Large scale rice germplasm screening for identification of novel brown planthopper resistance sources.

Rice (Oryza sativa L.) is a staple food crop globally. Brown planthopper (Nilaparvata lugens Stål, BPH) is the most destructive insect that threatens rice production annually. More than 40 BPH resistance genes have been identified so far, which provide valuable gene resources for marker-assisted breeding against BPH. However, it is still urgent to evaluate rice germplasms and to explore more new wide-spectrum BPH resistance genes to combat newly occurring virulent BPH populations. To this end, 560 germplasm accessions were collected from the International Rice Research Institute (IRRI), and their resistance to current BPH population of China was examined. A total of 105 highly resistant materials were identified. Molecular screening of BPH resistance genes in these rice germplasms was conducted by developing specific functional molecular markers of eight cloned resistance genes. Twenty-three resistant germplasms were found to contain none of the 8 cloned BPH resistance genes. These accessions also exhibited a variety of resistance mechanisms as indicated by an improved insect weight gain (WG) method, suggesting the existence of new resistance genes. One new BPH resistance gene, Bph44(t), was identified in rice accession IRGC 15344 and preliminarily mapped to a 0-2 Mb region on chromosome 4. This study systematically sorted out the corresponding relationships between BPH resistance genes and germplasm resources using a functional molecular marker system. Newly explored resistant germplasms will provide valualble donors for the identification of new resistance genes and BPH resistance breeding programs. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-023-01416-x.

Molecular and functional analysis of a brown planthopper resistance protein with two nucleotide-binding site domains.

The brown planthopper (Nilaparvata lugens Stål, BPH) resistance gene BPH9 encodes an unusual coiled-coil (CC) nucleotide-binding leucine-rich repeat (LRR) protein with two nucleotide-binding site (NBS) domains. To understand how this CC-NBS-NBS-LRR (CNNL) protein regulates defense signaling and BPH resistance, we dissected each domain's functions. The CC domain of BPH9 self-associated and was sufficient to induce cell death. The region of 97-115 residues in the CC domain is crucial for self-association and activation. NBS2, which contains a complete set of NBS function motifs and inhibits CC domain activation, rather than NBS1, acts as a molecular switch to regulate the activity of BPH9. We demonstrated that the CC domain, the NBS domain, and the LRR domain of BPH9 associate with each other and themselves in planta. Further domain swapping experiments revealed that the CC domains of BPH9 and susceptible alleles were similarly competent to induce resistance and the hypersensitive response, while the LRR domain of BPH9 confers resistance specificity to BPH. These findings provide new insights into the regulatory mechanisms governing the activity of CNNL proteins.

Combining next-generation sequencing and single-molecule sequencing to explore brown plant hopper responses to contrasting genotypes of japonica rice.

BACKGROUND: The brown plant hopper (BPH), Nilaparvata lugens, is one of the major pest of rice (Oryza sativa). Plant defenses against insect herbivores have been extensively studied, but our understanding of insect responses to host plants' resistance mechanisms is still limited. The purpose of this study is to characterize transcripts of BPH and reveal the responses of BPH insects to resistant rice at transcription level by using the advanced molecular techniques, the next-generation sequencing (NGS) and the single-molecule, real-time (SMRT) sequencing. RESULTS: The current study obtained 24,891 collapsed isoforms of full-length transcripts, and 20,662 were mapped to known annotated genes, including 17,175 novel transcripts. The current study also identified 915 fusion genes, 1794 novel genes, 2435 long non-coding RNAs (lncRNAs), and 20,356 alternative splicing events. Moreover, analysis of differentially expressed genes (DEGs) revealed that genes involved in metabolic and cell proliferation processes were significantly enriched in up-regulated and down-regulated sets, respectively, in BPH fed on resistant rice relative to BPH fed on susceptible wild type rice. Furthermore, the FoxO signaling pathway was involved and genes related to BPH starvation response (Nlbmm), apoptosis and autophagy (caspase 8, ATG13, BNIP3 and IAP), active oxygen elimination (catalase, MSR, ferritin) and detoxification (GST, CarE) were up-regulated in BPH responses to resistant rice. CONCLUSIONS: The current study provides the first demonstrations of the full diversity and complexity of the BPH transcriptome, and indicates that BPH responses to rice resistance, might be related to starvation stress responses, nutrient transformation, oxidative decomposition, and detoxification. The current result findings will facilitate further exploration of molecular mechanisms of interaction between BPH insects and host rice.

Xianglian pill alleviates ulcerative colitis by inhibiting M1 macrophage polarization via modulation of energy metabolite itaconate.

BACKGROUND: Xianglian pill (XLP) is a traditional Chinese medicine (TCM) that is widely used to treat ulcerative colitis (UC). However, its mechanism of action in UC is unclear. PURPOSE: This study aimed to investigate the mechanism of action of XLP in treating UC and role of M1 macrophage polarization in this process. STUDY DESIGN: In vivo experiments were performed using UC mice while in vitro experiments were conducted using RAW264.7 cells. METHODS: Mice were administered 3 % dextran sulfate to induce UC model and then treated with XLP. Changes in histopathology and pro-inflammatory cytokines were evaluated. The levels of M1 macrophages in mesenteric lymph nodes were detected by flow cytometry. Colon metabolite levels were analyzed using an energy metabolomic assay. To assess itaconate's impact, both in vivo (mice) and in vitro (RAW264.7 cells) models were employed. Immunofluorescence staining was used to measure the expression levels of TNF-α, IL-6, and iNOS, while qRT-PCR was utilized to quantify the mRNA levels of TET2, STAT1, and Nfkbiz. RESULTS: XLP alleviated ulcerative damage and reduced TNF-α and IL-6 levels in colon, and also downregulated the levels of M1 macrophages and modulated the state of energy metabolism. Specifically, XLP significantly increased ITA level in colonic tissue and this increase was significantly associated with decreased levels of M1 macrophages and alleviation of UC following XLP treatment. Moreover, ITA directly suppressed the polarization of macrophage from M0 to M1 phenotype, accompanied by the decrease of TNF-α, IL-6, and iNOS levels. Further, ITA decreased inflammatory responses in M1 macrophage by inhibiting the TET2/STAT1 and TET2/NF-κB signaling pathways. CONCLUSION: XLP can treat UC by suppressing M1 macrophage polarization via increasing the level of energy metabolite ITA.

Coptidis Rhizoma processed with Evodia Rutaecarpa improves the effect on ulcerative colitis by increasing intestinal energy metabolites alpha-ketoglutarate and Lactobacillus reuteri.

BACKGROUND: Evodia Rutaecarpa-processed Coptidis Rhizoma (ECR) is a traditional Chinese medicine for the treatment of ulcerative colitis (UC) in China. However, the mechanisms underlying the ECR processing are not elucidated. PURPOSE: Coptidis Rhizoma (CR) regulates the gut microbiota in the treatment of gastrointestinal diseases. This study explored the mechanism of action of ECR before and after processing in UC in view of the regulation of gut microecology. STUDY DESIGN: A preclinical experimental investigation was performed using a mouse model of UC to examine the regulatory effect of ECR and its mechanisms through gut microbiota analysis and metabolomic assays. METHODS: Mice received 4% dextran sulfate sodium to establish a UC model and treated with ECR and CR. Colonic histopathology and inflammatory changes were observed. Gut microbiota was analyzed using 16 s rRNA sequencing. Transplants of Lactobacillus reuteri were used to explore the correlation between ECR processing and the gut microbiota. The expression of mucin-2, Lgr5, and PCNA in colonic epithelial cells was measured using immunofluorescence. Wnt3a and ß-catenin levels were detected by western blotting. The metabolites in the colon tissue were analyzed using a targeted energy metabolomic assay. The effect of energy metabolite α-ketoglutarate (α-KG) on L. reuteri growth and UC were verified in mice. RESULTS: ECR improved the effects on UC in mice compared to CR, including alleviating colonic injury and inflammation, and modulating gut microbiota by increasing L. reuteri level. L. reuteri dose-dependently alleviated colonic injury, increased mucin-2 level, and promoted colonic epithelial regeneration by increasing Lgr5 and PCNA expression. This was consistent with the results before and after ECR processing. L. reuteri promoted epithelial regeneration by upregulating Wnt/ß-catenin pathway. Moreover, ECR increased metabolites levels (especially α-KG) to promote energy metabolism in the colon tissue compared to CR. α-KG treatment increased L. reuteri level and alleviated mucosal damage in UC mice. It promoted L. reuteri growth by increasing the energy metabolic status by enhancing α-KG dehydrogenase activity. CONCLUSION: ECR processing improves the therapeutic effects of UC via the α-KG-L. reuteri-epithelial regeneration axis.

Dual AuNPs detecting probe enhanced the NanoSPR effect for the high-throughput detection of the cancer microRNA21 biomarker.

The microRNA21 (miR-21), a specific tumor biomarker, is crucial for the diagnosis of several cancer types, and investigation of its overexpression pattern is important for cancer diagnosis. Herein, we report a low-cost, rapid, ultrasensitive, and convenient biosensing strategy for the detection of miR-21 using a nanoplasmonic array chip coupled with gold nanoparticles (AuNPs). This sensing platform combines the surface plasmon resonance effect of nanoplasmonics (NanoSPR) and the localized surface plasmon resonance (LSPR) effect, which allows the real-time monitoring of the subtle optical density (OD) changes caused by the variations in the dielectric constant in the process of the hybridization of the target miRNA. Using this method, the miRNA achieves a broad detection range from 100 aM to 1 µM, and with a limit of detection (LoD) of 1.85 aM. Furthermore, this assay also has a single-base resolution to discriminate the highly homologous miRNAs. More importantly, this platform has high throughput characteristics (96 samples can be detected simultaneously). This strategy exhibits more than 86.5 times enhancement in terms of sensitivity compared to that of traditional biosensors.

Xianglian pill modulates gut microbial production of succinate and induces regulatory T cells to alleviate ulcerative colitis in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Xianglian pill (XLP), a traditional Chinese formula, is widely used as treatment for ulcerative colitis (UC) in China. However, the mechanism of its therapeutic effect is still unclear. AIM OF THE STUDY: Our previous studies showed a low oral bioavailability and a predominant distribution of major XLP ingredients in the gut. In the present study, we aimed to explore the mechanism of action of XLP on UC with respect to the regulation of gut microecology. MATERIALS AND METHODS: UC model rats established using 5% dextran sulfate sodium were treated with XLP. After the treatment period, bodyweight, colon length, histopathology, and inflammatory changes were evaluated. Further, changes in gut microbiota structure were detected via 16S rRNA sequencing, and microbial metabolites in feces were analyzed via a metabolomic assay. Antibiotic intervention and fecal microbiota transplantation were also employed to explore the involvement of gut microbiota, while the level of regulatory T cells (Tregs) in mesenteric lymph nodes was determined via flow cytometry. Transcriptome sequencing was also performed to determine colonic gene changes. RESULTS: XLP alleviated colonic injury, inflammation, and gut microbial dysbiosis in UC model rats and also changed microbial metabolite levels. Particularly, it significantly decreased succinate level in the tyrosine pathway. We also observed that fecal microbiota derived from XLP-treated rats conferred resilience to UC model rats. However, this therapeutic effect of XLP on UC was inhibited by succinate. Moreover, XLP increased the level of anti-inflammatory cellular Tregs via gut microbiota. However, this beneficial effect was counteracted by succinate supplementation. Further, XLP induced the differentiation of Treg possibly by the regulation of the PHD2/HIF-1α pathway via decreasing microbial succinate production. CONCLUSIONS: Our findings indicated that XLP exerts its therapeutic effects on UC mainly via the gut microbiota-succinate-Treg differentiation axis.

Predicting gram-positive bacterial protein subcellular localization based on localization motifs.

The subcellular localization of proteins is closely related to their functions. In this work, we propose a novel approach based on localization motifs to improve the accuracy of predicting subcellular localization of Gram-positive bacterial proteins. Our approach performed well on a five-fold cross validation with an overall success rate of 89.5%. Besides, the overall success rate of an independent testing dataset was 97.7%. Moreover, our approach was tested using a new experimentally-determined set of Gram-positive bacteria proteins and achieved an overall success rate of 96.3%.