Exosomal miRNA-92a derived from cancer-associated fibroblasts promote invasion and metastasis in breast cancer by regulating G3BP2.

Cancer-associated Fibroblasts (CAFs) exert a tumor-promoting effect in various cancers, including breast cancer. CAFs secrete exosomes containing miRNA and proteins, influencing the tumor microenvironment. In this study, we identified CAF-derived exosomes that transport functional miR-92a from CAFs to tumor cells, thereby intensifying the aggressiveness of breast cancer. CAFs downregulate the expression of G3BP2 in breast cancer cells, and a significant elevation in miR-92a levels in CAF-derived exosomes was observed. Both in vitro and in vivo experiments demonstrate that miR-92a enhances breast cancer cell migration and invasion by directly targeting G3BP2, functioning as a tumor-promoting miRNA. We validated that the RNA-binding proteins SNRPA facilitate the transfer of CAF-derived exosomal miR-92a to breast cancer cells. The reduction of G3BP2 protein by CAF-derived exosomes releases TWIST1 into the nucleus, promoting epithelial-mesenchymal transition (EMT) and further exacerbating breast cancer progression. Moreover, CAF-derived exosomal miR-92a induces tumor invasion and metastasis in mice. Overall, our study reveals that CAF-derived exosomal miR-92a serves as a promoter in the migration and invasion of breast cancer cells by reducing G3BP2 and may represent a potential novel tumor marker for breast cancer.

Chiral BINOL-phosphate assembled single hexagonal nanotube in aqueous solution for confined rearrangement acceleration.

Creating microenvironments that mimic an enzyme's active site is a critical aspect of supramolecular confined catalysis. In this study, we employ the commonly used chiral 1,1'-bi-2-naphthol (BINOL) phosphates as subcomponents to construct supramolecular hollow nanotube in an aqueous medium through non-covalent intermolecular recognition and arrangement. The hexagonal nanotubular structure is characterized by various techniques, including X-ray, NMR, ESI-MS, AFM, and TEM, and is confirmed to exist in a homogeneous aqueous solution stably. The nanotube's length in solution depends on the concentration of chiral BINOL-phosphate as a monomer. Additionally, the assembled nanotube can accelerate the rate of the 3-aza-Cope rearrangement reaction by up to 85-fold due to the interior confinement effect. Based on the detailed kinetic and thermodynamic analyses, we propose that the chain-like substrates are constrained and pre-organized into a reactive chair-like conformation, which stabilizes the transition state of the reaction in the confined nanospace of the nanotube. Notably, due to the restricted conformer with less degrees of freedom, the entropic barrier is significantly reduced compared to the enthalpic barrier, resulting in a more pronounced acceleration effect.

What drives the spatial heterogeneity of cropping patterns in the Northeast China: The natural environment, the agricultural economy, or policy?

Understanding the spatiotemporal dynamic of crop cover types and the driving forces of cropping patterns in the Northeast China (NEC) is essential for establishing suitable and sustainable cropping patterns that are adapted to local conditions, and for promoting the optimal use of black soil resources. Here, we classified the major grain crop cover types and investigated their spatiotemporal dynamic in the NEC by combining multi-source remote sensing imagery and phenological information based on the Google Earth Engine (GEE) platform. A number of typical cropping patterns from 2017 to 2021 were defined and extracted, and the characteristics of their spatial heterogeneity were analyzed. Driving mechanisms for the spatial heterogeneity of cropping patterns were revealed using Geodetector. The results concluded that over the past five years (2017-2021), there has been a shift from soybean to maize in the NEC, while rice has remained stable in terms of spatiotemporal dynamics. Seven dominant cropping patterns showed high spatial heterogeneity and positive spatial agglomeration. The center of gravity of the cropping pattern shifted southwards as the frequency of maize planting increased, while the center of gravity shifted northwards as the frequency of soybean planting increased, while the rice cropping pattern remained stable. The interaction between black-soil productivity index (BPI) and total grain income trend (TGIT) exhibits the most pronounced impact on the spatial heterogeneity of cropping patterns, with a q statistic of 0.523. Following closely are the interactions of soybean subsidies trend (SST), rice subsidies trend (RST), and maize subsidies trend (MST) with TGIT, with q statistics of 0.481, 0.472, and 0.452, respectively. Among the seven dominant cropping patterns, the soybean-based cropping pattern had the highest level of TGIT and BPI, followed by the maize-based cropping pattern, while the rice-based cropping pattern had the lowest level. All of the natural environmental, agri-economic and policy factors have a synergistic effect in contributing to the spatial heterogeneity of cropping patterns. Natural environmental factors determine the overall spatial distribution of cropping patterns in the NEC, while economic and policy factors combine to influence farmers' decisions, resulting in diverse regional cropping patterns. It is recommended that maize-soybean rotations such as Maize-Soybean Alternate Cropping (MSAC) and Maize-Soybean Rotational Cropping (MSRC) should be promoted, especially in the central and southern regions of the NEC, to meet agricultural market demand and stabilize soil productivity.

Recognition of outer membrane proteins using multiple feature fusion.

Introduction: Outer membrane proteins are crucial in maintaining the structural stability and permeability of the outer membrane. Outer membrane proteins exhibit several functions such as antigenicity and strong immunogenicity, which have potential applications in clinical diagnosis and disease prevention. However, wet experiments for studying OMPs are time and capital-intensive, thereby necessitating the use of computational methods for their identification. Methods: In this study, we developed a computational model to predict outer membrane proteins. The non-redundant dataset consists of a positive set of 208 outer membrane proteins and a negative set of 876 non-outer membrane proteins. In this study, we employed the pseudo amino acid composition method to extract feature vectors and subsequently utilized the support vector machine for prediction. Results and Discussion: In the Jackknife cross-validation, the overall accuracy and the area under receiver operating characteristic curve were observed to be 93.19% and 0.966, respectively. These results demonstrate that our model can produce accurate predictions, and could serve as a valuable guide for experimental research on outer membrane proteins.

Prediction of apoptosis protein subcellular location based on amphiphilic pseudo amino acid composition.

Introduction: Apoptosis proteins play an important role in the process of cell apoptosis, which makes the rate of cell proliferation and death reach a relative balance. The function of apoptosis protein is closely related to its subcellular location, it is of great significance to study the subcellular locations of apoptosis proteins. Many efforts in bioinformatics research have been aimed at predicting their subcellular location. However, the subcellular localization of apoptotic proteins needs to be carefully studied. Methods: In this paper, based on amphiphilic pseudo amino acid composition and support vector machine algorithm, a new method was proposed for the prediction of apoptosis proteins\x{2019} subcellular location. Results and Discussion: The method achieved good performance on three data sets. The Jackknife test accuracy of the three data sets reached 90.5%, 93.9% and 84.0%, respectively. Compared with previous methods, the prediction accuracies of APACC_SVM were improved.

Enantioselective analyses of chloroquine and hydroxychloroquine in rat liver microsomes through chiral liquid chromatography-tandem mass spectrometry.

An efficient, sensitive and selective liquid chromatography-tandem mass spectrometry (LC-MS/MS) chiral analysis method was established for determination of chloroquine and hydroxychloroquine enantiomers in rat liver microsomes. Effects of polysaccharide chiral stationary phases and basic additives on chiral separations of two analytes were discussed in detail. Amylose tris(3, 5-dimethylphenylcarbamate)-coated chiral stationary phase showed the best separation performance for them with acetonitrile-diethylamine-ethanol-diethylamine mixture (90:0.1:10:0.1, v/v/v/v) among four chiral stationary phases. Then, multiple reaction monitoring mode was selected as the data acquisition for determination of two pairs of enantiomers. The proposed LC-MS/MS chiral analysis method was validated in terms of linearity, accuracy, precision, and specificity. Good linearity with correlation coefficient over 0.998 was obtained in the concentration range of 0.05-5 µM. Limits of quantification for chloroquine and hydroxychloroquine enantiomers were 5.0 and 1.0 nM, respectively. The recoveries ranged from 81.14% to 111.09%. The intra-day and inter-day relative standard deviation were less than 6.5%. Moreover, concentrations of chloroquine and hydroxychloroquine enantiomers in rat liver microsomes were determined through the proposed LC-MS/MS analysis method. After incubated with rat liver microsomes for 10 min, the enantiomeric factor of hydroxychloroquine decreased from 0.50 to 0.45 (p < 0.001). In brief, our developed determination method for chloroquine and hydroxychloroquine enantiomers through LC-MS/MS spectrometry showed the characteristics of high-efficiency, fast speed, and very low detection limit, and would be greatly beneficial for screening and quantitation of them in biological matrices.

Stereochemistry of chiral pesticide uniconazole and enantioselective metabolism in rat liver microsomes.

In this work, stereochemistry of uniconazole enantiomers and their metabolism behaviors in rat liver microsomes have been researched. Significance analysis has been applied in data processing. Absolute configurations of uniconazole enantiomers were identified through vibrational circular dichroism spectroscopy. According to their elution order from the chiral column using the CO2-methanol (80:20, v/v) mixture, two eluted fractions were determined to be (R)-uniconazole and (S)-uniconazole, respectively. A high-efficient and sensitive LC-MS/MS chiral analysis method was established for investigating the metabolism of uniconazole enantiomers in rat liver microsomes. The metabolic half-life of (R)-uniconazole (38.7 min) in rat liver microsomes was half that of (S)-enantiomer (74.5 min), and maximum velocity of metabolism, Michaelis constant of metabolism as well as the intrinsic metabolic clearance of (R)-uniconazole were significantly higher than (S)-enantiomer (p < 0.05), which indicated that (R)-uniconazole was preferentially metabolized in rat liver microsomes. By the virtue of molecular docking, (R)-uniconazole exhibited a higher binding affinity to cytochrome CYP2D2 than (S)-enantiomer, which corroborated well with the metabolism results. This work will shed light on the risk assessment of uniconazole toward human health and the ecological environment.

Enhancing the Curie temperature of two-dimensional monolayer CrI3 by introducing I-vacancies and interstitial H-atoms.

The discovery of two-dimensional monolayer CrI3 provides a promising possibility for developing spintronic devices. However, the low Curie temperature is an obstacle for practical applications. Here, based on the consideration of the superexchange interaction of ferromagnetic coupling, we investigate the effect of introducing I-vacancies and interstitial H-atoms on the Curie temperature of monolayer CrI3 by using first-principles calculations and Monte Carlo simulations. Our theoretical conclusions show that the Curie temperature of Cr8I23 (CrI2.875), Cr8I22 (CrI2.75) and Cr8I24H (CrI3H0.125) significantly increases to 97.0, 82.5 and 112.4 K, respectively. Moreover, the magnetic moment of the Cr atom increases from 3.10 to 3.45 and 3.46µB in monolayers Cr8I23 and Cr8I22, respectively. We provide more alternative approaches to effectively enhance the Curie temperature of monolayer CrI3, which will help both theoretical and experimental researchers to directly predict the change in Curie temperature of CrI3 and its analogs through structural information.

Orientational Alignment of Oxygen Vacancies: Electric-Field-Inducing Conductive Channels in TiO2 Film to Boost Photocatalytic Conversion of CO2 into CO.

Oxide semiconductors are widely used in the photocatalytic fields, and introducing oxygen vacancies is an effective strategy to improve their photocatalytic efficiency. However, oxygen vacancies in the bulk often act as the recombination centers of electron-hole pairs, which accelerates the recombination of electron-hole pairs. In this paper, we propose a strategy of electric field treatment and apply it to a TiO2 film with oxygen vacancies to promote the photocatalytic efficiency. After treatment by an electric field, the conductive channels consisting of oxygen vacancies are formed in the TiO2 film, which greatly decreases the resistance by almost 6 × 103 times. The yield of CO can reach up to 1.729 mmol gcat-1 h-1, which is one of the best performances among the reported TiO2-based catalysts. This work provides an effective and feasible way for enhancing photocatalytic activity through an electric field, and this method is promising for wide use in the field of catalysis.

Study on the spatial distribution patterns of histone modifications in Hippo pathway genes.

Hippo pathway can regulate cell division, differentiation and apoptosis, and control the shape and size of organs. To study the distribution patterns of histone modifications of Hippo pathway genes in embryonic stem cells is helpful to understand the molecular regulation mechanism of histone modification and Hippo pathway on stem cell self-renewal. In this study, 19 genes of Hippo pathway including YAP, TAZ, LATS1/2, MST1 and SAV1, and eight histone modifications in embryonic stem cells were chosen to study the spatial distribution patterns of histone modifications. It was found that there were obvious type specificity and the location preference of target regions in the distributions of histone modifications, and H3K4me3 and H3K36me3 played the most important regulatory roles. Through the correlation analysis of histone modifications, a histone modification functional cluster composed of H3K4ac, H3K4me3, H3K9ac and H3K27ac was detected in YAP. In addition, the spatial distribution patterns of histone modifications in Hippo pathway genes were obtained, which provided a new theoretical reference for elucidating the mechanism of histone modifications regulating the gene expression of Hippo pathway, and for revealing the molecular regulatory mechanism of histone modifications affecting the self-renewal of embryonic stem cells by regulating the Hippo pathway.

Linc00426 accelerates lung adenocarcinoma progression by regulating miR-455-5p as a molecular sponge.

Increasing lines of evidence indicate the role of long non-coding RNAs (LncRNAs) in gene regulation and tumor development. Hence, it is important to elucidate the mechanisms of LncRNAs underlying the proliferation, metastasis, and invasion of lung adenocarcinoma (LUAD). We employed microarrays to screen LncRNAs in LUAD tissues with and without lymph node metastasis and revealed their effects on LUAD. Among them, Linc00426 was selected for further exploration in its expression, the biological significance, and the underlying molecular mechanisms. Linc00426 exhibits ectopic expression in LUAD tissues and cells. The ectopic expression has been clinically linked to tumor size, lymphatic metastasis, and tumor differentiation of patients with LUAD. The deregulation of Linc00426 contributes to a notable impairment in proliferation, invasion, metastasis, and epithelial-mesenchymal transition (EMT) in vitro and in vivo. Mechanistically, the deregulation of Linc00426 could reduce cytoskeleton rearrangement and matrix metalloproteinase expression. Meanwhile, decreasing the level of Linc00426 or increasing miR-455-5p could down-regulate the level of UBE2V1. Thus, Linc00426 may act as a competing endogenous RNA (ceRNA) to abate miR-455-5p-dependent UBE2V1 reduction. We conclude that Linc00426 accelerates LUAD progression by acting as a molecular sponge to regulate miR-455-5p, and may be a potential novel tumor marker for LUAD.

BIM deletion polymorphism predicts poor response to EGFR-TKIs in nonsmall cell lung cancer: An updated meta-analysis.

BACKGROUND: A germline deletion in BIM (B cell lymphoma-2-like 11) gene has been shown to impair the apoptotic response to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) in vitro but its impact on response to EGFR-TKIs in patients of nonsmall cell lung cancer (NSCLC) remains controversial. METHODS: Eligible literature were searched and screened. Objective response rate (ORR) and disease control rate (DCR) were extracted and aggregated with odds ratio (OR). Hazard ratio (HR) and 95% confidence interval (CI) for progression-free survival (PFS) and overall survival (OS) were extracted and aggregated based on random-effect model. RESULTS: Fourteen studies including 2694 NSCLC patients were eligible. Individuals harboring BIM deletion polymorphism had inferior ORR (OR = 0.49, 95% CI: 0.34-0.70, P < .001), inferior DCR (OR = 0.50, 95% CI: 0.30-0.84, P = .009). Patients with BIM deletion had shorter OS despite of the heterogeneity between countries (in subgroup of South Korea and Taiwan, HR = 1.34, 95% CI: 1.18-1.53, P < .001; in subgroup of other countries, HR = 2.43, 95% CI: 2.03-2.91, P < .001). The pooled analysis of PFS showed great heterogeneity (I = 79%). All the reported characteristics did not account for the heterogeneity. However, 2 subgroups could be obtained through sensitivity analysis. In one subgroup, patients with BIM deletion polymorphism had shorter PFS (HR = 2.03, 95% CI: 1.71-2.40, P < .001), while in the other subgroup, no significant difference was observed (HR = 0.92, 95% CI: 0.79-1.06, P = .25). CONCLUSION: NSCLC patients with BIM deletion polymorphism show poor ORR, DCR, and OS after EGFR-TKIs treatment. BIM deletion polymorphism indicates poor response to EGFR-TKIs, and it could be used as a predictor to identify those who would benefit from EGFR-TKIs in NSCLC patients.

Elimination of Autofluorescence in Archival Formaldehyde-Fixed, Paraffin-Embedded Bone Marrow Biopsies.

CONTEXT.­: High levels of autofluorescence in bone marrow tissue constitute a major obstacle to immunofluorescence analysis of bone marrow biopsies. OBJECTIVE.­: To present a simple, efficient method to eliminate autofluorescence in bone marrow biopsies. DESIGN.­: Autofluorescence of paraffin bone marrow tissues was examined in different hematologic disorders with confocal laser scanning microscopy. Strong autofluorescence was observed in primary myelofibrosis and acute leukemia with reticulin myelofibrosis in 488-nm and 561-nm channels. To eliminate autofluorescence, AutoFluo Quencher was used on bone marrow sections with different incubation times. The effects of AutoFluo Quencher on immunofluorescence analysis of bone marrow biopsies was tested using antibodies tagged with different fluorophores. RESULTS.­: AutoFluo Quencher thoroughly eliminated the strong autofluorescence of bone marrow but did not decrease the intensity of fluorophores, leaving the specific signals of target proteins clearly visible. CONCLUSIONS.­: This study presents a simple, efficient method to eliminate autofluorescence in bone marrow paraffin tissue, and it opens the way to better results in the immunofluorescence analysis of bone marrow biopsies.

Relationship Between DNA Methylation in Key Region and the Differential Expressions of Genes in Human Breast Tumor Tissue.

Breast cancer has a high mortality rate for females. Aberrant DNA methylation plays a crucial role in the occurrence and progression of breast carcinoma. By comparing DNA methylation differences between tumor breast tissue and normal breast tissue, we calculate and analyze the distributions of the hyper- and hypomethylation sites in different function regions. Results indicate that enhancer regions are often hypomethylated in breast cancer. CpG islands (CGIs) are mainly hypermethylated, while the flanking CGI (shores and shelves) is more easily hypomethylated. The hypomethylation in gene body region is related to the upregulation of gene expression, and the hypomethylation of enhancer regions is closely associated with gene expression upregulation in breast cancer. Some key hypomethylation sites in enhancer regions and key hypermethylation sites in CGIs for regulating key genes are, respectively, found, such as oncogenes ESR1 and ERBB2 and tumor suppressor genes FBLN2, CEBPA, and FAT4. This suggests that the recognizing methylation status of these genes will be useful for the diagnosis of breast cancer.

Large-scale transcriptome comparison of sunflower genes responsive to Verticillium dahliae.

BACKGROUND: Sunflower Verticillium wilt (SVW) is a vascular disease caused by root infection with Verticillium dahliae (V. dahlia). It is a serious threat to the yield and quality of sunflower. However, chemical and agronomic measures for controlling this disease are not effective. The selection of more resistant genotypes is a desirable strategy to reduce contamination. A deeper knowledge of the molecular mechanisms and genetic basis underlying sunflower Verticillium wilt is necessary to accelerate breeding progress. RESULTS: An RNA-Seq approach was used to perform global transcriptome profiling on the roots of resistant (S18) and susceptible (P77) sunflower genotypes infected with V. dahlia. Different pairwise transcriptome comparisons were examined over a time course (6, 12 and 24 h, and 2, 3, 5 and 10 d post inoculation). In RD, SD and D datasets, 1231 genes were associated with SVW resistance in a genotype-common transcriptional pattern. Moreover, 759 and 511 genes were directly related to SVW resistance in the resistant and susceptible genotypes, respectively, in a genotype-specific transcriptional pattern. Most of the genes were demonstrated to participate in plant defense responses; these genes included peroxidase (POD), glutathione peroxidase, aquaporin PIP, chitinase, L-ascorbate oxidase, and LRR receptors. For the up-regulated genotype-specific differentially expressed genes (DEGs) in the resistant genotype, higher average fold-changes were observed in the resistant genotype compared to those in the susceptible genotype. An inverse effect was observed in the down-regulated genotype-specific DEGs in the resistant genotype. KEGG analyses showed that 98, 112 and 52 genes were classified into plant hormone signal transduction, plant-pathogen interaction and flavonoid biosynthesis categories, respectively. Many of these genes, such as CNGC, RBOH, FLS2, JAZ, MYC2 NPR1 and TGA, regulate crucial points in defense-related pathway and may contribute to V. dahliae resistance in sunflower. CONCLUSIONS: The transcriptome profiling results provided a clearer understanding of the transcripts associated with the crosstalk between sunflower and V. dahliae. The results identified several differentially expressed unigenes involved in the hyper sensitive response (HR) and the salicylic acid (SA)/jasmonic acid (JA)-mediated signal transduction pathway for resistance against V. dahliae. These results are useful for screening resistant sunflower genotypes.

Predicting gene expression level by the transcription factor binding signals in human embryonic stem cells.

The transcription factor (TF) binding signals play important role in the control of gene expression. In this study, to elucidate the relationship between the transcription factor binding signals and gene expression, we firstly analyze the distributions of 57 kinds of TFs' binding signals in human H1 embryonic stem cells. Their distributions in highly and lowly expressed genes are further compared. On this basis, a statistic model of predicting gene expression level is constructed by using 57 kinds of transcription factor synthetic indexes (TFSIs). Then, the TF's Down-regulated and Up-regulated genes are predicted and the statistics significance is estimated by one-sided Kolmogorov-Smirnov test. Based on the stepwise regression analysis, the "optimal" TFSIs are selected out, and the better results for predicting the expression level of genes with high CpG content promoters (HCPs) and low CpG content promoters (LCPs) are obtained.

Gene expression classification using epigenetic features and DNA sequence composition in the human embryonic stem cell line H1.

Epigenetic factors are known to correlate with gene expression in the existing studies. However, quantitative models that accurately classify the highly and lowly expressed genes based on epigenetic factors are currently lacking. In this study, a new machine learning method combines histone modifications, DNA methylation, DNA accessibility, transcription factors, and trinucleotide composition with support vector machines (SVM) is developed in the context of human embryonic stem cell line (H1). The results indicate that the predictive accuracy will be markedly improved when the epigenetic features are considered. The predictive accuracy and Matthews correlation coefficient of the best model are as high as 95.96% and 0.92 for 10-fold cross-validation test, and 95.58% and 0.92 for independent dataset test, respectively. Our model provides a good way to judge a gene is either highly or lowly expressed gene by using genetic and epigenetic data, when the expression data of the gene is lacking. And a web-server GECES for our analysis method is established at http://202.207.14.87:8032/fuwu/GECES/index.asp, so that other scientists can easily get their desired results by our web-server, without going through the mathematical details.

Association analysis between the distributions of histone modifications and gene expression in the human embryonic stem cell.

It is well known that histone modifications are associated with gene expression. In order to further study this relationship, 16 kinds of Chip-seq histone modification data and mRNA-seq data of the human embryonic stem cell H1 are chosen. The distributions of histone modifications in the regions flanking transcription start sites (TSSs) for highly expressed and lowly expressed genes are computed, respectively. And four types of distributions of histone modifications in regions flanking TSSs and the spatial patterning of the correlations between histone modifications and gene expression are detected. Our results suggest that the correlations between the regions overlapped by peaks are higher than the non-overlapped ones for each histone modification. In addition, to obtain the effect of the cooperative action of histone modification on gene expression, five histone modification clusters are found in highly expressed and lowly expressed genes, histone modification and gene expression interaction network is constructed. To further explore which region is the main target region for the specific histone modification, the human genes are divided into five functional regions. The results indicate that histone modifications are mostly located in the promoters of highly expressed genes versus the exons of lowly expressed genes, and exons have a smaller range of normalized tag counts than other gene elements in the two groups of genes. Finally, the type specificity and regional bias of histone modifications for 11 key transcription factor genes regulating the stem cell renewal are analyzed.

Discrimination of membrane transporter protein types using K-nearest neighbor method derived from the similarity distance of total diversity measure.

Membrane transporters play crucial roles in the fundamental cellular processes of living organisms. Computational techniques are very necessary to annotate the transporter functions. In this study, a multi-class K nearest neighbor classifier based on the increment of diversity (KNN-ID) was developed to discriminate the membrane transporter types when the increment of diversity (ID) was introduced as one of the novel similarity distances. Comparisons with multiple recently published methods showed that the proposed KNN-ID method outperformed the other methods, obtaining more than 20% improvement for overall accuracy. The overall prediction accuracy reached was 83.1%, when the K was selected as 2. The prediction sensitivity achieved 76.7%, 89.1%, 80.1% for channels/pores, electrochemical potential-driven transporters, primary active transporters, respectively. Discrimination and comparison between any two different classes of transporters further demonstrated that the proposed method is a potential classifier and will play a complementary role for facilitating the functional assignment of transporters.

[Effect of Gli1 gene silencing on proliferation of K562 cells and its mechanisms].

OBJECTIVE: To investigate the effect of Gli1 gene silencing by RNA interference (RNAi) on proliferation of K562 cells and its mechanisms. METHODS: The small interference RNA (siRNA) was synthesized in vitro. K562 cells were transfected with Gli1 siRNA by the way of lipofection (lipofectamine 2000). Non-specific siRNA transfected cells were used as control. Transfection efficiencies of different siRNA concentrations were detected by flow cytometry and the best siRNA concentration was selected. The silencing effect of siRNA was demonstrated by real time PCR and Westem blot analysis. Cell proliferation was measured by MTT method, cell cycle by PI assay, c-myc and p21 mRNA level was detected by real time PCR analysis. RESULTS: Transfection efficiency of siRNA was increased in a dose-dependent manner when siRNA concentration was below 200 pmol, and the highest transfection efficiency reached (80.11 ± 5.63)%. Both the mRNA and protein level of Gli1 was down-regulated in Gli1 specific siRNA group, the mRNA level was (52.60 ± 3.57)% of that of control group after 24 h (t = 20.33, P < 0.01) and the protein level was (79.31 ± 5.58)% of that of control group after 48 h (t = 6.54, P < 0.01). The cell proliferation rate in Gli1 siRNA group was (94.41 ± 3.58)% (t = 2.40, P = 0.05) and (90.22 ± 3.34)% (t = 4.37, P < 0.01) of that of control group after 24 h and 48 h, respectively. G(2)/M cell cycle arrest was observed, the mRNA level of c-myc was down-regulated while p21 was up-regulated in Gli1 siRNA group after 24 h and 48 h (P < 0.05). CONCLUSIONS: Targeted silencing of Gli1 gene by RNAi inhibits the proliferation of K562 cells, which acts through the down-regulation of c-myc and up-regulation of p21 expression.