Case report: Rubella virus-associated cutaneous granuloma in an adult with TAP1 deficiency.

Rubella virus-associated granulomas commonly occur in immunocompromised individuals, exhibiting a diverse range of clinical presentations. These manifestations can vary from predominantly superficial cutaneous plaques or nonulcerative nodules to more severe deep ulcerative lesions, often accompanied by extensive necrosis and significant tissue destruction. TAP1 deficiency, an exceedingly rare primary immune-deficiency disorder, presents with severe chronic sino-pulmonary infection and cutaneous granulomas. This report highlights the occurrence of rubella virus-associated cutaneous granulomas in patients with TAP1 deficiency. Notably, the pathogenic mutation responsible for TAP1 deficiency stems from a novel genetic alteration that has not been previously reported. This novel observation holds potential significance for the field of diagnosis and investigative efforts in the context of immunodeficiency disorders.

An alternative splicing isoform of wheat TaYRG1 resistance protein activates immunity by interacting with dynamin-related proteins.

Wheat (Triticum aestivum) is a commercially important crop and its production is seriously threatened by the fungal pathogen Puccinia striiformis f. sp. tritici West (Pst). Resistance (R) genes are critical factors that facilitate plant immune responses. Here, we report a wheat R gene NB-ARC-LRR ortholog, TaYRG1, that is associated with distinct alternative splicing events in wheat infected by Pst. The native splice variant, TaYRG1.6, encodes internal-motif-deleted polypeptides with the same N- and C-termini as TaYRG1.1, resulting in gain of function. Transient expression of protein variants in Nicotiana benthamiana showed that the NB and ARC domains, and TaYRG1.6 (half LRR domain), stimulate robust elicitor-independent cell death based on a signal peptide, although the activity was negatively modulated by the CC and complete LRR domains. Furthermore, molecular genetic analyses indicated that TaYRG1.6 enhanced resistance to Pst in wheat. Moreover, we provide multiple lines of evidence that TaYRG1.6 interacts with a dynamin-related protein, TaDrp1. Proteome profiling suggested that the TaYRG1.6-TaDrp1-DNM complex in the membrane trafficking systems may trigger cell death by mobilizing lipid and kinase signaling in the endocytosis pathway. Our findings reveal a unique mechanism by which TaYRG1 activates cell death and enhances disease resistance by reconfiguring protein structure through alternative splicing.

CRISPR-based detection of Helicobacter pylori in stool samples.

BACKGROUND: Noninvasive detection of Helicobacter pylori plays an important role in clinical practice. However, few noninvasive methods have been applied in epidemiological studies due to the requirement for expensive equipment and complicated processes. The aim of this study was to establish a reliable, fast, and inexpensive noninvasive method based on CRISPR-Cas12a technology for the detection of Helicobacter pylori in stool specimens. METHOD: A novel detection method based on CRISPR-Cas12a technology was established and validated with 41 stool specimens collected from Zhujiang Hospital and compared with reliable Helicobacter pylori detection assays, such as the rapid urease test and urea breath test. RESULT: A CRISPR-Cas12a system-based method was established, and its sensitivity and specificity were evaluated. Utilizing a lateral flow biosensor, the limit of detection was 5 copies/µl, and our method could successfully distinguish Helicobacter pylori from other pathogens, suggesting no cross-reactivity with other pathogens. Furthermore, lateral flow biosensor strips were utilized to test stool specimens, which could display the detection results in an accessible way. CONCLUSION: Our CRISPR-Cas12a system-based method successfully detected Helicobacter pylori in stool specimens. It is a rapid, simple, and inexpensive method for the detection and screening of Helicobacter pylori, which makes it a very promising supplemental test. However, its sensitivity and specificity compared with those of the gold standard test still need to be examined.

Discovery of Natural Products Targeting NQO1 via an Approach Combining Network-Based Inference and Identification of Privileged Substructures.

NAD(P)H:quinone oxidoreductase 1 (NQO1) has been shown to be a potential therapeutic target for various human diseases, such as cancer and neurodegenerative disorders. Recent advances in computational methods, especially network-based methods, have made it possible to identify novel compounds for a target with high efficiency and low cost. In this study, we designed a workflow combining network-based methods and identification of privileged substructures to discover new compounds targeting NQO1 from a natural product library. According to the prediction results, we purchased 56 compounds for experimental validation. Without the assistance of privileged substructures, 31 compounds (31/56 = 55.4%) showed IC50 < 100 µM, and 11 compounds (11/56 = 19.6%) showed IC50 < 10 µM. With the assistance of privileged substructures, the two success rates were increased to 61.8 and 26.5%, respectively. Seven natural products further showed inhibitory activity on NQO1 at the cellular level, which may serve as lead compounds for further development. Moreover, network analysis revealed that osthole may exert anticancer effects against multiple cancer types by inhibiting not only carbonic anhydrases IX and XII but also NQO1. Our workflow and computational methods can be easily applied in other targets and become useful tools in drug discovery and development.

Effects of endogenous and exogenous dissolved organic matter on sorption behaviors of bisphenol A onto soils.

The transport of organic contaminants in groundwater might be greatly affected by coexistence of dissolved organic matter (DOM) from different sources. In this study, the effects of endogenous and exogenous DOMs (referred to as DOMen and DOMex, respectively) on sorption behavior of bisphenol A (BPA) onto two reference soils were investigated by batch experiments and microscopic characterization. The results showed that BPA sorption onto soils was dominated by soil organic matter content and affected by DOM properties. The effect of DOMen on BPA sorption was also related to the inorganic components of the two soils. The decrease of organic matter content reduced the sorption capacity of fluvo-aquic soil. However, because the content of available inorganic components in black soil was high, after removing DOMen, more inorganic sites were exposed to increase the sorption capacity. In addition, DOMen could form complexes with BPA in solution, thus the removal of DOMen promoted BPA sorption onto black soil. Under the experimental conditions, contribution of DOMex to the total sorption of BPA onto both soils was not more than 30%. Results of dialysis experiments and soil sorption experiments indicated that effects of coexisting DOMex on BPA sorption was related to the affinity of DOMex to soils and complexation of BPA and DOMex. Since the affinity of DOMex to fluvo-aquic soil was relatively low, the complex of BPA and DOMex in solution was the main inhibition mechanism for BPA sorption. For black soil, higher complexation proportion of BPA with DOMex adsorbed onto soil which promoted BPA sorption onto soil. The findings are of significance for understanding the co-migration of DOM with BPA through soils.

Differences in the dielectric properties of various benign and malignant thyroid nodules.

PURPOSE: This experiment was conducted to investigate the dielectric properties of different types of thyroid nodules. Our goal was to find a simple and fast method to detect thyroid diseases at different stages from the dielectric properties of thyroid nodules. METHODS: We used the open-ended coaxial line method to measure the dielectric permittivities of thyroid tissues from 155 patients at frequencies ranging from 1 to 4000 MHz. Tissues that were investigated included normal thyroid tissue and benign and malignant thyroid nodules (nodular goiter, follicular adenoma, papillary carcinoma, and follicular carcinoma), as determined from pathological reports. Differences in dielectric properties were measured between each nodule and the surrounding 1 cm of tissue. RESULTS: The analysis results revealed that the dielectric permittivity and conductivity values were positively correlated with the degree of malignancy of the nodule (normal < benign < malignant; all differences P < 0.05). This was more obvious at frequencies within 20~70 MHz, following the order normal tissue < nodular goiter < follicular adenoma < papillary carcinoma < follicular carcinoma. A significant difference (P < 0.05) in dielectric permittivity and conductivity was found when comparing these nodules with the surrounding 1 cm of tissue. CONCLUSIONS: Normal, benign, and malignant nodules were successfully distinguished from one another, and dielectric permittivity was found to be a more sensitive parameter than conductivity. In particular, different disease types can be distinguished at a stimulation frequency of 20~70 MHz, which shows that dielectric properties have application prospects for the detection and diagnosis of cancer. At the same time, the dielectric parameter differences between the surrounding 1 cm of tissue and the diseased nodule can distinguish the tumor and its surrounding tissues in real time during surgery to determine the tumor boundary.

Large-Scale Cloning and Comparative Analysis of TaNAC Genes in Response to Stripe Rust and Powdery Mildew in Wheat (Triticum aestivum L.).

The NAM, ATAF1/2, and CUC2 (NAC) transcription factors (TFs) constitute the largest plant-specific TF superfamily, and play important roles in various physiological processes, including stress responses. Stripe rust and powdery mildew are the most damaging of the fungal diseases that afflict wheat (Triticum aestivum L.). However, studies on Triticum aestivum NAC (TaNAC)s' role in resistance to the two diseases are still limited, especially in an overall comparative analysis of TaNACs responding or not to fungal stress. In the present study, 186 TaNAC transcripts were obtained from the resistant hexaploid wheat line N9134 under fungal stress, and 180 new transcripts were submitted to GenBank. Statistical results show that 35.1% (54/154) of TaNAC genes responded to stripe rust and powdery mildew in the seedling stage. "Abnormal" coding transcripts of differentially expressed (DE)-TaNAC genes in wheat responding to fungal stress were found in a significantly higher proportion (24/117 vs. 8/69, p = 0.0098) than in non-DE-NACs. This hinted that the alternative splicing of TaNAC genes was active in transcriptional or post-transcriptional regulation during plant-pathogen interactions. Full-length NAC proteins were classified into nine groups via phylogenetic analysis. Multiple-sequence alignment revealed diversity in the C-terminal structural organization, but the differentially expressed gene (DEG)-encoding proteins enriched in Subgroups VI and VII were conserved, with WV[L/V]CR amino acid residues in Motif 7 following the NAM domain. Our data that showed TaNAC TFs responded to fungal disease, which was affected by expression levels and by the regulation of multifarious transcript variants. These data for TaNAC responses to stripe rust and/or powdery mildew and their numerous structural variants provide a good resource for NAC function-mechanism analysis in the context of biotic-stress tolerance in wheat.

Gene co-expression network analysis provides a novel insight into the dynamic response of wheat to powdery mildew stress.

Powdery mildew (Blumeria graminis f. sp. Tritici, (Bgt)) is an important worldwide fungal foliar disease of wheat (Triticum aestivum) responsible for severe yield losses. The development of resistance genes and dissection of the resistance mechanism will therefore be beneficial in wheat breeding. The Bgt resistance gene PmAS846 was transferred to the hexaploid wheat lines N9134 from Triticum dicoccoides, and it is still one of the most effective resistance genes. Here, by RNA sequencing, we identified three co-expressed gene modules using pairwise comparisons and weighted gene co-expression network analysis during wheat-Bgt interactions compared with mock-infected plants. Hub genes of stress-specific modules were significantly enriched in spliceosomes, phagosomes, the mRNA surveillance pathway, protein processing in the endoplasmic reticulum, and endocytosis. Induced module genes located on chromosome 5BL were selected to construct a protein-protein interaction network. Several proteins were predicted as the key hub node, including Hsp70, DEAD/DEAH box RNA helicase PRH75, elongation factor EF-2, cell division cycle 5, ARF guanine-nucleotide exchange factor GNOM-like, and protein phosphatase 2C 70 protein, which interacted with several disease resistance proteins such as RLP37, RPP13 and RPS2 analogues. Gene ontology enrichment results showed that wheat could activate binding functional genes via an mRNA transcription mechanism in response to Bgt stress. Of these node genes, GNOM-like, PP2C isoform X1 and transmembrane 9 superfamily member 9 were mapped onto the genetic fragment of PmAS846 with a distance of 4.8 Mb. This work provides the foundations for understanding the resistance mechanism and cloning the resistance gene PmAS846.

Illuminating NAD+ Metabolism in Live Cells and In Vivo Using a Genetically Encoded Fluorescent Sensor.

Understanding of NAD+ metabolism provides many critical insights into health and diseases, yet highly sensitive and specific detection of NAD+ metabolism in live cells and in vivo remains difficult. Here, we present ratiometric, highly responsive genetically encoded fluorescent indicators, FiNad, for monitoring NAD+ dynamics in living cells and animals. FiNad sensors cover physiologically relevant NAD+ concentrations and sensitively respond to increases and decreases in NAD+. Utilizing FiNad, we performed a head-to-head comparison study of common NAD+ precursors in various organisms and mapped their biochemical roles in enhancing NAD+ levels. Moreover, we showed that increased NAD+ synthesis controls morphofunctional changes of activated macrophages, and directly imaged NAD+ declines during aging in situ. The broad utility of the FiNad sensors will expand our mechanistic understanding of numerous NAD+-associated physiological and pathological processes and facilitate screening for drug or gene candidates that affect uptake, efflux, and metabolism of this important cofactor.

Multicoloured fluorescent indicators for live-cell and in vivo imaging of inorganic mercury dynamics.

Engineered fluorescent indicators for visualizing mercury ion (Hg2+) are powerful tools to illustrate the intracellular distribution and serious toxicity of the ion. However, the sensitive and specific detection of Hg2+ in living cells and in vivo is challenging. This paper reported the development of fluorescent indicators for Hg2+ in green or red color by inserting a circularly permuted fluorescent protein into a highly mercury-specific repressor. These sensors provided a rapid, sensitive, specific, and real-time read-out of Hg2+ dynamics in solutions, bacteria, subcellular organelles of mammalian cells, and zebrafish, thereby providing a useful new method for Hg2+ detection and bioimaging. In conjunction with the hydrogen peroxide sensor HyPer, we found mercury uptake would trigger subcellular oxidative events at the single-cell level, and provided visual evidence of the causality of mercury and oxidative damage. These sensors would paint the landscape of mercury toxicity to cell functions.

Short communication: Improving the activity of bile salt hydrolases in Lactobacillus casei based on in silico molecular docking and heterologous expression.

Bile salt hydrolase (BSH) plays an essential role in the cholesterol-removing effect of lactic acid bacteria, which hydrolyze conjugated bile salts to amino acid and deconjugated bile salts. However, Lactobacillus casei lacks the bsh gene, which may make it highly sensitive to bile salt stress. We wanted to improve the BSH activity of L. casei for various food-industry applications (e.g., milk fermentation). Plate assay testing indicated that Lactobacillus plantarum AR113 has the highest BSH activity. We cloned and sequenced 4 bsh genes from the genome of L. plantarum AR113. Structure modeling and molecular docking of BSH indicated that BSH1 and BSH3 could react efficiently with bile salts, so we selected BSH1 and BSH3 for heterologous expression in L. casei. Compared with single expression of BSH1 or BSH3, co-expression of both protein sequences showed the highest hydrolysis activity by HPLC analysis. Our results suggested that heterologous expression of BSH in L. casei can significantly improve host activity against bile salts, and in silico molecular docking could be an efficient method of rapid screening for BSH with high activity.

Stereoselective synthesis of highly functionalized nitrocyclopropanes through the organocatalyic Michael-addition-initiated cyclization of bromonitromethane and ß,γ-unsaturated α-ketoesters.

A highly diastereo- and enantioselective cyclopropanation of ß,γ-unsaturated α-ketoesters with bromonitromethane has been successfully developed through a domino Michael-addition/intramolecular-alkylation strategy. Acceptable yields (up to 89%) and enantioselectivities (up to 96% ee) have been obtained.