Effect of smoking-related features and 731 immune cell phenotypes on esophageal cancer: a two-sample and mediated Mendelian randomized study.

Introduction: Numerous observational studies have indicated that smoking is a substantial risk factor for esophageal cancer. However, there is a shortage of research that delves into the specific causal relationship and potential mediators between the two. Our study aims to validate the correlation between smoking-related traits and esophageal cancer while exploring the possible mediating effects of immune factors. Methods: Initially, we conducted bidirectional univariate Mendelian Randomization (MR) analyses to forecast the causal effects linking smoking-related traits and esophageal cancer. Subsequently, we employed a two-step MR analysis to scrutinize immune cell phenotypes that could mediate these effects. Finally, the coefficient product method was employed to determine the precise mediating impact. Additionally, we have refined our sensitivity analysis to ensure the reliability of the outcomes. Results: After analysis, Smoking status: Never had a significant negative association with the incidence of esophageal cancer (inverse-variance weighted (IVW) method, p=1.82e-05, OR=0.10, 95%CI=0.04~0.29). Ever smoked (IVW, p=1.49e-02, OR=4.31, 95%CI=1.33~13.94) and Current tobacco smoking (IVW, p=1.49e-02, OR=4.31, 95%CI=1.33~13.94) showed the promoting effect on the pathogenesis of esophageal cancer. Through further examination, researchers discovered 21 immune cell phenotypes that have a causal relationship with esophageal cancer. After careful screening, two immune cell phenotypes were found to have potential mediating effects. In particular, it was observed that in the case of the preventive effect of Smoking status: Never on esophageal cancer, the absolute count of CD62L plasmacytoid dendritic cells mediated a reduction of 4.21%, while the mediating effect of CD27 in CD20-CD38-B cells was -4.12%. In addition, sensitivity analyses did not reveal significant heterogeneity or level pleiotropy. Conclusion: The study provides new evidence for the causal relationship between smoking-related features and esophageal cancer and proposes immune factors with potential mediating effects. However, this finding needs to be further demonstrated by more extensive clinical studies.

Artificial evolution of OsEPSPS through an improved dual cytosine and adenine base editor generated a novel allele conferring rice glyphosate tolerance.

Exploiting novel endogenous glyphosate-tolerant alleles is highly desirable and has promising potential for weed control in rice breeding. Here, through fusions of different effective cytosine and adenine deaminases with nCas9-NG, we engineered an effective surrogate two-component composite base editing system, STCBE-2, with improved C-to-T and A-to-G base editing efficiency and expanded the editing window. Furthermore, we targeted a rice endogenous OsEPSPS gene for artificial evolution through STCBE-2-mediated near-saturated mutagenesis. After hygromycin and glyphosate selection, we identified a novel OsEPSPS allele with an Asp-213-Asn (D213N) mutation (OsEPSPS-D213N) in the predicted glyphosate-binding domain, which conferred rice plants reliable glyphosate tolerance and had not been reported or applied in rice breeding. Collectively, we developed a novel dual base editor which will be valuable for artificial evolution of important genes in crops. And the novel glyphosate-tolerant rice germplasm generated in this study will benefit weeds management in rice paddy fields.

Pre-operative prognostic nutrition index and post-operative pneumonia in aneurysmal subarachnoid hemorrhage patients.

Background and objective: Post-operative pneumonia (POP), a common complication, may be associated with prolonged hospitalization and long-term mortality in aneurysmal subarachnoid hemorrhage (aSAH) patients. This study aimed to explore the association between pre-operative prognostic nutrition index (PNI) and POP in aSAH patients. Methods: A total of 280 aSAH patients were enrolled from Nanjing Drum Tower Hospital. PNI was calculated as follows: [10 × albumin(gr/dl)] + [0.005 × absolute pre-operative lymphocyte count (per mm3)]. We utilized multivariate analyses, restricted cubic spline, net reclassification improvement (NRI), and integrated discrimination improvement (IDI) to elucidate the role of PNI in POP. Results: Pre-operative PNI levels in the POP group were higher, compared with the non-POP group (41.0 [39.0, 45.4] vs. 44.4 [40.5, 47.3], P = 0.001). When we included PNI as a categorical variable in the multivariate analysis, the levels of PNI were associated with POP (odds ratio, 0.433; 95% confidence interval, 0.253-0.743; P=0.002). In addition, when we included PNI as a continuous variable in the multivariate analysis, the PNI levels were also associated with POP (odds ratio, 0.942; 95% confidence interval, 0.892-0.994; P = 0.028). The level of albumin was also a predictor of the occurrence of POP, with a lower diagnostic power than PNI [AUC: 0.611 (95% confidence interval, 0.549-0.682; P = 0.001) for PNI vs. 0.584 (95% confidence interval, 0.517-0.650; P = 0.017) for albumin]. Multivariable-adjusted spline regression indicated a linear dose-response association between PNI and POP in aSAH participants (P for linearity = 0.027; P for non-linearity = 0.130). Reclassification assessed by IDI and NRI was significantly improved with the addition of PNI to the conventional model of POP in aSAH patients (NRI: 0.322 [0.089-0.555], P = 0.007; IDI: 0.016 [0.001-0.031], P = 0.040). Conclusion: The lower levels of pre-operative PNI may be associated with the higher incidence of POP in aSAH patients. Neurosurgeons are supposed to pay more attention to pre-operative nutrition status in aSAH patients.

Plant base editing and prime editing: The current status and future perspectives.

Precise replacement of an allele with an elite allele controlling an important agronomic trait in a predefined manner by gene editing technologies is highly desirable in crop improvement. Base editing and prime editing are two newly developed precision gene editing systems which can introduce the substitution of a single base and install the desired short indels to the target loci in the absence of double-strand breaks and donor repair templates, respectively. Since their discoveries, various strategies have been attempted to optimize both base editor (BE) and prime editor (PE) in order to improve the precise editing efficacy, specificity, and expand the targeting scopes. Here, we summarize the latest development of various BEs and PEs, as well as their applications in plants. Based on these progresses, we recommend the appropriate BEs and PEs for both basic plant research and crop improvement. Moreover, we propose the perspectives for further optimization of these two editors. We envision that both BEs and PEs will become the routine and customized precise gene editing tools for both plant biological research and crop improvement in the near future.

A VQ-motif-containing protein fine-tunes rice immunity and growth by a hierarchical regulatory mechanism.

Rice blast and bacterial blight, caused by the fungus Magnaporthe oryzae and the bacterium Xanthomonas oryzae pv. oryzae (Xoo), respectively, are devastating diseases affecting rice. Here, we report that a rice valine-glutamine (VQ) motif-containing protein, OsVQ25, balances broad-spectrum disease resistance and plant growth by interacting with a U-Box E3 ligase, OsPUB73, and a transcription factor, OsWRKY53. We show that OsPUB73 positively regulates rice resistance against M. oryzae and Xoo by interacting with and promoting OsVQ25 degradation via the 26S proteasome pathway. Knockout mutants of OsVQ25 exhibit enhanced resistance to both pathogens without a growth penalty. Furthermore, OsVQ25 interacts with and suppresses the transcriptional activity of OsWRKY53, a positive regulator of plant immunity. OsWRKY53 downstream defense-related genes and brassinosteroid signaling genes are upregulated in osvq25 mutants. Our findings reveal a ubiquitin E3 ligase-VQ protein-transcription factor module that fine-tunes plant immunity and growth at the transcriptional and posttranslational levels.

An update on precision genome editing by homology-directed repair in plants.

Beneficial alleles derived from local landraces or related species, or even orthologs from other plant species, are often caused by differences of one or several single-nucleotide polymorphisms or indels in either the promoter region or the encoding region of a gene and often account for major differences in agriculturally important traits. Clustered regularly interspaced short palindromic repeats-associated endonuclease Cas9 system (CRISPR/Cas9)-mediated precision genome editing enables targeted allele replacement or insertion of flag or foreign genes at specific loci via homology-directed repair (HDR); however, HDR efficiency is low due to the intrinsic rare occurrence of HDR and insufficient DNA repair template in the proximity of a double-stranded break (DSB). Precise replacement of the targeted gene with elite alleles from landraces or relatives into a commercial variety through genome editing has been a holy grail in the crop genome editing field. In this update, we briefly summarize CRISPR/Cas-mediated HDR in plants. We describe diverse strategies to improve HDR efficiency by manipulating the DNA repair pathway, timing DSB induction, and donor delivery, and so on. Lastly, we outline open questions and challenges in HDR-mediated precision genome editing in both plant biological research and crop improvement.

Present and future prospects for wheat improvement through genome editing and advanced technologies.

Wheat (Triticum aestivum, 2n = 6x = 42, AABBDD) is one of the most important staple food crops in the world. Despite the fact that wheat production has significantly increased over the past decades, future wheat production will face unprecedented challenges from global climate change, increasing world population, and water shortages in arid and semi-arid lands. Furthermore, excessive applications of diverse fertilizers and pesticides are exacerbating environmental pollution and ecological deterioration. To ensure global food and ecosystem security, it is essential to enhance the resilience of wheat production while minimizing environmental pollution through the use of cutting-edge technologies. However, the hexaploid genome and gene redundancy complicate advances in genetic research and precision gene modifications for wheat improvement, thus impeding the breeding of elite wheat cultivars. In this review, we first introduce state-of-the-art genome-editing technologies in crop plants, especially wheat, for both functional genomics and genetic improvement. We then outline applications of other technologies, such as GWAS, high-throughput genotyping and phenotyping, speed breeding, and synthetic biology, in wheat. Finally, we discuss existing challenges in wheat genome editing and future prospects for precision gene modifications using advanced genome-editing technologies. We conclude that the combination of genome editing and other molecular breeding strategies will greatly facilitate genetic improvement of wheat for sustainable global production.

Increasing yield potential through manipulating of an ARE1 ortholog related to nitrogen use efficiency in wheat by CRISPR/Cas9.

Wheat (Triticum aestivum L.) is a staple food crop consumed by more than 30% of world population. Nitrogen (N) fertilizer has been applied broadly in agriculture practice to improve wheat yield to meet the growing demands for food production. However, undue N fertilizer application and the low N use efficiency (NUE) of modern wheat varieties are aggravating environmental pollution and ecological deterioration. Under nitrogen-limiting conditions, the rice (Oryza sativa) abnormal cytokinin response1 repressor1 (are1) mutant exhibits increased NUE, delayed senescence and consequently, increased grain yield. However, the function of ARE1 ortholog in wheat remains unknown. Here, we isolated and characterized three TaARE1 homoeologs from the elite Chinese winter wheat cultivar ZhengMai 7698. We then used CRISPR/Cas9-mediated targeted mutagenesis to generate a series of transgene-free mutant lines either with partial or triple-null taare1 alleles. All transgene-free mutant lines showed enhanced tolerance to N starvation, and showed delayed senescence and increased grain yield in field conditions. In particular, the AABBdd and aabbDD mutant lines exhibited delayed senescence and significantly increased grain yield without growth defects compared to the wild-type control. Together, our results underscore the potential to manipulate ARE1 orthologs through gene editing for breeding of high-yield wheat as well as other cereal crops with improved NUE.

Regulation of biochar mediated catalytic degradation of quinolone antibiotics: Important role of environmentally persistent free radicals.

Antibiotic pollution threatens aquatic ecosystems and water supplies, so analysis of ecofriendly remediation approaches like biochars with catalytic degradation abilities is a top priority. In this work, quinolone antibiotics were degraded by activating oxidants to generate transient radicals using the environmentally persistent free radicals (EPFRs) carried by biochar. The physical and chemical characterization confirmed that biochar is suitable for the removal of organic pollutants. By regulating biochar preparation parameters, it was found that EPFR generation peaked at 500 °C. As the temperature increased from 300 °C to 500 °C, the EPFRs changed from oxygen-centered radicals (g > 2.0040) to carbon-centered radicals (g < 2.0030). The catalytic degradation efficiencies of the EPFR activated oxidants from large to small were: peroxydisulfate (PDS), peroxymonosulfate (PMS), H2O2 and flowing O2. The combined actions of SO4•- and •OH effectively degraded antibiotics. The results showed that biochar activating persulfate is a promising technique for the degradation of antibiotics.

Precise gene replacement in plants through CRISPR/Cas genome editing technology: current status and future perspectives.

CRISPR/Cas, as a simple, versatile, robust and cost-effective system for genome manipulation, has dominated the genome editing field over the past few years. The application of CRISPR/Cas in crop improvement is particularly important in the context of global climate change, as well as diverse agricultural, environmental and ecological challenges. Various CRISPR/Cas toolboxes have been developed and allow for targeted mutagenesis at specific genome loci, transcriptome regulation and epigenome editing, base editing, and precise targeted gene/allele replacement or tagging in plants. In particular, precise replacement of an existing allele with an elite allele in a commercial variety through homology-directed repair (HDR) is a holy grail in genome editing for crop improvement as it has been very difficult, laborious and time-consuming to introgress the elite alleles into commercial varieties without any linkage drag from parental lines within a few generations in crop breeding practice. However, it still remains very challenging in crop plants. This review intends to provide an informative summary of the latest development and breakthroughs in gene replacement using CRISPR/Cas technology, with a focus on achievements, potential mechanisms and future perspectives in plant biological science as well as crop improvement.

A barley stripe mosaic virus-based guide RNA delivery system for targeted mutagenesis in wheat and maize.

Plant RNA virus-based guide RNA (gRNA) delivery has substantial advantages compared to that of the conventional constitutive promoter-driven expression due to the rapid and robust amplification of gRNAs during virus replication and movement. To date, virus-induced genome editing tools have not been developed for wheat and maize. In this study, we engineered a barley stripe mosaic virus (BSMV)-based gRNA delivery system for clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9-mediated targeted mutagenesis in wheat and maize. BSMV-based delivery of single gRNAs for targeted mutagenesis was first validated in Nicotiana benthamiana. To extend this work, we transformed wheat and maize with the Cas9 nuclease gene and selected the wheat TaGASR7 and maize ZmTMS5 genes as targets to assess the feasibility and efficiency of BSMV-mediated mutagenesis. Positive targeted mutagenesis of the TaGASR7 and ZmTMS5 genes was achieved for wheat and maize with efficiencies of up to 78% and 48%. Our results provide a useful tool for fast and efficient delivery of gRNAs into economically important crops.

Precise gene replacement in rice by RNA transcript-templated homologous recombination.

One of the main obstacles to gene replacement in plants is efficient delivery of a donor repair template (DRT) into the nucleus for homology-directed DNA repair (HDR) of double-stranded DNA breaks. Production of RNA templates in vivo for transcript-templated HDR (TT-HDR) could overcome this problem, but primary transcripts are often processed and transported to the cytosol, rendering them unavailable for HDR. We show that coupling CRISPR-Cpf1 (CRISPR from Prevotella and Francisella 1) to a CRISPR RNA (crRNA) array flanked with ribozymes, along with a DRT flanked with either ribozymes or crRNA targets, produces primary transcripts that self-process to release the crRNAs and DRT inside the nucleus. We replaced the rice acetolactate synthase gene (ALS) with a mutated version using a DNA-free ribonucleoprotein complex that contains the recombinant Cpf1, crRNAs, and DRT transcripts. We also produced stable lines with two desired mutations in the ALS gene using TT-HDR.

Synthesis-dependent repair of Cpf1-induced double strand DNA breaks enables targeted gene replacement in rice.

The recently developed CRISPR (clustered regularly interspaced short palindromic repeats)/Cpf1 system expands the range of genome editing and is emerging as an alternative powerful tool for both plant functional genomics and crop improvement. Cpf1-CRISPR RNA (crRNA) produces double strand DNA breaks (DSBs) with long 5'-protruding ends, which may facilitate the pairing and insertion of repair templates through homology-directed repair (HDR) for targeted gene replacement and introduction of the desired DNA elements at specific gene loci for crop improvement. However, the potential mechanism underlying HDR of DSBs generated by Cpf1-crRNA remains to be investigated, and the inherent low efficiency of HDR and poor availability of exogenous donor DNA as repair templates strongly impede the use of HDR for precise genome editing in crop plants. Here, we provide evidence of synthesis-dependent repair of Cpf1-induced DSBs, which enables us precisely to replace the wild-type ALS gene with the intended mutant version that carries two discrete point mutations conferring herbicide resistance to rice plants. Our observation that the donor repair template (DRT) with only the left homologous arm is sufficient for precise targeted allele replacement offers a better understanding of the mechanism underlying HDR in plants, and greatly simplifies the design of DRTs for precision genome editing in crop improvement.

Efficient allelic replacement in rice by gene editing: A case study of the NRT1.1B gene.

Precise replacement of an existing allele in commercial cultivars with an elite allele is a major goal in crop breeding. A single nucleotide polymorphism in the NRT1.1B gene between japonica and indica rice is responsible for the improved nitrogen use efficiency in indica rice. Herein, we precisely replaced the japonica NRT1.1B allele with the indica allele, in just one generation, using CRISPR/Cas9 gene-editing technology. No additional selective pressure was needed to enrich the precise replacement events. This work demonstrates the feasibility of replacing any genes with elite alleles within one generation, greatly expanding our ability to improve agriculturally important traits.