI is present in greater amounts in the B-granules of CS than in Bd21; two phosphorylation sites (Thr183 and Tyr185) were found in Triticum and Aegilops; these sites were not phosphorylated in Bd21. GBSSI phosphorylation may reflect its importance in amylose synthesis.