Regulation of carbohydrate metabolism during anther development in a thermo-sensitive genic male-sterile wheat line.

Bainong sterility (BNS) is a thermo-sensitive genic male sterile wheat line, characterised by anther fertility transformation in response to low temperature (LT) stress during meiosis, the failure of vacuole decomposition and the absence of starch accumulation in sterile bicellular pollen. Our study demonstrates that the late microspore (LM) stage marks the transition from the anther growth to anther maturation phase, characterised by the changes in anther structure, carbohydrate metabolism and the main transport pathway of sucrose (Suc). Fructan is a main storage polysaccharide in wheat anther, and its synthesis and remobilisation are crucial for anther development. Moreover, the process of pollen amylogenesis and the fate of the large vacuole in pollen are closely intertwined with fructan synthesis and remobilisation. LT disrupts the normal physiological metabolism of BNS anthers during meiosis, particularly affecting carbohydrate metabolism, thus determining the fate of male gametophytes and pollen abortion. Disruption of fructan synthesis and remobilisation regulation serves as a decisive event that results in anther abortion. Sterile pollen exhibits common traits of pollen starvation and impaired starch accumulation due to the inhibition of apoplastic transport starting from the LM stage, which is regulated by cell wall invertase TaIVR1 and Suc transporter TaSUT1.

Kir4.1 deletion prevents salt-sensitive hypertension in early streptozotocin-induced diabetic mice via Na + -Cl - cotransporter in the distal convoluted tubule.

OBJECTIVES: Functional impairment of renal sodium handling and blood pressure (BP) homeostasis is an early characteristic manifestation of type 1 diabetes. However, the underlying mechanisms remain unclear. METHODS: Metabolic cages, radio-telemetry, immunoblotting, and electrophysiology were utilized to examine effects of high salt (8% NaCl, HS) intake on Na + /K + balance, BP, Na + -Cl - cotransporter (NCC) function, and basolateral K + channel activity in the distal convoluted tubule (DCT) under diabetic conditions. RESULTS: Improper Na + balance, hypernatremia, and a mild but significant increase in BP were found in streptozotocin (STZ)-induced diabetic mice in response to HS intake for 7 days. Compared to the vehicle, STZ mice showed increased Kir4.1 expression and activity in the DCT, a more negative membrane potential, higher NCC abundance, and enhanced hydrochlorothiazide-induced natriuretic effect. However, HS had no significant effect on basolateral Kir4.1 expression/activity and DCT membrane potential, or NCC activity under diabetic conditions, despite a downregulation in phosphorylated NCC abundance. In contrast, HS significantly downregulated the expression of Na + -H + exchanger 3 (NHE3) and cleaved epithelial sodium channel-γ in STZ mice, despite an increase in NHE3 abundance after STZ treatment. Kir4.1 deletion largely abolished STZ-induced upregulation of NCC expression and prevented BP elevation during HS intake. Interestingly, HS causes severe hypokalemia in STZ-treated kidney-specific Kir4.1 knockout (Ks-Kir4.1 KO) mice and lead to death within a few days, which could be attributed to a higher circulating aldosterone level. CONCLUSIONS: We concluded that Kir4.1 is required for upregulating NCC activity and may be essential for developing salt-sensitive hypertension in early STZ-induced diabetes.

Medicinal Values and Potential Risks Evaluation of Ginkgo biloba Leaf Extract (GBE) Drinks Made from the Leaves in Autumn as Dietary Supplements.

Ginkgo tea and ginkgo wine are two familiar Ginkgo biloba leaf extract (GBE) drinks in the form of dietary supplements (DS) used for healthcare in east Asia. Nevertheless, a comprehensive evaluation of their safety and efficacy is still lacking. In this study, GBE drinks were prepared from naturally newly senescent yellow leaves (YL) and green leaves (GL) in autumn. Their total flavonoids, antioxidant capacity and prescribed ingredients were investigated. In brief, the proportions of total flavonoids, total flavonol glycosides (TFs), total terpene trilactones (TTLs) and ginkgolic acids in the GBE drinks all did not meet the standards of worldwide pharmacopoeias. Specifically, the levels of TFs in the ginkgo tea prepared from YL were significantly higher than that prepared from GL. Further analyses revealed a substandard ratio of isorhamnetin/quercetin and an accumulation of leaf-age-related compounds, which were both unqualified. The proportions of specific TTLs varied between the ginkgo tea and ginkgo wine, although no significant differences were detected in terms of the total levels of TTLs. Noticeably, numerous biflavones and thousands of times over the limiting concentration of ginkgolic acids, including newly identified types, were only detected in ginkgo wine. Finally, the use of the GBE drinks as DSs was comprehensively evaluated according to the acceptable daily intake. This study showed the limited healthcare effects of GBE drinks despite their powerful antioxidant capacity.

Comparison of oxidative stress-mitochondria-mediated tenderization in two different bovine muscles during aging.

The aim of this study was to investigate the differences in the effects of mitochondria-involved energy metabolism and caspases activation on postmortem tenderness in different muscle fiber types. Beef Longissimus thoracis (LT) and Psoas major (PM) muscles showed significant difference in mitochondrial function. Our data revealed that PM suffered from higher levels of reactive oxygen species (ROS) earlier than LT, causing faster mitochondrial swelling and rupture. Additionally, faster metabolism of ATP-related compounds and activation of caspase-9 appeared in PM, but the activity of caspase-3 in PM was lower than that in LT. Differences in myofibril fragmentation index (MFI) of LT and PM at different aging stages suggested that energy metabolism and caspases activities may play a role in tenderness at different aging stages. These results indicated that oxidative stress-mitochondria-mediated tenderization process could be muscle-specific.

Identification of ankyrin-transmembrane-type subfamily genes in Triticeae species reveals TaANKTM2A-5 regulates powdery mildew resistance in wheat.

The ankyrin-transmembrane (ANKTM) subfamily is the most abundant subgroup of the ANK superfamily, with critical roles in pathogen defense. However, the function of ANKTM proteins in wheat immunity remains largely unexplored. Here, a total of 381 ANKTMs were identified from five Triticeae species and Arabidopsis, constituting five classes. Among them, class a only contains proteins from Triticeae species and the number of ANKTM in class a of wheat is significantly larger than expected, even after consideration of the ploidy level. Tandem duplication analysis of ANKTM indicates that Triticum urartu, Triticum dicoccoides and wheat all had experienced tandem duplication events which in wheat-produced ANKTM genes all clustered in class a. The above suggests that not only did the genome polyploidization result in the increase of ANKTM gene number, but that tandem duplication is also a mechanism for the expansion of this subfamily. Micro-collinearity analysis of Triticeae ANKTMs indicates that some ANKTM type genes evolved into other types of ANKs in the evolution process. Public RNA-seq data showed that most of the genes in class d and class e are expressed, and some of them show differential responses to biotic stresses. Furthermore, qRT-PCR results showed that some ANKTMs in class d and class e responded to powdery mildew. Silencing of TaANKTM2A-5 by barley stripe mosaic virus-induced gene silencing compromised powdery mildew resistance in common wheat Bainongaikang58. Findings in this study not only help to understand the evolutionary process of ANKTM genes, but also form the basis for exploring disease resistance genes in the ANKTM gene family.

The Deterioration of Agronomical Traits of the Continuous Cropping of Stevia Is Associated With the Dynamics of Soil Bacterial Community.

Stevia rebaudiana Bertoni is grown worldwide as an important, natural sweetener resource plant. The yield of steviol glycosides (SVglys) is greatly influenced by continuous cropping. In this study, we collected the roots, rhizosphere soils, and bulk soils from 2 years of continuous cropping (Y2) and 8 years of continuous cropping (Y8). A high-throughput sequencing technology based on Illumina Hiseq 2500 platform was used to study the structure and diversity of bacterial communities in the roots and soils of stevia with different years of continuous cropping. The results demonstrated that although the content of a group of SVglys was significantly increased in stevia of long-term continuous cropping, it inhibited the growth of plants and lowered the leaf dry weight; as a result, the total amount of SVglys was significantly decreased. Meanwhile, continuous cropping changed the physicochemical properties and the bacterial composition communities of soil. The different sampling sources of the root, rhizosphere soil, and bulk soil had no impact on the richness of bacterial communities, while it exhibited obvious effects on the diversity of bacterial communities. Continuous cropping had a stronger effect on the bacterial community composition in rhizosphere soil than in root and bulk soil. Based on linear discriminant analysis effect size (LEfSe), in the rhizosphere soil of Y8, the relative abundance of some beneficial bacterial genera of Sphingomonas, Devosia, Streptomyces, and Flavobacterium decreased significantly, while the relative abundance of Polycyclovorans, Haliangium, and Nitrospira greatly increased. Moreover, the soil pH and nutrient content, especially the soil organic matter, were correlated with the relative abundance of predominant bacteria at the genus level. This study provides a theoretical basis for uncovering the mechanism of obstacles in continuous stevia cropping and provides guidance for the sustainable development of stevia.

Genome-Wide Analysis of Serine Hydroxymethyltransferase Genes in Triticeae Species Reveals That TaSHMT3A-1 Regulates Fusarium Head Blight Resistance in Wheat.

Serine hydroxymethyltransferase (SHMT) plays a pivotal role in cellular one-carbon, photorespiration pathways and it influences the resistance to biotic and abiotic stresses. However, the function of SHMT proteins in wheat remains largely unexplored. In the present study, SHMT genes in five Triticeae species, Oryza sativa, and four dicotyledon species were identified based on whole genome information. The origin history of the target gene was traced by micro-collinearity analysis. Gene expression patterns of TaSHMTs in different tissues, various biotic stresses, exogenous hormones, and two biotic stresses were determined by Quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR). The function of the selected TaSHMT3A-1 was studied by barley stripe mosaic virus-induced gene silencing in common wheat Bainong207. A total of 64 SHMT members were identified and further classified into two main classes based on the structure of SHMT proteins. The gene structure and motif composition analyses revealed that SHMTs kept relatively conserved within the same subclasses. Interestingly, there was a gene, TdSHMT7B-1, on chromosome 7B of Triticum dicoccoides, but there was no SHMT gene on chromosome 7 of other analyzed Triticeae species; TdSHMT7B-1 had fewer exons and conserved motifs than the genes in the same subclass, suggesting that the gene of TdSHMT7B-1 has a notable evolutionary progress. The micro-collinearity relationship showed that no homologs of TaSHMT3A-1 and its two neighboring genes were found in the collinearity region of Triticum urartu, and there were 27 genes inserted into the collinearity region of T. urartu. Furthermore, qRT-PCR results showed that TaSHMT3A-1 was responsive to abiotic stresses (NaCl and cold), abscisic acid, methyl jasmonate, and hydrogen peroxide. Significantly, upon Fusarium graminearum infection, the expression of TaSHMT3A-1 was highly upregulated in resistant cultivar Sumai3. More importantly, silencing of TaSHMT3A-1 compromises Fusarium head blight resistance in common wheat Bainong207. Our new findings suggest that the TaSHMT3A-1 gene in wheat plays an important role in resistance to Fusarium head blight. This provides a valuable reference for further study on the function of this gene family.

The Quality Changes and Proteomic Analysis of Cattle Muscle Postmortem during Rigor Mortis.

Rigor mortis occurs in a relatively early postmortem period and is a complex biochemical process in the conversion of muscle to meat. Understanding the quality changes and biomarkers during rigor mortis can provide a theoretical basis for maintaining and improving meat quality. Herein, a tandem mass tag proteomic method is used to investigate the effects of differentially expressed proteins on the meat quality of cattle Longissimus lumborum muscle postmortem (0, 6, and 24 h). The pH, total sulfhydryl content and sarcomere length decrease significantly during storage. In contrast, meat color values (L*, a*, and b*) and the myofibril fragmentation index increase significantly. Altogether, 147 differentially expressed proteins are identified, most being categorized as metabolic enzymes, mitochondrial proteins, necroptosis and ferroptosis proteins and structural proteins. The results also reveal additional proteins that are potentially involved in rigor mortis, such as cardiac phospholamban, acetyl-coenzyme A acyltransferase, and ankyrin repeat domain 2. The current results provide proteomic insights into the changes in meat quality during rigor mortis.

Proteomic analyses of mitochondrial damage in postmortem beef muscles.

BACKGROUND: The objective of the study was to examine the expression profiles of mitochondrial proteins in at-death and 24 h postmortem (PM) using tandem mass tag (TMT) approach to characterize the mitochondria possible mechanisms that are affiliated with tenderization. RESULTS: Results showed that the tender meat at 24 h PM emerged with more serious mitochondrial damage. Altogether 456 mitochondrial proteins were identified, including 442 down-regulated and 14 up-regulated proteins. These differentially-expressed proteins were primarily involved in the progress of PM energy metabolism, apoptosis, and the morphological alterations of mitochondrial. Among them, 47 subunits (such as NDUFA2, COX4I1, and ATP5PB) were annotated into the oxidative phosphorylation pathway. VDAC1, VDAC2, and VDAC3 involving in the damage of MPTP, and IMMT, CHCHD3, APOL and APOO modulating the morphology of mitochondria, and DIABLO and AIFM1 released from mitochondria affect caspase's activation. HSPD1 and HSPE1 involved in apoptosis, mitochondrial physiological and morphological alterations. CONCLUSION: The earlier-mentioned proteins were validated as potential indicators of tenderness regulated by mitochondrial damage. These results highlighted that mitochondrial damage possibly participate in PM tenderization of beef muscles by energy metabolism and cell apoptosis status. © 2022 Society of Chemical Industry.

A rapid, simple, and highly efficient method for VIGS and in vitro-inoculation of plant virus by INABS applied to crops that develop axillary buds and can survive from cuttings.

BACKGROUND: Virus-induced gene silencing (VIGS) is one of the most convenient and powerful methods of reverse genetics. In vitro-inoculation of plant virus is an important method for studying the interactions between viruses and plants. Agrobacterium-based infiltration has been widely adopted as a tool for VIGS and in vitro-inoculation of plant virus. Most agrobacterium-based infiltration methods applied to VIGS and virus inoculation have the characteristics of low transformation efficiencies, long plant growth time, large amounts of plant tissue, large test spaces, and complex preparation procedures. Therefore, a rapid, simple, economical, and highly efficient VIGS and virus inoculation method is in need. Previous studies have shown that the selection of suitable plant tissues and inoculation sites is the key to successful infection. RESULTS: In this study, Tobacco rattle virus (TRV) mediated VIGS and Tomato yellow leaf curl virus (TYLCV) for virus inoculation were developed in tomato plants based on the agrobacterium tumefaciens-based infiltration by injection of the no-apical-bud stem section (INABS). The no-apical-bud stem section had a "Y- type" asymmetric structure and contained an axillary bud that was about 1-3 cm in length. This protocol provides high transformation (56.7%) and inoculation efficiency (68.3%), which generates VIGS transformants or diseased plants in a very short period (8 dpi). Moreover, it greatly reduces the required experimental space. This method will facilitate functional genomic studies and large-scale disease resistance screening. CONCLUSIONS: Overall, a rapid, simple, and highly efficient method for VIGS and virus inoculation by INABS was developed in tomato. It was reasonable to believe that it can be used as a reference for the other virus inoculation methods and for the application of VIGS to other crops (such as sweet potato, potato, cassava and tobacco) that develop axillary buds and can survive from cuttings.

A Ricin B-Like Lectin Protein Physically Interacts with TaPFT and Is Involved in Resistance to Fusarium Head Blight in Wheat.

Fusarium head blight (FHB), mainly caused by Fusarium graminearum, has become one of the most serious diseases that damage wheat. The TaPFT (pore-forming toxin-like) and TaHRC (histidine-rich calcium-binding protein) genes at the quantitative trait locus Fhb1 were identified to confer resistance to FHB in the wheat cultivar Sumai 3. In this study, a wheat ricin B-like lectin gene (designated TaRBL) that interacted with TaPFT was isolated by a yeast two-hybrid screen of a wheat cDNA library. A yeast two-hybrid and bimolecular fluorescence complementation study further verified that TaRBL interacted with TaPFT but not with TaHRC. Gene expression studies showed that upon F. graminearum infection, TaRBL expression was upregulated in resistant cultivars but downregulated in susceptible cultivars. Furthermore, knockdown of TaRBL expression by barley stripe mosaic virus-induced gene silencing significantly reduced the resistance of wheat to FHB in both the resistant cultivar Sumai 3 and the susceptible cultivar Jimai 22. Thus, we conclude that TaRBL encodes a ricin B-like lectin protein that interacts with TaPFT and is involved in resistance to FHB in wheat.

Influence of oxidation on heat shock protein 27 translocation, caspase-3 and calpain activities and myofibrils degradation in postmortem beef muscles.

The objective of this study was to investigate the influence of oxidation on heat shock protein 27 (HSP27) and cytochrome c translocation, myofibrils degradation and endogenous enzymes activities, perfecting tenderization mechanism after slaughter. Bovine muscle (longissimus thoracis) was obtained at 30 min postmortem. Bovine muscle was cut and exposed to saline solution with or without H2O2 at 4 °C for 0.25, 1, 3 and 5 days, followed by detection of proteins degradation, location and enzymes activities. Results showed that oxidation promoted the translocation of HSP27 and cytochrome c from the cytoplasm to the cell membrane, which reduced µ-calpain activity, but increased caspase-3 activity through mediating the interaction with the two enzymes. Oxidation retarded troponin-T degradation, but accelerated desmin degradation, which is probably because oxidative modification of myofibrils induced different susceptibility to proteolysis. Therefore, oxidation leads to different regulatory mechanism on µ-calpain and caspase-3, as well as the degree of degradation of myofibrillar proteins, possibly through mediating HSP27 and cytochrome c.

Clay nanosheet-mediated delivery of recombinant plasmids expressing artificial miRNAs via leaf spray to prevent infection by plant DNA viruses.

Whitefly-transmitted begomoviruses are economically important plant pathogens that cause severe problems in many crop plants, such as tomato, papaya, cotton, and tobacco. Tomato yellow leaf curl virus (TYLCV) is a typical monopartite begomovirus that has been extensively studied, but methods that can efficiently control begomoviruses are still scarce. In this study, we combined artificial microRNA (amiRNA)-mediated silencing technology and clay nanosheet-mediated delivery by spraying and developed a method for efficiently preventing TYLCV infection in tomato plants. We designed three amiRNAs that target different regions of TYLCV to silence virus-produced transcripts. Three plant expression vectors expressing pre-amiRNAs were constructed, and recombinant plasmid DNAs (pDNAs) were loaded onto nontoxic and degradable layered double hydroxide (LDH) clay nanosheets. LDH nanosheets containing multiple pDNAs were sprayed onto plant leaves. We found that the designed amiRNAs were significantly accumulated in leaves 7 days after spraying, while the pDNAs were sustainably detected for 35 days after the spray, suggesting that the LDH nanosheets released pDNAs in a sustained manner, protected pDNAs from degradation and efficiently delivered pDNAs into plant cells. Importantly, when the LDH nanosheets coated with pDNAs were sprayed onto plants infected by TYLCV, both the disease severity and TYLCV viral concentration in sprayed plants were significantly decreased during the 35 days, while the levels of H2O2 were significantly increased in those plants. Taken together, these results indicate that LDH nanosheets loaded with pDNAs expressing amiRNAs can be a sustainable and promising tool for begomovirus control.

Quantitatively Identifying the Roles of Interfacial Water and Solid Surface in Governing Peptide Adsorption.

Understanding the molecular mechanism of protein adsorption on solids is critical to their applications in materials synthesis and tissue engineering. Although the water phase at the surface/water interface has been recognized as three types: bulk water, intermediate water phase and surface-bound water layers, the roles of the water and surface in determining the protein adsorption are not clearly identified, particularly at the quantitative level. Herein, we provide a methodology involving the combination of microsecond strengthen sampling simulation and force integration to quantitatively characterize the water-induced contribution and the peptide-surface interactions into the adsorption free energy. Using hydroxyapatite and graphene surfaces as examples, we demonstrate how the distinct interfacial features dominate the delicate force balance between these two thermodynamics parameters, leading to surface preference/resistance to peptide adsorption. Specifically, the water layer provides sustained repelling force against peptide adsorption, as indicated by a monotonic increase in the water-induced free energy profile, whereas the contribution from the surface-peptide interactions is thermodynamically favorable to peptide adsorptions. More importantly, the revealed adsorption mechanism is critically dictated by the distribution of water phase, which plays a crucial role in establishing the force balance between the interactions of the peptide with the water layer and the surface. For the HAP surface, the charged peptide exhibits strong binding affinity to the surface, due to the controlling contribution of peptide-surface interaction in the intermediate water phase. The surface-bound water layers are observed as the origin of bioresistance of solid surfaces toward the adsorption of charge-neutral peptides. The preferred peptide adsorption on the graphene, however, is dominated by the surface-induced component at the water layers adjacent to the surface. Our results further elucidate that the intermediate water phase significantly shortens the effective range of the surface dispersion force, in contrast to the observation on the hydrophilic surface.

Method Evaluations for Adsorption Free Energy Calculations at the Solid/Water Interface through Metadynamics, Umbrella Sampling, and Jarzynski's Equality.

Considerable interest in characterizing protein/peptide-surface interactions has prompted extensive computational studies on calculations of adsorption free energy. However, in many cases, each individual study has focused on the application of free energy calculations to a specific system; therefore, it is difficult to combine the results into a general picture for choosing an appropriate strategy for the system of interest. Herein, three well-established computational algorithms are systemically compared and evaluated to compute the adsorption free energy of small molecules on two representative surfaces. The results clearly demonstrate that the characteristics of studied interfacial systems have crucial effects on the accuracy and efficiency of the adsorption free energy calculations. For the hydrophobic surface, steered molecular dynamics exhibits the highest efficiency, which appears to be a favorable method of choice for enhanced sampling simulations. However, for the charged surface, only the umbrella sampling method has the ability to accurately explore the adsorption free energy surface. The affinity of the water layer to the surface significantly affects the performance of free energy calculation methods, especially at the region close to the surface. Therefore, a general principle of how to discriminate between methodological and sampling issues based on the interfacial characteristics of the system under investigation is proposed.