It is now widely known that C-X3-C motif ligand 1 (CX3CL1) plays an essential part in the process of regulating pro-inflammatory cells migration across a wide range of inflammatory disorders, ...including a number of malignancies. However, there has been no comprehensive study on the correlation between CX3CL1 and cancers on the basis of clinical features. In order to investigate the potential function of CX3CL1 in the clinical prognosis and immunotherapy, I evaluated the expression of CX3CL1 in numerous cancer types, methylation levels and genetic alterations. I found CX3CL1 was differentially expressed in numerous cancer types, which indicated CX3CL1 may plays a potential role in tumor progression. Furthermore, CX3CL1 was variably expressed in methylation levels and gene alterations in most cancers according to The Cancer Genome Atlas (TCGA). CX3CL1 was robustly associated with clinical characteristics and pathological stages, suggesting that it was related to the degree of tumor malignancy and the physical function of patients. As determined by the Kaplan-Meier method of estimating survival, high CX3CL1 expression was associated with either favorable or unfavorable outcomes depending on the different types of cancer. It suggests the correlation between CX3CL1 and tumor prognosis. Significant positive correlations of CX3CL1 expression with CD4
T cells, M1 macrophage cells and activated mast cells have been established in the majority of TCGA malignancies. Which indicates CX3CL1 plays an important role in tumor immune microenvironment. Gene Ontology (GO) terms and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis suggested that the chemokine signaling pathway may shed light on the pathway for CX3CL1 to exert function. In a conclusion, our study comprehensively summarizes the potential role of CX3CL1 in clinical prognosis and immunotherapy, suggesting that CX3CL1 may represent a promising pharmacological treatment target of tumors.
Genome editing holds promise for correcting pathogenic mutations. However, it is difficult to determine off-target effects of editing due to single-nucleotide polymorphism in individuals. Here we ...developed a method named GOTI (genome-wide off-target analysis by two-cell embryo injection) to detect off-target mutations by editing one blastomere of two-cell mouse embryos using either CRISPR-Cas9 or base editors. Comparison of the whole-genome sequences of progeny cells of edited and nonedited blastomeres at embryonic day 14.5 showed that off-target single-nucleotide variants (SNVs) were rare in embryos edited by CRISPR-Cas9 or adenine base editor, with a frequency close to the spontaneous mutation rate. By contrast, cytosine base editing induced SNVs at more than 20-fold higher frequencies, requiring a solution to address its fidelity.
Extratropical cyclones (ETCs) over East Asia and Northwest Pacific are identified and tracked by applying an objective algorithm to the 850-hPa relative vorticity fields from the ERA-Interim ...reanalysis. A total of 2866 ETCs originating at the western side of the date line have been identified in the extended November–March winters from 1979 to 2018. The ETC tracks are counted and visualized using a hexagonal tessellation rather than the regular longitude–latitude grids. Two generalized linear models (GLMs), Poisson regression model and Gamma regression model, are firstly applied to investigate the associations of wintertime ETCs with three atmospheric teleconnection patterns. The West Pacific (WP) pattern and the Pacific/North American (PNA) pattern are more responsible for the meridional variability of ETC activities in the North Pacific, while the influence of the Polar/Eurasia pattern on ETC activities is negligible. Results of composite analysis are qualitatively consistent with that of regression analysis. Composite maps of differences of jet stream, thermal gradient and mid-tropospheric baroclinicity in the positive and negative phases of teleconnection patterns also support the close associations of ETC activities with WP and PNA teleconnection patterns.
Cytosine base editors (CBEs) offer a powerful tool for correcting point mutations, yet their DNA and RNA off-target activities have caused concerns in biomedical applications. We describe screens of ...23 rationally engineered CBE variants, which reveal mutation residues in the predicted DNA-binding site can dramatically decrease the Cas9-independent off-target effects. Furthermore, we obtained a CBE variant-YE1-BE3-FNLS-that retains high on-target editing efficiency while causing extremely low off-target edits and bystander edits.
Conversion of glial cells into functional neurons represents a potential therapeutic approach for replenishing neuronal loss associated with neurodegenerative diseases and brain injury. Previous ...attempts in this area using expression of transcription factors were hindered by the low conversion efficiency and failure of generating desired neuronal types in vivo. Here, we report that downregulation of a single RNA-binding protein, polypyrimidine tract-binding protein 1 (Ptbp1), using in vivo viral delivery of a recently developed RNA-targeting CRISPR system CasRx, resulted in the conversion of Müller glia into retinal ganglion cells (RGCs) with a high efficiency, leading to the alleviation of disease symptoms associated with RGC loss. Furthermore, this approach also induced neurons with dopaminergic features in the striatum and alleviated motor defects in a Parkinson’s disease mouse model. Thus, glia-to-neuron conversion by CasRx-mediated Ptbp1 knockdown represents a promising in vivo genetic approach for treating a variety of disorders due to neuronal loss.
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•Knockdown of Ptbp1 converts Müller glia into retinal ganglion cells in mature retinas•Central projections of converted retinal ganglion cells restore visual responses•Induction of neurons with dopaminergic features in PD model mice•Induced neurons alleviated motor dysfunctions in PD mice
In vivo CasRx-mediated downregulation of a single RNA-binding protein, Ptbp1, locally converts glia to neurons and shows promise for treating disorders due to neuronal loss in mice.
Clathrin-mediated endocytosis is an essential cellular process that involves the concerted assembly and disassembly of many different proteins at the plasma membrane. In yeast, live-cell imaging has ...shown that the spatiotemporal dynamics of these proteins is highly stereotypical. Recent work has focused on determining how the timing and functions of endocytic proteins are regulated. In this Cell Science at a Glance article and accompanying poster, we review our current knowledge of the timeline of endocytic site maturation and discuss recent works focusing on how phosphorylation, ubiquitylation and lipids regulate various aspects of the process.
Abstract
Efficient and precise base editors (BEs) for C-to-G transversion are highly desirable. However, the sequence context affecting editing outcome largely remains unclear. Here we report ...engineered C-to-G BEs of high efficiency and fidelity, with the sequence context predictable via machine-learning methods. By changing the species origin and relative position of uracil-DNA glycosylase and deaminase, together with codon optimization, we obtain optimized C-to-G BEs (OPTI-CGBEs) for efficient C-to-G transversion. The motif preference of OPTI-CGBEs for editing 100 endogenous sites is determined in HEK293T cells. Using a sgRNA library comprising 41,388 sequences, we develop a deep-learning model that accurately predicts the OPTI-CGBE editing outcome for targeted sites with specific sequence context. These OPTI-CGBEs are further shown to be capable of efficient base editing in mouse embryos for generating
Tyr
-edited offspring. Thus, these engineered CGBEs are useful for efficient and precise base editing, with outcome predictable based on sequence context of targeted sites.
Understanding the distribution of water and nitrate nitrogen in the soil profile is crucial for the reasonable operation of fertigation, and it is also fundamental for controlling and regulating ...nitrate nitrogen in the root zone, thereby meeting a crop’s requirements. The application rates of fertilizer and water directly influence this distribution of water and nitrate nitrogen. However, the effects in Aeolian sandy soil, a type of developing soil bordering deserts, remain ambiguous. In this study, field experiments for different drip fertigation treatments in Aeolian sandy soil were conducted to investigate the soil water distribution, as well as that of nitrate nitrogen. A completely randomized experimental design was implemented, encompassing three levels of irrigation amount: low (W1), medium (W2), and high (W3), and three levels of nitrogen application rate: low (F1), medium (F2), high (F3). After the completion of each irrigation treatment, soil samples were extracted at 10–20 cm intervals. The soil water and nitrate nitrogen contents in the profiles of these samples were measured. The experimental results revealed that increasing the nitrogen application rate facilitated the retention of greater amounts of water and nitrate nitrogen in the soil profile. However, with an increase in the nitrogen application rate, both soil water and nitrate nitrogen exhibited a radial tendency to move away from the drip emitter. Some moved upward and accumulated in surface soil near a ridge furrow, while some moved downward and remained in a deeper area approximately 30 cm horizontally from the emitter at depths of 40–60 cm. The uniformity of the water distribution decreased with increasing nitrogen application under low water conditions, with a reversal of this trend observed in medium and high water treatments. The effect of nitrogen application level on the uniformity of the nitrate nitrogen distribution was not significant. There was no significant correlation between the average soil water content and nitrate nitrogen content along the horizontal direction, however, a positive correlation existed in the vertical direction. In the whole profile, increasing the nitrogen application enhanced the correlation under low water conditions, but under medium and high water conditions, this trend was the opposite. This implies that, to avoid nitrate nitrogen leaching or limiting in a specific area, a moderate nitrogen application level is advisable. Under low water conditions, nitrogen application showed a positive effect on the nitrate nitrogen content, and a higher application is recommended. In cases of substantial water irrigation or rainy years, the nitrogen application rate should be decreased.
The CRISPR/Cas9 system has become an efficient gene editing method for generating cells carrying precise gene mutations, including the rearrangement and deletion of chromosomal segments. However, ...whether an entire chromosome could be eliminated by this technology is still unknown.
Here we demonstrate the use of the CRISPR/Cas9 system to eliminate targeted chromosomes. Using either multiple cleavages induced by a single-guide RNA (sgRNA) that targets multiple chromosome-specific sites or a cocktail of multiple sgRNAs, each targeting one specific site, we found that a sex chromosome could be selectively eliminated in cultured cells, embryos, and tissues in vivo. Furthermore, this approach was able to produce a targeted autosome loss in aneuploid mouse embryonic stem cells with an extra human chromosome and human induced pluripotent stem cells with trisomy 21, as well as cancer cells.
CRISPR/Cas9-mediated targeted chromosome elimination offers a new approach to develop animal models with chromosome deletions, and a potential therapeutic strategy for human aneuploidy diseases involving additional chromosomes.
Genome editing holds great potential for correcting pathogenic mutations. We developed a method called GOTI (genome-wide off-target analysis by two-cell embryo injection) to detect off-target ...mutations by editing one blastomere of two-cell mouse embryos using either CRISPR-Cas9 or base editors. GOTI directly compares edited and non-edited cells without the interference of genetic background and thus could detect potential off-target variants with high sensitivity. Notably, the GOTI method was designed to detect potential off-target variants of any genome editing tools by the combination of experimental and computational approaches, which is critical for accurate evaluation of the safety of genome editing tools. Here we provide a detailed protocol for GOTI, including mice mating, two-cell embryo injection, embryonic day 14.5 embryo digestion, fluorescence-activated cell sorting, whole-genome sequencing and data analysis. To enhance the utility of GOTI, we also include a computational workflow called GOTI-seq (https://github.com/sydaileen/GOTI-seq) for the sequencing data analysis, which can generate the final genome-wide off-target variants from raw sequencing data directly. The protocol typically takes 20 d from the mice mating to sequencing and 7 d for sequencing data analysis.
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