Combined Metabolomic and Quantitative RT-PCR Analyses Revealed the Synthetic Differences of 2-Acetyl-1-pyrroline in Aromatic and Non-Aromatic Vegetable Soybeans

Aroma is an important economic trait of vegetable soybeans, which greatly influences their market value. The 2-acetyl-1-pyrroline (2AP) is considered as an important substance affecting the aroma of plants. Although the 2AP synthesis pathway has been resolved, the differences of the 2AP synthesis in the aromatic and non-aromatic vegetable soybeans are unknown. In this study, a broad targeted metabolome analysis including measurement of metabolites levels and gene expression levels was performed to reveal pathways of aroma formation in the two developmental stages of vegetable soybean grains [35 (S5) and 40 (S6) days after anthesis] of the ‘Zhexian No. 8’ (ZX8, non-aromatic) and ZK1754 (aromatic). The results showed that the differentially accumulated metabolites (DAMs) of the two varieties can be classified into nine main categories including flavonoids, lipids, amino acids and derivatives, saccharides and alcohols, organic acids, nucleotides and derivatives, phenolic acids, alkaloids and vitamin, which mainly contributed to their phenotypic differences. Furthermore, in combination with the 2AP synthesis pathway, the differences of amino acids and derivatives were mainly involved in the 2AP synthesis. Furthermore, 2AP precursors’ analysis revealed that the accumulation of 2AP mainly occurred from 1-pyrroline-5-carboxylate (P5C), not 4-aminobutyraldehyde (GABald). The quantitative RT-PCR showed that the associated synthetic genes were 1-pyrroline-5-carboxylate dehydrogenase (P5CDH), ∆¹-pyrroline-5-carboxylate synthetase (P5CS), proline dehydrogenase (PRODH) and pyrroline-5-carboxylate reductase (P5CR), which further verified the synthetic pathway of 2AP. Furthermore, the betaine aldehyde dehydrogenase 2 (GmBADH2) mutant was not only vital for the occurrence of 2AP, but also for the synthesis of 4-aminobutyric acid (GABA) in vegetable soybean. Therefore, the differences of 2AP accumulation in aromatic and non-aromatic vegetable soybeans have been revealed, and it also provides an important theoretical basis for aromatic vegetable soybean breeding.     

Mechanism of [CO2] Enrichment Alleviated Drought Stress in the Roots of Cucumber Seedlings Revealed via Proteomic and Biochemical Analysis

Cucumber is one of the most widely cultivated greenhouse vegetables, and its quality and yield are threatened by drought stress. Studies have shown that carbon dioxide concentration ([CO₂]) enrichment can alleviate drought stress in cucumber seedlings; however the mechanism of this [CO₂] enrichment effect on root drought stress is not clear. In this study, the effects of different drought stresses (simulated with 0, 5% and 10% PEG 6000, i.e., no, moderate, and severe drought stress) and [CO₂] (400 μmol·mol⁻¹ and 800 ± 40 μmol·mol⁻¹) on the cucumber seedling root proteome were analyzed using the tandem mass tag (TMT) quantitative proteomics method. The results showed that after [CO₂] enrichment, 346 differentially accumulating proteins (DAPs) were found only under moderate drought stress, 27 DAPs only under severe drought stress, and 34 DAPs under both moderate and severe drought stress. [CO₂] enrichment promoted energy metabolism, amino acid metabolism, and secondary metabolism, induced the expression of proteins related to root cell wall and cytoskeleton metabolism, effectively maintained the balance of protein processing and degradation, and enhanced the cell wall regulation ability. However, the extent to which [CO₂] enrichment alleviated drought stress in cucumber seedling roots was limited under severe drought stress, which may be due to excessive damage to the seedlings.     

Increasing the Grain Yield and Grain Protein Content of Common Wheat (Triticum aestivum) by Introducing Missense Mutations in the Q Gene

Grain yield (GY) and grain protein content (GPC) are important traits for wheat breeding and production; however, they are usually negatively correlated. The Q gene is the most important domestication gene in cultivated wheat because it influences many traits, including GY and GPC. Allelic variations in the Q gene may positively affect both GY and GPC. Accordingly, we characterized two new Q alleles (Q^s1 and Q^c1-N8) obtained through ethyl methanesulfonate-induced mutagenesis. Compared with the wild-type Q allele, Q^s1 contains a missense mutation in the sequence encoding the first AP₂ domain, whereas Q^c1-N8 has two missense mutations: one in the sequence encoding the second AP₂ domain and the other in the microRNA172-binding site. The Q^s1 allele did not significantly affect GPC or other processing quality parameters, but it adversely affected GY by decreasing the thousand kernel weight and grain number per spike. In contrast, Q^c1-N8 positively affected GPC and GY by increasing the thousand kernel weight and grain number per spike. Thus, we generated novel germplasm relevant for wheat breeding. A specific molecular marker was developed to facilitate the use of the Q^c1-N8 allele in breeding. Furthermore, our findings provide useful new information for enhancing cereal crops via non-transgenic approaches.     

CircTCF4 Suppresses Proliferation and Differentiation of Goat Skeletal Muscle Satellite Cells Independent from AGO2 Binding

The proliferation and differentiation of mammalian skeletal muscle satellite cells (MuSCs) are highly complicated. Apart from the regulatory signaling cascade driven by the protein-coding genes, non-coding RNAs such as microRNAs (miRNA) and circular RNAs (circRNAs) play essential roles in this biological process. However, circRNA functions in MuSCs proliferation and differentiation remain largely to be elucidated. Here, we screened for an exonic circTCF4 based on our previous RNA-Seq data, specifically expressed during the development of the longest dorsal muscle in goats. Subsequently, the circular structure and whole sequence of circTCF4 were verified using Sanger sequencing. Besides, circTCF4 was spatiotemporally expressed in multiple tissues from goats but strikingly enriched in muscles. Furthermore, circTCF4 suppressed MuSCs proliferation and differentiation, independent of AGO2 binding. Finally, we conducted Poly(A) RNA-Seq using cells treated with small interfering RNA targeting circTCF4 and found that circTCF4 would affect multiple signaling pathways, including the insulin signaling pathway and AMPK signaling pathway related to muscle differentiation. Our results provide additional solid evidence for circRNA regulating skeletal muscle formation.     

Ontogenetic Variation in Macrocyclic and Hemicyclic Poplar Rust Fungi

Melampsora larici-populina (Mlp), M. medusae (Mmed), M. magnusiana (Mmag), and M. pruinosae (Mpr) are epidemic rust fungi in China. The first two are macrocyclic rust fungi distributed in temperate humid environments. The latter two are hemicyclic rusts, mainly distributed in arid and semi-arid areas. Ontogenetic variation that comes with this arid-resistance is of great interest—and may help us predict the influence of a warmer, drier, climate on fungal phylogeny. To compare the differences in the life history and ontogeny between the two types of rust, we cloned mating type genes, STE3.4 and STE3.3 using RACE-smart technology. Protein structures, functions, and mutant loci were compared across each species. We also used microscopy to compare visible cytological differences at each life stage for the fungal species, looking for variation in structure and developmental timing. Quantitative PCR technology was used to check the expression of nuclear fusion and division genes downstream of STE3.3 and STE3.4. Encoding amino acids of STE3.3 and STE3.4 in hemicyclic rusts are shorter than these in the macrocyclic rusts. Both STE3.3 and STE3.4 interact with a protein kinase superfamily member {"type":"entrez-protein","attrs":{"text":"EGG12818","term_id":"328863719","term_text":"EGG12818"}}EGG12818 and an E3 ubiquitin protein ligase {"type":"entrez-protein","attrs":{"text":"EGG09709","term_id":"328860604","term_text":"EGG09709"}}EGG09709 directly, and activating G-beta conformational changes. The mutation at site 74th amino acid in the conserved transmembrane domain of STE3.3 ascribes to a positive selection, in which alanine (Ala) is changed to phenylalanine (Phe) in hemicyclic rusts, and a mutation with Tyr lost at site 387th in STE3.4, where it is the binding site for β-D-Glucan. These mutants are speculated corresponding to the insensitivity of hemicyclic rust pheromone receptors to interact with MFa pheromones, and lead to Mnd1 unexpressed in teliospora, and they result in the diploid nuclei division failure and the sexual stage missing in the life cycle. A Phylogenic tree based on STE3.4 gene suggests these two rust types diverged about 14.36 million years ago. Although these rusts share a similar uredia and telia stage, they show markedly different wintering strategies. Hemicyclic rusts overwinter in the poplar buds endophytically, their urediniospores developing thicker cell walls. They form haustoria with a collar-like extrahaustorial membrane neck and induce host thickened callose cell walls, all ontogenetic adaptations to arid environments.     

Serum SELENBP1 and VCL Are Effective Biomarkers for Clinical and Forensic Diagnosis of Coronary Artery Spasm

Coronary artery spasm (CAS) plays an important role in the pathogenesis of many ischemic heart entities; however, there are no established diagnostic biomarkers for CAS in clinical and forensic settings. This present study aimed to identify such serum biomarkers by establishing a rabbit CAS provocation model and integrating quantitative serum proteomics, parallel reaction monitoring/mass spectrometry-based targeted proteomics, and partial least-squares discriminant analysis (PLS-DA). Our results suggested that SELENBP1 and VCL were potential candidate biomarkers for CAS. In independent clinical samples, SELENBP1 and VCL were validated to be significantly lower in serum but not blood cells from CAS patients, with the reasons for this possibly due to the decreased secretion from cardiomyocytes. The areas under the curve of the receiver operating characteristics (ROC) analysis were 0.9384 for SELENBP1 and 0.9180 for VCL when diagnosing CAS. The CAS risk decreased by 32.3% and 53.6% for every 10 unit increases in the serum SELENBP1 and VCL, respectively. In forensic samples, serum SELENBP1 alone diagnosed CAS-induced deaths at a sensitivity of 100.0% and specificity of 72.73%, and its combination with VCL yielded a diagnostic specificity of 100.0%, which was superior to the traditional biomarkers of cTnI and CK-MB. Therefore, serum SELENBP1 and VCL could be effective biomarkers for both the clinical and forensic diagnosis of CAS.     

Performance Analysis of a HT-PEMFC System with 6FPBI Membranes Doped with Cross-Linkable Polymeric Ionic Liquid

In this paper, a high-temperature proton-exchange membrane fuel cell (HT-PEMFC) system using fluorine-containing polybenzimidazole (6FPBI) composite membranes doped with cross-linkable polymer ionic liquid (cPIL) is developed and studied. The reliability of the model is verified by a comparison with the experimental data. The performance of the HT-PEMFC system using 6FPBI membranes with different levels of cPIL is analyzed. The results show that when the HT-PEMFC uses 6FPBI membranes with a cPIL content of 20 wt % (6FPBI-cPIL 20 membranes), the single cell power density is 4952.3 W·m−2. The excessive cPIL content will lead to HT-PEMFC performance degradation. The HT-PEMFC system using the 6FPBI-cPIL 20 membranes shows a higher performance, even at higher temperatures and pressures, than the systems using 6FPBI membranes. In addition, the parametric study results suggest that the HT-PEMFC system should be operated at a higher inlet temperature and hydrogen pressure to increase system output power and efficiency. The oxygen inlet pressure should be reduced to decrease the power consumption of the ancillary equipment and improve system efficiency. The proposed model can provide a prediction for the performance of HT-PEMFC systems with the application of phosphoric-acid-doped polybenzimidazole (PA-PBI) membranes.     

Agrobacterium sp. ZX09 β-Glucan Attenuates Enterotoxigenic Escherichia coli-Induced Disruption of Intestinal Epithelium in Weaned Pigs

To explore the protective effect of dietary β-glucan (BGL) supplementation on intestinal epithelium exposure to enterotoxigenic Escherichia coli (ETEC), thirty-two weaned pigs were assigned to four groups. Pigs were fed with a basal diet or basal diet containing 500 mg/kg BGL, and were orally infused with ETEC or culture medium. Results showed BGL supplementation had no influence on growth performance in weaned pigs. However, BGL supplementation increased the absorption of D-xylose, and significantly decreased the serum concentrations of D-lactate and diamine oxidase (DAO) in the ETEC-challenged pigs (p < 0.05). Interestingly, BGL significantly increased the abundance of the zonula occludens-1-(ZO-1) in the jejunal epithelium upon ETEC challenge (p < 0.05). BGL supplementation also increased the number of S-phase cells and the number of sIgA-positive cells, but significantly decreased the number of total apoptotic cells in the jejunal epithelium upon ETEC challenge (p < 0.05). Moreover, BGL significantly increased the duodenal catalase (CAT) activity and the ileal total superoxide dismutase (T-SOD) activity in the ETEC-challenged pigs (p < 0.05). Importantly, BGL significantly decreased the expression levels of critical inflammation related proteins such as the tumor necrosis factor-α (TNF-α), interlukin-6 (IL-6), myeloid differentiation factor 88 (MyD88), and nuclear factor-κB (NF-κB) in the jejunal and ileal mucosa upon ETEC challenge (p < 0.05). BGL also elevated the propanoic acid content and the abundance of Lactobacillus and Bacillus in the colon upon ETEC challenge (p < 0.05). These results suggested BGL could alleviate the ETEC-induced intestinal epithelium injury, which may be associated with suppressed inflammation and improved intestinal immunity and antioxidant capacity, as well as the improved intestinal macrobiotic.