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1.
Article in English | MEDLINE | ID: mdl-38696218

ABSTRACT

The newly emerging liquid metal flexible electronics are gaining increasing applications over the world due to their outstanding adaptability and printability. Here, we proposed a generalized purpose thermo-activated hybrid transfer printing method for the rapid fabrication of multifunctional soft electronics, which can significantly reduce the difficulty facing existing technologies. This printing involves two delivery and deposition processes of liquid metals and the allied inks (toners) based on their adhesion selection mechanisms. Through developing office supplies, the laser printer could directly print toner masks on soft substrates, such as polydimethylsiloxane film. The heating plate was applied to remove the toner sacrificial mask after rolling liquid metal inks, resulting in retaining the liquid metal circuits on the target substrate. For illustration, diverse materials and inks are adapted to the strategy of constructing flexible electronics. Particularly, colorful circuits, flexible heaters, transparent circuitry, and soft programmable light-emitting diode array displays with multilayer circuits have been fabricated and tested. This general and easily accessible method allows for the rapid acquisition of user-designed soft electronics and is expected to promote the widespread use of flexible electronics in e-skin, sensing, displays, etc.

2.
Mitochondrial DNA B Resour ; 9(4): 465-469, 2024.
Article in English | MEDLINE | ID: mdl-38591052

ABSTRACT

Strobilanthes dalzielii of Acanthaceae is an herb species with potentially extensive applications for its pharmaceutical and ornamental values. Due to taxonomic complications and limited genetic information, the structural characteristics, and phylogenetic relationships of the S. dalzielii chloroplast genome were assembled and characterized here for the first time. The complete chloroplast genome of S. dalzielii was 144,580 bp in length. The genome is quadripartite in structure and consists of a large single-copy region (92,137 bp) and a small single-copy region (17,669 bp), which are separated by a pair of inverted repeats (each 17,387 bp). A total of 125 genes were annotated, including 80 protein-coding, 37 transfer RNA, and eight ribosomal RNA genes. The overall GC content was 36.4%. Phylogenetic analysis based on the complete chloroplast genome sequence of 21 taxa within the tribe Ruellieae of Acanthaceae using the maximum likelihood and Bayesian inference methods revealed that Strobilanthes diverged after Ruellia; S. dalzielii is closely related to S. tonkinensis. The genomic data obtained from this study will serve as valuable information to the species delimitation and genetic classification of Strobilanthes.

3.
World J Gastroenterol ; 30(7): 774-778, 2024 Feb 21.
Article in English | MEDLINE | ID: mdl-38515953

ABSTRACT

This research aimed to examine the diagnostic accuracy and clinical significance of endoscopic ultrasonography (EUS) in the context of small rectal neuroendocrine neoplasms (NENs). A total of 108 patients with rectal subepithelial lesions (SELs) with a diameter of < 20 mm were included in the analysis. The diagnosis and depth assessment of EUS was compared to the histology findings. The prevalence of NENs in rectal SELs was 78.7% (85/108). The sensitivity of EUS in detecting rectal NENs was 98.9% (84/85), while the specificity was 52.2% (12/23). Overall, the diagnostic accuracy of EUS in identifying rectal NENs was 88.9% (96/108). The overall accuracy rate for EUS in assessing the depth of invasion in rectal NENs was 92.9% (78/84). Therefore, EUS demonstrates reasonable diagnostic accuracy in detecting small rectal NENs, with good sensitivity but inferior specificity. EUS may also assist physicians in assessing the depth of invasion in small rectal NENs before endoscopic excision.


Subject(s)
Neuroendocrine Tumors , Rectal Neoplasms , Humans , Endosonography , Clinical Relevance , Neuroendocrine Tumors/pathology , Rectal Neoplasms/pathology , Rectum/diagnostic imaging , Rectum/pathology
4.
Microbiol Res ; 282: 127651, 2024 May.
Article in English | MEDLINE | ID: mdl-38430888

ABSTRACT

Climate change and anthropogenic disturbances are known to influence soil biodiversity. The objectives of this study were to compare the community composition, species coexistence patterns, and ecological assembly processes of soil microbial communities in a paired setting featuring a natural and an anthropogenic ecosystem facing each other at identical climatic, pedological, and vegetational conditions. A transect gradient from forest to seashore allowed for sampling across different habitats within both sites. The field survey was carried out at two adjacent strips of land within the Po River delta lagoon system (Veneto, Italy) one of which is protected within a natural preserve and the other has been converted for decades into a tourist resort. The anthropogenic pressure interestingly led to an increase in the α-diversity of soil microbes but was accompanied by a reduction in ß-diversity. The community assembly mechanisms of microbial communities differentiate in natural and anthropic ecosystems: for bacteria, in natural ecosystems deterministic variables and homogeneous selection play a main role (51.92%), while stochastic dispersal limitation (52.15%) is critical in anthropized ecosystems; for fungi, stochastic dispersal limitation increases from 38.1% to 66.09% passing from natural to anthropized ecosystems. We are on calcareous sandy soils and in more natural ecosystems a variation of topsoil pH favors the deterministic selection of bacterial communities, while a divergence of K availability favors stochastic selection. In more anthropized ecosystems, the deterministic variable selection is influenced by the values of SOC. Microbial networks in the natural system exhibited higher numbers of nodes and network edges, as well as higher averages of path length, weighted degree, clustering coefficient, and density than its equivalent sites in the more anthropically impacted environment. The latter on the other hand presented a stronger modularity. Although the influence of stochastic processes increases in anthropized habitats, niche-based selection also proves to impose constraints on communities. Overall, the functionality of the relationships between groups of microorganisms co-existing in communities appeared more relevant to the concept of functional biodiversity in comparison to the plain number of their different taxa. Fewer but functionally more organized lineages displayed traits underscoring a better use of the resources than higher absolute numbers of taxa when those are not equally interconnected in their habitat exploitation. However, considering that network complexity can have important implications for microbial stability and ecosystem multifunctionality, the extinction of complex ecological interactions in anthropogenic habitats may impair important ecosystem services that soils provide us.


Subject(s)
Ecosystem , Microbiota , Soil Microbiology , Biodiversity , Forests , Soil/chemistry , Bacteria/genetics
5.
Anal Methods ; 16(8): 1252-1260, 2024 Feb 22.
Article in English | MEDLINE | ID: mdl-38323334

ABSTRACT

Acute pancreatitis (AP) is a surgical abdominal disease for which the Dachengqi Decoction (DCQD) of traditional Chinese medicine (TCM) is widely used in China. This study aims to analyse the pharmacodynamic interactions and quantitative relationship of DCQD in the treatment of AP based on orthogonal partial least squares (OPLS) analysis. The experimental data show organic chemical components as candidate pharmacodynamic substances (PS) in the blood and include pharmacodynamic indicators (PIs). Taking each PI as the target and using OPLS method to construct three types of mathematical equations, including the mathematical relationship between the pharmacodynamic substances and each target pharmacodynamic indicator (PS-TPI); the mathematical relationship between the pharmacodynamic substances, the pharmacodynamics indicators and each target pharmacodynamic indicator (PS, PI-TPI); and the mathematical relationship between the pharmacodynamic indicators and each target pharmacodynamic indicator (PI-TPI). Through analysis, we find that the R2Y(cum) values and VIP values indicate that PS and PI are the follow-up factors of TPI; the coefficient value indicates that there is a quantitative relationship between the PS and the TPI; and there also is a quantitative relationship between PI and TPI. The results demonstrated that PS and other PIs are the important influencing factors of TPI, and that there are interactions and quantitative relationships among the PIs.


Subject(s)
Pancreatitis , Rats , Animals , Pancreatitis/drug therapy , Medicine, Chinese Traditional , Least-Squares Analysis , Acute Disease , Rats, Sprague-Dawley
6.
J Exp Med ; 221(3)2024 Mar 04.
Article in English | MEDLINE | ID: mdl-38284990

ABSTRACT

Human lung adenosquamous cell carcinoma (LUAS), containing both adenomatous and squamous pathologies, exhibits strong cancer plasticity. We find that ALK rearrangement is detectable in 5.1-7.5% of human LUAS, and transgenic expression of EML4-ALK drives lung adenocarcinoma (LUAD) formation initially and squamous transition at late stage. We identify club cells as the main cell-of-origin for squamous transition. Through recapitulating lineage transition in organoid system, we identify JAK-STAT signaling, activated by EML4-ALK phase separation, significantly promotes squamous transition. Integrative study with scRNA-seq and immunostaining identify a plastic cell subpopulation in ALK-rearranged human LUAD showing squamous biomarker expression. Moreover, those relapsed ALK-rearranged LUAD show notable upregulation of squamous biomarkers. Consistently, mouse squamous tumors or LUAD with squamous signature display certain resistance to ALK inhibitor, which can be overcome by combined JAK1/2 inhibitor treatment. This study uncovers strong plasticity of ALK-rearranged tumors in orchestrating phenotypic transition and drug resistance and proposes a potentially effective therapeutic strategy.


Subject(s)
Adenocarcinoma of Lung , Carcinoma, Squamous Cell , Lung Neoplasms , Humans , Animals , Mice , Lung Neoplasms/genetics , Lung , Receptor Protein-Tyrosine Kinases , Oncogene Proteins, Fusion/genetics
7.
Nat Struct Mol Biol ; 31(1): 42-53, 2024 Jan.
Article in English | MEDLINE | ID: mdl-38177668

ABSTRACT

DNA cytosine methylation plays a vital role in repressing retrotransposons, and such derepression is linked with developmental failure, tumorigenesis and aging. DNA methylation patterns are formed by precisely regulated actions of DNA methylation writers (DNA methyltransferases) and erasers (TET, ten-eleven translocation dioxygenases). However, the mechanisms underlying target-specific oxidation of 5mC by TET dioxygenases remain largely unexplored. Here we show that a large low-complexity domain (LCD), located in the catalytic part of Tet enzymes, negatively regulates the dioxygenase activity. Recombinant Tet3 lacking LCD is shown to be hyperactive in converting 5mC into oxidized species in vitro. Endogenous expression of the hyperactive Tet3 mutant in mouse oocytes results in genome-wide 5mC oxidation. Notably, the occurrence of aberrant 5mC oxidation correlates with a consequent loss of the repressive histone mark H3K9me3 at ERVK retrotransposons. The erosion of both 5mC and H3K9me3 causes ERVK derepression along with upregulation of their neighboring genes, potentially leading to the impairment of oocyte development. These findings suggest that Tet dioxygenases use an intrinsic auto-regulatory mechanism to tightly regulate their enzymatic activity, thus achieving spatiotemporal specificity of methylome reprogramming, and highlight the importance of methylome integrity for development.


Subject(s)
5-Methylcytosine , Dioxygenases , Animals , Mice , 5-Methylcytosine/metabolism , Dioxygenases/genetics , Dioxygenases/metabolism , Retroelements/genetics , DNA Methylation , Oocytes/metabolism , Demethylation
8.
Zhongguo Zhong Yao Za Zhi ; 48(19): 5345-5355, 2023 Oct.
Article in Chinese | MEDLINE | ID: mdl-38114124

ABSTRACT

The study investigated the effect of Buyang Huanwu Decoction(BYHWD) on endogenous biomarkers in the urine of rats with chronic inflammation induced by lipopolysaccharide(LPS) using ultra-high performance liquid chromatography-quadrupole-time-of-flight-mass spectrometry(UPLC-Q-TOF-MS), aiming to elucidate the molecular mechanism underlying the therapeutic effect of BYHWD on chronic inflammation from a metabolomics perspective. Male SD rats were randomly divided into a normal group, a model group, and low-, medium-, and high-dose BYHWD groups(7.5, 15, and 30 g·kg~(-1)). The model group and BYHWD groups received tail intravenous injection of LPS(200 µg·kg~(-1)) on the first day of each week, followed by oral administration of BYHWD once a day for four consecutive weeks. Urine samples were collected at the end of the administration period, and UPLC-Q-TOF-MS was used to analyze the metabolic profiles of the rat urine in each group. Multivariate statistical analysis methods such as principal component analysis(PCA), partial least squares-discriminant analysis(PLS-DA), and orthogonal partial least squares-discriminant analysis(OPLS-DA) were used to analyze the effect of BYHWD on endogenous metabolites. One-way ANOVA and variable importance for the projection(VIP) were used to screen for potential biomarkers related to chronic inflammation. The identified biomarkers were subjected to pathway and enrichment analysis using MetaboAnalyst 5.0. A total of 25 potential biomarkers were screened and identified in the rat urine in this experiment. Compared with the normal group, the model group showed significant increases in the levels of 14 substances(P<0.05) and significant decreases in the levels of 11 substances(P<0.05). BYHWD was able to effectively reverse the trend of most endogenous biomarkers. Compared with the model group, BYHWD significantly down-regulated 13 biomarkers(P<0.05) and up-regulated 10 biomarkers(P<0.05). The metabolic products were mainly related to the biosynthesis of pantothenic acid and coenzyme A, tryptophan metabolism, retinol metabolism, and propionate metabolism. BYHWD has therapeutic effect on chronic inflammation induced by LPS, which may be related to its ability to improve the levels of endogenous metabolites, enhance the body's anti-inflammatory and antioxidant capabilities, and restore normal metabolic activity.


Subject(s)
Lipopolysaccharides , Metabolomics , Rats , Male , Animals , Chromatography, High Pressure Liquid/methods , Rats, Sprague-Dawley , Metabolomics/methods , Inflammation/drug therapy , Biomarkers/urine
10.
Mitochondrial DNA B Resour ; 8(10): 1137-1140, 2023.
Article in English | MEDLINE | ID: mdl-37928400

ABSTRACT

Oreocharis argyreia var. angustifolia of Gesneriaceae is widely distributed in South China, including Guangdong, Guangxi, Hunan, and Jiangxi provinces. However, genetic information of this species is limited, further contributing to the taxonomic complications surrounding this species. Thus, in this study, we assembled and characterized the complete chloroplast genome of O. argyreia var. angustifolia as a genomic resource for future studies. The complete plastid genome was 154,675 bp in size, with a pair of inverted repeat regions of 25,329 bp each, separating the 85,977-bp large and 18,040-bp small single copy regions. A total of 131 genes were predicted, consisting of 86 protein-coding, 37 tRNA, and eight rRNA genes. The overall GC content was 37.6%. Phylogenetic analysis based on 79 shared unique CDS resulted in a fully resolved phylogenetic tree using both the maximum likelihood and Bayesian inference methods. Based on current circumscription, both methods indicated that Oreocharis is monophyletic; O. argyreia var. angustifolia diverged after O. chienii, which then followed by the divergence of the other three species included namely, O. continifolia, O. esquirolii, and O. mileensis. The genomic data obtained will be useful for future studies on the phylogenetics and evolution of Gesneriaceae.

11.
RSC Adv ; 13(45): 31772-31784, 2023 Oct 26.
Article in English | MEDLINE | ID: mdl-37908648

ABSTRACT

The objective of this study was to investigate the anticancer activities of biodegradable polymeric micelles composed of monomethoxy poly(ethylene glycol), polylactic acid, and nitric oxide (mPEG-PLA-NO) loaded with paclitaxel (PTX) as a nanomedicine delivery system. We aimed to compare the anticancer effects of these NO/PTX micelles with PTX alone and elucidate their mechanism of action. We evaluated the impact of NO/PTX and PTX on cell viability using Cell Counting Kit-8 (CCK8) assays conducted on the Bel-7402 liver cancer cell line. Additionally, we employed H22 xenografted mice to assess the in vivo tumor growth inhibitory activity of NO/PTX. To examine the cytotoxicity of NO/PTX, the intracellular levels of reactive oxygen species (ROS), and the expression of ferroptosis-related proteins, we conducted experiments in the presence of the ferroptosis inhibitor ferrostatin-1 (Fer-1) or the ROS inhibitor N-acetyl cysteine (NAC). Furthermore, we investigated the expression of endoplasmic reticulum stress (ERS) and apoptosis-associated proteins. Our results demonstrated that NO/PTX exhibited enhanced anticancer effects compared to PTX alone in both Bel-7402 cells and H22 xenografted mice. The addition of Fer-1 or NAC reduced the anticancer activity of NO/PTX, indicating the involvement of ferroptosis and ROS in its mechanism of action. Furthermore, NO/PTX modulated the expression of proteins related to ERS and apoptosis, indicating the activation of these cellular pathways. The anticancer effects of NO/PTX in liver cancer cells were mediated through the induction of ferroptosis, pyroptosis, ERS, and apoptosis-associated networks. Ferroptosis and pyroptosis were activated by treatment of NO/PTX at low concentration, whereas ERS was induced to trigger apoptosis at high concentration. The superior anti-tumor effect of NO/PTX may be attributed to the downregulation of a multidrug resistance transporter and the sensitization of cells to PTX chemotherapy. In summary, our study highlights the potential of mPEG-PLA-NO micelles loaded with PTX as a nanomedicine delivery system for liver cancer treatment. The observed enhancement in anticancer activity, combined with the modulation of key cellular pathways, provides valuable insights into the therapeutic potential of NO/PTX in overcoming resistance and improving treatment outcomes in liver cancer patients.

12.
Leukemia ; 37(12): 2457-2467, 2023 12.
Article in English | MEDLINE | ID: mdl-37816954

ABSTRACT

Somatic loss-of-function mutations of the dioxygenase Ten-eleven translocation-2 (TET2) occur frequently in individuals with clonal hematopoiesis (CH) and acute myeloid leukemia (AML). These common hematopoietic disorders can be recapitulated in mouse models. However, the underlying mechanisms by which the deficiency in TET2 promotes these disorders remain unclear. Here we show that the cyclic guanosine monophosphate-adenosine monophosphate synthase (cGAS)-stimulator of interferon genes (STING) pathway is activated to mediate the effect of TET2 deficiency in dysregulated hematopoiesis in mouse models. DNA damage arising in Tet2-deficient hematopoietic stem/progenitor cells (HSPCs) leads to activation of the cGAS-STING pathway which in turn promotes the enhanced self-renewal and development of CH. Notably, both pharmacological inhibition and genetic deletion of STING suppresses Tet2 mutation-induced aberrant hematopoiesis. In patient-derived xenograft (PDX) models, STING inhibition specifically attenuates the proliferation of leukemia cells from TET2-mutated individuals. These observations suggest that the development of CH associated with TET2 mutations is powered through chronic inflammation dependent on the activated cGAS-STING pathway and that STING may represent a potential target for intervention of relevant hematopoietic diseases.


Subject(s)
Dioxygenases , Hematologic Diseases , Mice , Animals , Humans , Cell Transformation, Neoplastic/genetics , Translocation, Genetic , Hematopoiesis/genetics , Nucleotidyltransferases/genetics , Nucleotidyltransferases/pharmacology , Stem Cells/metabolism , DNA-Binding Proteins/metabolism , Dioxygenases/genetics
13.
Phys Chem Chem Phys ; 25(41): 28230-28240, 2023 Oct 25.
Article in English | MEDLINE | ID: mdl-37823325

ABSTRACT

Carbon-quantum-dot-based fluorescence sensing of Hg2+ is a well-known cost-effective tactic with fast response and high sensitivity, while rationally constructing heteroatom-doped carbon quantum dots with improved fluorescence sensing performances through tuning the electronic and chemical structures of the reactive site still remains a challenging project for monitoring trace Hg2+ in aquatic ecosystems to avoid harm resulting from its high toxicity, nonbiodegradabilty and accumulative effects on human health. Herein, intriguing N,S-codoped carbon quantum dots were synthesized via a facile one-step hydrothermal procedure. As an admirable fluorescent probe with plentiful heteroatom-related functional groups, these N,S-codoped carbon quantum dots can exhibit an absolute fluorescence quantum yield as high as 11.6%, excellent solubility and stability over three months, remarkable sensitivity for Hg2+ detection with an attractive detection limit of 0.27 µg L-1 and admirable selectivity for Hg2+ against thirteen other metal ions. Density functional theory calculations reveal that electron-enriched meta-S of the unique graphitic N with homocyclic meta-thiophene sulfur structure can regulate this N site to have more electrons and preferable affinity towards Hg, hence achieving enhanced fluorescence quenching due to greater charge transfer from N to Hg after the coordination interaction. This strategy provides a promising avenue for precisely designing purpose-made quantum dots with the dedicated fluorescence sensing applications.

14.
Signal Transduct Target Ther ; 8(1): 343, 2023 09 12.
Article in English | MEDLINE | ID: mdl-37696812

ABSTRACT

Chromobox protein homolog 4 (CBX4) is a component of the Polycomb group (PcG) multiprotein Polycomb repressive complexes 1 (PRC1), which is participated in several processes including growth, senescence, immunity, and tissue repair. CBX4 has been shown to have diverse, even opposite functions in different types of tissue and malignancy in previous studies. In this study, we found that CBX4 deletion promoted lung adenocarcinoma (LUAD) proliferation and progression in KrasG12D mutated background. In vitro, over 50% Cbx4L/L, KrasG12D mouse embryonic fibroblasts (MEFs) underwent apoptosis in the initial period after Adeno-Cre virus treatment, while a small portion of survival cells got increased proliferation and transformation abilities, which we called selected Cbx4-/-, KrasG12D cells. Karyotype analysis and RNA-seq data revealed chromosome instability and genome changes in selected Cbx4-/-, KrasG12D cells compared with KrasG12D cells. Further study showed that P15, P16 and other apoptosis-related genes were upregulated in the primary Cbx4-/-, KrasG12D cells due to chromosome instability, which led to the large population of cell apoptosis. In addition, multiple pathways including Hippo pathway and basal cell cancer-related signatures were altered in selected Cbx4-/-, KrasG12D cells, ultimately leading to cancer. We also found that low expression of CBX4 in LUAD was associated with poorer prognosis under Kras mutation background from the human clinical data. To sum up, CBX4 deletion causes genomic instability to induce tumorigenesis under KrasG12D background. Our study demonstrates that CBX4 plays an emerging role in tumorigenesis, which is of great importance in guiding the clinical treatment of lung adenocarcinoma.


Subject(s)
Adenocarcinoma of Lung , Ligases , Lung Neoplasms , Polycomb Repressive Complex 1 , Animals , Humans , Mice , Adenocarcinoma of Lung/genetics , Carcinogenesis/genetics , Cell Transformation, Neoplastic/genetics , Chromosomal Instability , Fibroblasts , Genomic Instability/genetics , Ligases/genetics , Lung Neoplasms/genetics , Polycomb Repressive Complex 1/genetics
15.
J Med Virol ; 95(8): e29041, 2023 08.
Article in English | MEDLINE | ID: mdl-37621182

ABSTRACT

The emerging outbreak of monkeypox is closely associated with the viral infection and spreading, threatening global public health. Virus-induced cell migration facilitates viral transmission. However, the mechanism underlying this type of cell migration remains unclear. Here we investigate the motility of cells infected by vaccinia virus (VACV), a close relative of monkeypox, through combining multi-omics analyses and high-resolution live-cell imaging. We find that, upon VACV infection, the epithelial cells undergo epithelial-mesenchymal transition-like transformation, during which they lose intercellular junctions and acquire the migratory capacity to promote viral spreading. After transformation, VACV-hijacked RhoA signaling significantly alters cellular morphology and rearranges the actin cytoskeleton involving the depolymerization of robust actin stress fibers, leading-edge protrusion formation, and the rear-edge recontraction, which coordinates VACV-induced cell migration. Our study reveals how poxviruses alter the epithelial phenotype and regulate RhoA signaling to induce fast migration, providing a unique perspective to understand the pathogenesis of poxviruses.


Subject(s)
Mpox (monkeypox) , Vaccinia virus , Humans , Cell Movement , Disease Outbreaks , Epithelial Cells
16.
Polymers (Basel) ; 15(14)2023 Jul 17.
Article in English | MEDLINE | ID: mdl-37514462

ABSTRACT

In this work, a model for predicting the leakage rate was developed to investigate the effect of irradiation on the sealing performance of ethylene propylene diene monomer (EPDM) O-rings. The model is based on a mesoscopic interfacial gap flow simulation and accurately predicts the sealing performance of irradiated and non-irradiated materials by utilizing the gap height as an indicator in a mechanical simulation of the O-ring under operating conditions. A comparison with vacuum test results indicates that the model is a good predictor of leak initiation. The positive pressure leakage of the O-rings was investigated numerically. The results show the following. The sealing performance of the non-irradiated O-ring is much better than that of the irradiated one. The sealing performance is the worst at 0. 713 MGy and the best at 1.43 MGy, and the seal is maintained at an absorbed dose of 3.55 MGy. A theoretical analysis of the non-monotonic variation using the proposed model shows that the leakage behavior of the O-rings depends not only on the material properties but also on the roughness and prestressing properties. Finally, a method was proposed to classify the sealing performance, using the maximum allowable leakage rate as an indicator.

17.
Plant J ; 115(3): 846-865, 2023 08.
Article in English | MEDLINE | ID: mdl-37310200

ABSTRACT

Precise gene-editing using CRISPR/Cas9 technology remains a long-standing challenge, especially for genes with low expression and no selectable phenotypes in Chlamydomonas reinhardtii, a classic model for photosynthesis and cilia research. Here, we developed a multi-type and precise genetic manipulation method in which a DNA break was generated by Cas9 nuclease and the repair was mediated using a homologous DNA template. The efficacy of this method was demonstrated for several types of gene editing, including inactivation of two low-expression genes (CrTET1 and CrKU80), the introduction of a FLAG-HA epitope tag into VIPP1, IFT46, CrTET1 and CrKU80 genes, and placing a YFP tag into VIPP1 and IFT46 for live-cell imaging. We also successfully performed a single amino acid substitution for the FLA3, FLA10 and FTSY genes, and documented the attainment of the anticipated phenotypes. Lastly, we demonstrated that precise fragment deletion from the 3'-UTR of MAA7 and VIPP1 resulted in a stable knock-down effect. Overall, our study has established efficient methods for multiple types of precise gene editing in Chlamydomonas, enabling substitution, insertion and deletion at the base resolution, thus improving the potential of this alga in both basic research and industrial applications.


Subject(s)
Chlamydomonas reinhardtii , Chlamydomonas , CRISPR-Cas Systems , Chlamydomonas/genetics , Gene Editing/methods , Chlamydomonas reinhardtii/genetics
19.
Epigenetics ; 18(1): 2216005, 2023 12.
Article in English | MEDLINE | ID: mdl-37246786

ABSTRACT

While epigenetic mechanisms such as DNA methylation and histone modification are known to be important for gene suppression, relatively little is still understood about the interplay between these systems. The UHRF1 protein can interact with both DNA methylation and repressive chromatin marks, but its primary function in humans has been unclear. To determine what that was, we first established stable UHRF1 knockdowns (KD) in normal, immortalized human fibroblasts using targeting shRNA, since CRISPR knockouts (KO) were lethal. Although these showed a loss of DNA methylation across the whole genome, transcriptional changes were dominated by the activation of genes involved in innate immune signalling, consistent with the presence of viral RNA from retrotransposable elements (REs). We confirmed using mechanistic approaches that 1) REs were demethylated and transcriptionally activated; 2) this was accompanied by activation of interferons and interferon-stimulated genes and 3) the pathway was conserved across other adult cell types. Restoring UHRF1 in either transient or stable KD systems could abrogate RE reactivation and the interferon response. Notably, UHRF1 itself could also re-impose RE suppression independent of DNA methylation, but not if the protein contained point mutations affecting histone 3 with trimethylated lysine 9 (H3K9me3) binding. Our results therefore show for the first time that UHRF1 can act as a key regulator of retrotransposon silencing independent of DNA methylation.


Subject(s)
DNA Methylation , RNA, Viral , Humans , RNA, Viral/metabolism , Ubiquitin-Protein Ligases/genetics , Ubiquitin-Protein Ligases/metabolism , CCAAT-Enhancer-Binding Proteins/genetics , CCAAT-Enhancer-Binding Proteins/metabolism , Immunity, Innate/genetics , Interferons/metabolism
20.
PhytoKeys ; 226: 1-16, 2023.
Article in English | MEDLINE | ID: mdl-37207080

ABSTRACT

Primulinajiulianshanensis F.Wen & G.L.Xu, a new species of Gesneriaceae from Jiulianshan National Nature Reserve of Jiangxi Province, China, is described and illustrated here. Molecular evidence showed it was sister to P.wenii Jian Li & L.J.Yan, while the morphological observation found clear differences between them, petiole, both sides of leaf blades, adaxial surface of the calyx lobes, corolla inside toward the bottom, bract margins covered glandular-pubescent hairs in P.jiulianshanensis (vs. no glandular-pubescent hairs in P.wenii); lateral bracts 4-9 × ca. 2 mm, the central one 2-5 × 1-1.5 mm, adaxially glabrous but sparsely pubescent at apex (vs. lateral bracts 14-16 × 2.5-3.0 mm, the central one 10-12 × 1.3-1.6 mm, all adaxially pubescent); calyx lobes 8-11 × ca. 2 mm, each side with several brown serrate teeth at apex (vs. 14-15 × ca. 2.5 mm, margin entire); filaments and staminodes sparsely yellow glandular-puberulent (vs. white, glabrous).

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