Knowledge-guided data mining on the standardized architecture of NRPS: Subtypes, novel motifs, and sequence entanglements.

Non-ribosomal peptide synthetase (NRPS) is a diverse family of biosynthetic enzymes for the assembly of bioactive peptides. Despite advances in microbial sequencing, the lack of a consistent standard for annotating NRPS domains and modules has made data-driven discoveries challenging. To address this, we introduced a standardized architecture for NRPS, by using known conserved motifs to partition typical domains. This motif-and-intermotif standardization allowed for systematic evaluations of sequence properties from a large number of NRPS pathways, resulting in the most comprehensive cross-kingdom C domain subtype classifications to date, as well as the discovery and experimental validation of novel conserved motifs with functional significance. Furthermore, our coevolution analysis revealed important barriers associated with re-engineering NRPSs and uncovered the entanglement between phylogeny and substrate specificity in NRPS sequences. Our findings provide a comprehensive and statistically insightful analysis of NRPS sequences, opening avenues for future data-driven discoveries.

Knockout of the delta11-desaturase SfruDES1 disrupts sex pheromone biosynthesis, mating and oviposition in the fall armyworm, Spodoptera frugiperda.

Moth insects rely on sex pheromones for long distance attraction and searching for sex partners. The biosynthesis of moth sex pheromones involves the catalytic action of multiple enzymes, with desaturases playing a crucial role in the process of carbon chain desaturation. However, the specific desaturases involved in sex pheromone biosynthesis in fall armyworm (FAW), Spodoptera frugiperda, have not been clarified. In this study, a Δ11 desaturase (SfruDES1) gene in FAW was knocked out using the CRISPR/Cas9 genome editing system. A homozygous mutant of SfruDES1 was obtained through genetic crosses. The gas chromatography-mass spectrometry (GC-MS) analysis results showed that the three main sex pheromone components (Z7-12:Ac, Z9-14:Ac, and Z11-16:Ac) and the three minor components (Z9-14:Ald, E11-14:Ac and Z11-14:Ac) of FAW were not detected in homozygous mutant females compared to the wild type. Furthermore, behavioral assay demonstrated that the loss of SfruDES1 resulted in a significant reduction in the attractiveness of females to males, along with disruptions in mating behavior and oviposition. Additionally, in a heterologous expression system, recombinant SfruDES1 could introduce a cis double bond at the Δ11 position in palmitic acid, which resulted in the changes in components of the synthesized products. These findings suggest desaturase plays a key role in the biosynthesis of sex pheromones, and knockout of the SfruDES1 disrupts sex pheromone biosynthesis and mating behavior in FAW. The SfruDES1 could serve as tool to develop a control method for S. frugiperda.

Core Microbiota in the Rhizosphere of Heavy Metal Accumulators and Its Contribution to Plant Performance.

Persistent microbial symbioses can confer greater fitness to their host under unfavorable conditions, but manipulating such beneficial interactions necessitates a mechanistic understanding of the consistently important microbiomes for the plant. Here, we examined the phylogenetic profiles and plant-beneficial traits of the core microbiota that consistently inhabits the rhizosphere of four divergent Cd hyperaccumulators and an accumulator. We evidenced the existence of a conserved core rhizosphere microbiota in each plant distinct from that in the non-hyperaccumulating plant. Members of Burkholderiaceae and Sphingomonas were the shared cores across hyperaccumulators and accumulators. Several keystone taxa in the rhizosphere networks were part of the core microbiota, the abundance of which was an important predictor of plant Cd accumulation. Furthermore, an inoculation experiment with synthetic communities comprising isolates belonging to the shared cores indicated that core microorganisms could facilitate plant growth and metal tolerance. Using RNA-based stable isotope probing, we discovered that abundant core taxa overlapped with active rhizobacteria utilizing root exudates, implying that the core rhizosphere microbiota assimilating plant-derived carbon may provide benefits to plant growth and host phenotype such as Cd accumulation. Our study suggests common principles underpinning hyperaccumulator-microbiome interactions, where plants consistently interact with a core set of microbes contributing to host fitness and plant performance. These findings lay the foundation for harnessing the persistent root microbiomes to accelerate the restoration of metal-disturbed soils.

Plant volatile compound methyl benzoate is highly effective against Spodoptera frugiperda and safe to non-target organisms as an eco-friendly botanical-insecticide.

Recent studies have indicated that the plant volatile methyl benzoate (MB) exhibits significant insecticidal bioactivity against several common insects. However, the potential environmental hazards of MB and its safety to non-target organisms is poorly understood. In the present study, these characteristics were investigated through laboratory experiments and field investigations. The results revealed that MB was highly toxic to the agricultural pest, fall armyworm Spodoptera frugiperda. Compared with the commercial pesticide lambda-cyhalothrin, the toxicities of MB against S. frugiperda larvae and adults were comparable and 3.41 times higher, respectively. Behavioral bioassays showed that the percentage repellency of MB to S. frugiperda larvae was 56.72 %, and MB induced 69.40 % oviposition deterrence rate in S. frugiperda female adults. Furthermore, in terms of median lethal concentration (LC50) and median lethal doses (LD50), MB exhibited non-toxic effects on non-target animals with 3-d LC50 of > 1 % to natural predators (Coccinella septempunctata and Harmonia axyridis), 3-d LD50 of 467.86 µg/bee to the bumblebee Bombus terrestris, 14-d LC50 of 971.09 mg/kg to the earthworm Eisenia fetida, and 4-d LC50 of 47.30 mg/L to the zebrafish Brachydanio rerio. The accumulation of MB in the soil and earthworms was found to be extremely limited. Our comparative study clearly demonstrated that MB is effective as a selective botanical pesticide against S. frugiperda and it is safe to use in the tested environment, with no toxic effects on non-target animals and natural predators.

Integrative analysis of AR-mediated transcriptional regulatory network reveals IRF1 as an inhibitor of prostate cancer progression.

BACKGROUND: Androgen receptor (AR) is crucial for prostate cancer (PCa) initiation and malignant progression. Only half of androgen-responsive genes have been identified as having androgen-responsive elements, suggesting that AR regulates downstream genes through other transcriptional factors. However, whether and how AR regulates the progression via regulating these androgen-responsive genes remains unclear. METHODS: Androgen-responsive and activity-changed (AC) transcriptional factors (TFs) were identified based on the time-course gene-expression array and gene promoter regions analysis. The intersection of androgen-responsive and AC TFs was selected the core TFs, which were used to construct the core transcriptional regulatory network. GO enrichment analysis, cell proliferation assays, glycolysis experiments, and reverse transcription polymerase chain reaction analysis were used to analyze and validate the functions of the network. As one of the core TFs, the function and mechanism of IRF1 have been further explored. RESULTS: We devised a new integrated approach to select core TFs and construct core transcriptional regulatory network in PCa. The 24 core TFs and core transcriptional regulatory network participate in regulating PCa cell proliferation, RNA splicing, and cancer metabolism. Further validations showed that AR signaling could promote glycolysis via inducing glycolytic enzymes in PCa cells. IRF1, a novel target of AR, served as a tumor suppressor by inhibiting PCa proliferation, cell cycle, and glycolysis. CONCLUSIONS: It is the first time to demonstrate the regulating role of the AR-mediated transcriptional regulatory network in a series of important biological processes in PCa cells. IRF1, an AR-regulated TF, acts as tumor suppressor in this core transcriptional regulatory network, which highlights the therapeutic potential of targeting this regulatory network for PCa.

Associations between ambient air pollution and daily incidence of pediatric hand, foot and mouth disease in Ningbo, 2014-2016: a distributed lag nonlinear model.

Hand, foot and mouth disease (HFMD) has high prevalence around the world, with serious consequences for children. Due to the long survival period of HFMD virus in ambient air, air pollutants may play a critical role in HFMD epidemics. We collected data on daily cases of HFMD among children aged 0-14 years in Ningbo City between 2014 and 2016. Distributed lag nonlinear models were used to assess the effects of particulate matter (PM2.5), sulphur dioxide (SO2), nitrogen dioxide (NO2) and ozone (O3) on the daily incidence of HFMD among children, with analyses stratified by gender and age. Compared with moderate levels of air pollution, high SO2 levels had a relative risk (RR) of 2.32 (95% CI 1.42-3.79) and high NO2 levels had a RR of 2.01 (95% CI 1.22-3.31). The RR of O3 was 2.12 (95% CI 1.47-3.05) and that of PM2.5 was 0.77 (95% CI 0.64-0.92) at moderate levels of air pollution. Specifically, high levels of SO2 and NO2 had RRs of 2.39 (95% CI 1.44-3.96) and 2.02 (95% CI 1.21-3.39), respectively, among 0-4-year-old children, while high O3 had an RR of 2.31 (95% CI 1.09-4.89) among 5-14-year-old children. Our findings suggest significant associations of high SO2 and NO2 levels and moderate O3 levels in HFMD epidemics, and also indicate that air pollution causes lagged effects on HFMD epidemics. Our study provides practical and useful data for targeted prevention and control of HMFD based on environmental evidence.

A long non-coding RNA regulates cadherin transcription and susceptibility to Bt toxin Cry1Ac in pink bollworm, Pectinophora gossypiella.

Extensive planting of transgenic crops producing insecticidal proteins from the bacterium Bacillus thuringiensis (Bt) has spurred increasingly rapid evolution of resistance in pests. In the pink bollworm, Pectinophora gossypiella, a devastating global pest, resistance to Bt toxin Cry1Ac produced by transgenic cotton is linked with mutations in a gene (PgCad1) encoding a cadherin protein that binds Cry1Ac in the larval midgut. We previously reported a long non-coding RNA (lncRNA) in intron 20 of cadherin alleles associated with both resistance and susceptibility to Cry1Ac. Here we tested the hypothesis that reducing expression of this lncRNA decreases transcription of PgCad1 and susceptibility to Cry1Ac. Quantitative RT-PCR showed that feeding susceptible neonates small interfering RNAs (siRNAs) targeting this lncRNA but not PgCad1 decreased the abundance of transcripts of both the lncRNA and PgCad1. Moreover, neonates fed the siRNAs had lower susceptibility to Cry1Ac. The results imply that the lncRNA increases transcription of PgCad1 and susceptibility of pink bollworm to Cry1Ac. The results suggest that disruption of lncRNA expression could be a novel mechanism of pest resistance to Bt toxins.

Resource stoichiometry shapes community invasion resistance via productivity-mediated species identity effects.

Diversity-invasion resistance relationships are often variable and sensitive to environmental conditions such as resource availability. Resource stoichiometry, the relative concentration of different elements in the environment, has been shown to have strong effects on the physiology and interactions between different species. Yet, its role for diversity-invasion resistance relationships is still poorly understood. Here, we explored how the ratio of nitrogen (N) and phosphorus affects the productivity and invasion resistance of constructed microbial communities by a plant pathogenic bacterium, Ralstonia solanacearum. We found that resource stoichiometry and species identity effects affected the invasion resistance of communities. Both high N concentration and resident community diversity constrained invasions, and two resident species, in particular, had strong negative effects on the relative density of the invader and the resident community productivity. While resource stoichiometry did not affect the mean productivity of the resident community, it favoured the growth of two species that strongly constrained invasions turning the slope of productivity-invasion resistance relationship more negative. Together our findings suggest that alterations in resource stoichiometry can change the community resistance to invasions by having disproportionate effects on species growth, potentially explaining changes in microbial community composition under eutrophication.

The terpene synthase gene family in Gossypium hirsutum harbors a linalool synthase GhTPS12 implicated in direct defence responses against herbivores.

Herbivore-induced terpenes have been reported to function as ecological signals in plant-insect interactions. Here, we showed that insect-induced cotton volatile blends contained 16 terpenoid compounds with a relatively high level of linalool. The high diversity of terpene production is derived from a large terpene synthase (TPS) gene family. The TPS gene family of Gossypium hirsutum and Gossypium raimondii consist of 46 and 41 members, respectively. Twelve TPS genes (GhTPS4-15) could be isolated, and protein expression in Escherichia coli revealed catalytic activity for eight GhTPS. The upregulation of the majority of these eight genes additionally supports the function of these genes in herbivore-induced volatile biosynthesis. Furthermore, transgenic Nicotiana tabacum plants overexpressing GhTPS12 were generated, which produced relatively large amounts of (3S)-linalool. In choice tests, female adults of Helicoverpa armigera laid fewer eggs on transgenic plants compared with non-transformed controls. Meanwhile, Myzus persicae preferred feeding on wild-type leaves over leaves of transgenic plants. Our findings demonstrate that transcript accumulation of multiple TPS genes is mainly responsible for the production and diversity of herbivore-induced volatile terpenes in cotton. Also, these genes might play roles in plant defence, in particular, direct defence responses against herbivores.

Useful Bicistronic Reporter System for Studying Poly(A) Site-Defining cis Elements and Regulation of Alternative Polyadenylation.

The link between polyadenylation (pA) and various biological, behavioral, and pathological events of eukaryotes underlines the need to develop in vivo polyadenylation assay methods for characterization of the cis-acting elements, trans-acting factors and environmental stimuli that affect polyadenylation efficiency and/or relative usage of two alternative polyadenylation (APA) sites. The current protein-based CAT or luciferase reporter systems can measure the polyadenylation efficiency of a single pA site or candidate cis element but not the choice of two APA sites. To address this issue, we developed a set of four new bicistronic reporter vectors that harbor either two luciferase or fluorescence protein open reading frames connected with one Internal Ribosome Entry Site (IRES). Transfection of single or dual insertion constructs of these vectors into mammalian cells demonstrated that they could be utilized not only to quantify the strength of a single candidate pA site or cis element, but also to accurately measure the relative usage of two APA sites at both the mRNA (qRT-PCR) and protein levels. This represents the first reporter system that can study polyadenylation efficiency of a single pA site or element and regulation of two APA sites at both the mRNA and protein levels.

RNAi-Induced Electrophysiological and Behavioral Changes Reveal two Pheromone Binding Proteins of Helicoverpa armigera Involved in the Perception of the Main Sex Pheromone Component Z11-16:Ald.

Pheromone binding proteins (PBPs) are thought to play key roles in insect sex pheromone recognition; however, there is little in vivo evidence to support this viewpoint in comparison to abundant biochemical data in vitro. In the present study, two noctuid PBP genes HarmPBP1 and HarmPBP2 of the serious agricultural pest, Helicoverpa armigera were selected to be knocked down by RNA interference, and then the changes in electrophysiological and behavioral responses of male mutants to their major sex pheromone component (Z)-11-hexadecenal (Z11-16:Ald) were recorded. There were no significant electrophysiological or behavioral changes of tested male moths in response to Z11-16:Ald when either single PBP gene was knocked down. However, decreased sensitivity of male moths in response to Z11-16:Ald was observed when both HarmPBP1 and HarmPBP2 genes were silenced. These results reveal that both HarmPBP1 and HarmPBP2 are required for the recognition of the main sex pheromone component Z11-16:Ald in H. armigera. Furthermore, these findings may help clarify physiological roles of moth PBPs in the sex pheromone recognition pathway, which in turn could facilitate pest control by exploring sex pheromone blocking agents.

Aminopeptidase N5 (APN5) as a Putative Functional Receptor of Cry1Ac Toxin in the Larvae of Athetis lepigone.

Athetis lepigone was a new lepidopteran pest and caused severe damage to maize crops in China. We have detected that Cry1Ac protoxin and toxin were highly active against the larvae of A. lepigone. However, there is no report about the mode of action of Bt Cry1Ac toxin against this pest until now. A 110 kDa APN5 protein from BBMV of A. lepigone was identified as the binding receptor of Cry1Ac toxin using Ligand blotting. The Cry1Ac receptor APN5 was cloned from A. lepigone larval midgut mRNA and named as AlAPN5 (GenBank accession no.: KU950745). AlAPN5 had a GATEN motif and been classified to Class 5 APNs. 79.2% reduction in mortality was observed when A. lepigone larvae were injected with siRNA of the AlAPN5 gene and treated with Cry1Ac toxin. These data demonstrate that AlAPN5 is a putative functional receptor and maybe the only receptor of Cry1Ac in A. lepigone.

Investigation of TtrD, an expressing recombinant fusion tag, in Escherichia coli.

Escherichia coli is widely used for expressing recombinant proteins, and several tags have been developed to improve protein solubility. However, expressing and purifying protein from other organisms is not always successful. In this study, we investigated the possibility of using TtrD as an expressing fusion tag in E. coli. Twenty RING finger domain containing human genes were expressed in E. coli grown at 37 °C and 18 °C and tested with four other fusion tags, namely His, SUMO, GST and MBP, for comparison. The results indicated that the soluble expressing ability of the tags was MBP, GST, TtrD, SUMO, and His in descending order. A one-column refolding process was used to purify the expressed proteins in inclusion bodies, and TtrD showed the strongest refolding ability. The results suggested that the TtrD tag enhanced recombinant protein solubility and refolding ability and might be a useful tag for protein expression in E. coli.

Heat-related illness in China, summer of 2013.

Extreme heat events have occurred more frequently in China in recent years, leading to serious impacts on human life and the health care system. To identify the characteristics of individuals with heat-related illnesses in China during the summer of 2013, we collected the data from the Heat-related Illness Surveillance System in Chinese Center for Disease Control and Prevention (China CDC). A total of 5758 cases were reported in the summer of 2013, mostly concentrated in urban areas around the middle and lower reaches of the Yangtze River. We found a difference in age distribution of percentage of deaths from heat-related illness between males and females. Severe cases in males mostly occurred in the age group 45-74 years but in females mostly in the age group over 75. A distributed lag non-linear model had been used to identify population vulnerabilities in Ningbo and Chongqing. The results show that there was a clear positive relationship between maximum temperature and heat-related illness, and the heat effect was nonlinear and could last for 3 days. The elderly and males in the range of 45-64 years old might be the most vulnerable people of heat-related illness in China. We also highlighted some deficiencies of the surveillance system, such that the reported data were not accurate, comprehensive, or timely enough at this stage.

Association of CDC25 phosphatase family polymorphisms with the efficacy/toxicity of platinum-based chemotherapy in Chinese advanced NSCLC patients.

AIM: To explore relationships between single nucleotide polymorphisms (SNPs) of the CDC25 protein family and the survival and chemotherapy responses of patients with advanced non-small-cell lung cancer (NSCLC). METHODS & MATERIALS: We genotyped 14 SNPs of the CDC25 family in 663 Chinese patients with advanced NSCLC who were treated with first-line platinum-based chemotherapy and, in evaluable patients, analyzed relationships between the CDC25 family and the efficacy of platinum-based chemotherapy. RESULTS: CDC25A rs3731513 and rs1380053, CDC25C rs6861656, CDC25A haplotype T/A/A/A/C and CDC25C haplotype A/G/G/G/C were significantly associated with the patients' progression-free survival. In addition, CDC25B rs3761218 and haplotype G/T/G/G were associated with the occurrence of severe toxicity with platinum-based chemotherapy, especially gastrointestinal and hematological toxicity. CONCLUSION: These findings reveal a relationship between genetic variations of the CDC25 family and the efficacy and toxicity of platinum-based chemotherapy in patients with advanced NSCLC, especially in those with non-squamous-cell carcinoma.

The effects of summer temperature and heat waves on heat-related illness in a coastal city of China, 2011-2013.

BACKGROUND: Devastating health effects from recent heat waves in China have highlighted the importance of understanding health consequences from extreme heat stress. Despite the increasing mortality from extreme heat, very limited studies have quantified the effects of summer extreme temperature on heat-related illnesses in China. METHODS: The associations between extreme heat and daily heat-related illnesses that occurred in the summers of 2011-2013 in Ningbo, China, have been examined, using a distributed lag non-linear model (DLNM) based on 3862 cases. The excess morbidities of heat-related illness during each heat wave have been calculated separately and the cumulative heat wave effects on age-, sex-, and cause-specific illnesses in each year along lags have been estimated as well. RESULTS: After controlling the effect of relative humidity, it is found that maximum temperature, rather than heat index, was a better predictor of heat-related illnesses in summers. A positive association between maximum temperatures and occurrence of heat-related diseases was apparent, especially at short lag effects. Six heat waves during the period of 2011-2013 were identified and all associated with excess heat-related illnesses. Relative to the average values for the corresponding periods in 2011 and 2012, a total estimated 679 extra heat-related illnesses occurred during three heat waves in 2013. The significant prolonged heat wave effects on total heat-related illnesses during heat waves in three study years have also been identified. The strongest cumulative effect of heat waves was on severe heat diseases in 2013, with a 10-fold increased risk. More males than females, individuals with more severe forms of illness, were more affected by the heat. However, all age groups were vulnerable. CONCLUSIONS: Recent heat waves had a substantial and delayed effect on heat illnesses in Ningbo. Relevant active well-organized public health initiatives should be implemented to reduce the adverse effects of heat extremes on the illnesses.

Keap1 Negatively Regulates Transcription of Three Counter-Defense Genes and Susceptibility to Plant Toxin Gossypol in Helicoverpa armigera.

Expressions of a wide range of cytoprotective counter-defense genes are mainly regulated by the Keap1-Nrf2-ARE signaling pathway in response to oxidative stress from xenobiotics. Gossypol is the major antiherbivore secondary metabolite of cotton, but how the polyphagous pest Helicoverpa armigera copes with this phytochemical to utilize its favorite host plant cotton remains largely elusive. In this study, we first suppressed the Keap1 gene in newly hatched larvae of cotton bollworm by feeding them the siRNA diet for 4 days. All of the larvae were subsequently fed the artificial diet supplied with gossypol or the control diet for 5 days. We identified that the knockdown of the Keap1 gene significantly decreased larval mortality and significantly increased the percentages of larval survival, reaching the fourth instar, compared with ncsiRNA when exposed to a diet containing gossypol. Three counter-defense genes CYP9A17, CYP4L11 and UGT41B3, which were related to the induction or metabolism of gossypol according to the report before, were all significantly up-regulated after the knockdown of the Keap1 gene. The Antioxidant Response Elements (AREs) were also detected in the promoter regions of the three counter-defense genes above. These data indicate that the suppression of the Keap1 gene activates the Keap1-Nrf2-ARE signaling pathway, up-regulates the expressions of counter-defense genes involved in the resistance of oxidative stress and finally contributes to reducing the susceptibility of gossypol. Our results provide more knowledge about the transcriptional regulation mechanisms of counter-defense genes that enable the cotton bollworm to adapt to the diversity of host plants including cotton.

Two cuticle-enriched chemosensory proteins confer multi-insecticide resistance in Spodoptera frugiperda.

Recent studies revealed that insect chemosensory proteins (CSPs) both play essential roles in insect olfaction and insect resistance. However, functional evidence supporting the crosslink between CSP and insecticide resistance remains unexplored. In the present study, 22 SfruCSP transcripts were identified from the fall armyworm (FAW) and SfruCSP1 and SfruCSP2 are enriched in the larval cuticle and could be induced by multiple insecticides. Both SfruCSP1 and SfruCSP2 are highly expressed in the larval inner endocuticle and outer epicuticle, and these two proteins exhibited high binding affinities with three insecticides (chlorfenapyr, chlorpyrifos and indoxacarb). The knockdown of SfruCSP1 and SfruCSP2 increased the susceptibility of FAW larvae to the above three insecticides, and significantly increased the penetration ratios of these insecticides. Our in vitro and in vivo evidence suggests that SfruCSP1 and SfruCSP2 are insecticide binding proteins and confer FAW larval resistance to chlorfenapyr, chlorpyrifos and indoxacarb by an insecticide sequestration mechanism. The study should aid in the exploration of larval cuticle-enriched CSPs for insect resistance management.

SIDERITE: Unveiling hidden siderophore diversity in the chemical space through digital exploration.

In this work, we introduced a siderophore information database (SIDERTE), a digitized siderophore information database containing 649 unique structures. Leveraging this digitalized data set, we gained a systematic overview of siderophores by their clustering patterns in the chemical space. Building upon this, we developed a functional group-based method for predicting new iron-binding molecules with experimental validation. Expanding our approach to the collection of open natural products (COCONUT) database, we predicted a staggering 3199 siderophore candidates, showcasing remarkable structure diversity that is largely unexplored. Our study provides a valuable resource for accelerating the discovery of novel iron-binding molecules and advancing our understanding of siderophores.

The Microbiome Protocols eBook initiative: Building a bridge to microbiome research.

The Microbiome Protocols eBook (MPB) serves as a crucial bridge, filling gaps in microbiome protocols for both wet experiments and data analysis. The first edition, launched in 2020, featured 152 meticulously curated protocols, garnering widespread acclaim. We now extend a sincere invitation to researchers to participate in the upcoming 2nd version of MPB, contributing their valuable protocols to advance microbiome research.