Journal: Bioinformatics Advances

Article Title: easyEWAS: a flexible and user-friendly R package for epigenome-wide association study

doi: 10.1093/bioadv/vbaf026

Figure Lengend Snippet: The workflow chart of the easyEWAS. Abbreviations: DNAm, DNA methylation; DMP, differentially methylated position; DMR, differentially methylated region; GLM, general linear model; LMM, linear mixed-effects model; CoxPH, Cox proportional hazards model; QQ: quantile–quantile; Boot.CI, bootstrap-derived CI; KEGG, Kyoto Encyclopedia of Genes and Genomes.

Article Snippet: In conclusion, easyEWAS is an R package that can be easily integrated into various DNA methylation microarrays detected by Illumina HumanMethylation Bead Chip, significantly enhancing the accessibility of EWAS.

Techniques: DNA Methylation Assay, Methylation, Derivative Assay

Journal: Bioinformatics Advances

Article Title: easyEWAS: a flexible and user-friendly R package for epigenome-wide association study

doi: 10.1093/bioadv/vbaf026

Figure Lengend Snippet: The function names and their corresponding functionalities within the easyEWAS package.

Article Snippet: In conclusion, easyEWAS is an R package that can be easily integrated into various DNA methylation microarrays detected by Illumina HumanMethylation Bead Chip, significantly enhancing the accessibility of EWAS.

Techniques: DNA Methylation Assay, Methylation, Biomarker Discovery

Journal: Nature Communications

Article Title: SLC13A3 is a major effector downstream of activated β-catenin in liver cancer pathogenesis

doi: 10.1038/s41467-024-51860-2

Figure Lengend Snippet: a , b mRNA expression of TBX3 , GLUL , and SLC13A3 , as well as protein levels of β-catenin and SLC13A3 in CTNNB1 -overexpressing or CTNNB1 -knockdown HCC cells. Huh7 and HLF cells were transfected with pT3-EF1αH plasmid (empty vector, EV , gray) or pT3-EF1αH plasmid containing ΔN90-β-catenin mutant fragment ( CTNNB1 , red). HepG2 and SNU398 cells were infected with shRNA lentivirus using pLKO.1 plasmid containing either scramble shRNA (negative control shRNA, sh NC , gray) or sh CTNNB1 (blue) sequences. Cells were collected at 24 h for qRT-PCR and 48 h for western blot. The experiments were performed three times on different days. Each western blot represented one biological replicate (two technical repeats per group). Data are presented as the mean ± SEM ( n = 3 independent experiments). Statistical analysis was performed using two-tailed Student’s t test. c Luciferase reporter assay for the identification of β-catenin binding sites in the SLC13A3 gene promoter region (~1.0 kb from transcription start site, TSS). A series of fragments in the SLC13A3 promoter region were schematized. HEK293T cells were transfected with the respective promoter plasmid, pCMV-renilla, and EV - or CTNNB1 -overexpressing plasmid for 24 h. Data are presented as the mean ± SEM ( n = 3 independent experiments). Statistical analysis was performed using one-way ANOVA test. d Chromatin immunoprecipitation (ChIP)-PCR detection of the SLC13A3 promoter. DNA was isolated by anti-β-catenin antibody (orange), anti-TCF4 antibody (blue), or negative control IgG (gray). Input DNA which equaled 10% total DNA samples prior to immunoprecipitation was used as positive control. DNAs were respectively amplified using SLC13A3 promoter primers#1 (for binding site 1), #2 (for binding site 2), and #3 (spans −2100 to −2081 bp upstream of the TSS), as well as negative GAPDH primers and positive MYC primers. Data are presented as the mean ± SEM ( n = 3 independent experiments). Statistical analysis was performed using two-tailed Student’s t test. e In vitro EMSA analysis of TCF4 protein binding with two putative β-catenin binding sites in HepG2 nuclear extracts. The protein and DNA interactions were abolished by adding unlabeled wild-type probes, but could not be abrogated by mutant probes. Left panel: Adding anti-TCF4 antibody resulted in a supershift band to TCF4 protein. Right panel: No supershift band after adding anti-TCF4 antibody. SLC13A3 probe #1 was for the bind site 1, and SLC13A3 probe #2 was for the bind site 2. Each experiment was independently repeated three times. f Relative intracellular levels of malate, succinate, and fumarate. The data were obtained from the untargeted metabolomics in CTNNB1 -overexpressing Huh7 cells (red) and CTNNB1 -knockdown HepG2 cells (blue). Data are presented as the mean ± SEM ( n = 6 independent experiments). Statistical analysis was performed using two-tailed Student’s t test. Source data are provided as a Source Data file.

Article Snippet: HepG2-sh NC and -sh SLC13A3 cells were seeded into 10 cm dishes and then cultured for 24 h. MeDIP assays were performed using a commercial methylated DNA immunoprecipitation (MeDIP) ChIP kit (ab117135, Abcam, Cambridge, MA).

Techniques: Expressing, Knockdown, Transfection, Plasmid Preparation, Mutagenesis, Infection, shRNA, Negative Control, Quantitative RT-PCR, Western Blot, Two Tailed Test, Luciferase, Reporter Assay, Binding Assay, Chromatin Immunoprecipitation, Isolation, Immunoprecipitation, Positive Control, Amplification, In Vitro, Protein Binding

Journal: Nature Communications

Article Title: SLC13A3 is a major effector downstream of activated β-catenin in liver cancer pathogenesis

doi: 10.1038/s41467-024-51860-2

Figure Lengend Snippet: a Cellular accumulation of 13 C 2 , 15 N-glutathione (GSH) in HEK293 cells. HEK293- SLC13A3 and HEK- EV cells were incubated with isotope-labeled GSH at concentrations from 10 nM to 10 mM for 20 min. Saturable GSH uptake was calculated using the difference of GSH accumulation in HEK293- SLC13A3 and HEK- EV cells. The data were fit to a Michaelis–Menten equation. Data are presented as the mean ± SEM for a representative experiment (n = 3 independent experiments). b Cellular uptake of 13 C 2 , 15 N-glutathione (GSH) (5 and 10 μM) was significantly higher in HEK- SLC13A3 cells (red) compared to HEK293- EV cells (gray). Cells were incubated in the uptake buffer containing GSH for 20 min. Data are presented as the mean ± SEM ( n = 3 independent experiments). Statistical analysis was performed using two-tailed Student’s t test. c Representative western blots of SLC13A3, SLC7A5, and SLC3A2 in HepG2 and SNU398 cell. Stably control (sh NC ) and SLC13A3 -knockdown (sh#1) cells were established by infection with shRNA lentivirus. Blots were representative of three independent experiments (two replicates per group for each experiment). d Cellular uptake of 2 mM leucine in HepG2 and SNU398 cells. Stability control (sh NC , gray) and SLC13A3 -knockdown cells (sh#1, blue) were incubated with 2 mM leucine for 20 min. Data are presented as the mean ± SEM ( n = 3 independent experiments). Statistical analysis was performed using two-tailed Student’s t test. e qRT-PCR and western blot analyses of MYC expression in HepG2 and SNU398 cells. Stably control (sh NC , gray) and SLC13A3 -knockdown (sh#1, blue and sh#2, dark blue) cells were established using respective shRNA lentivirus. Data are presented as the mean ± SEM ( n = 3 independent experiments). Statistical analysis was performed using one-way ANOVA test. Blots were representative of three independent experiments. f Methyl-DNA immunoprecipitation (MeDIP)-PCR detection of the MYC promoter methylation. DNA was immunoprecipitated with 5-methylcytosine antibody (blue) or IgG (gray), and purified according to the manufacturer’s protocol. Input DNA prior to immunoprecipitation was used as the positive control. Precipitated and input DNAs were amplified using primers for the CpG islands in the MYC promoter region, and PCR products were visualized by gel electrophoresis. Data are presented as the mean ± SEM ( n = 3 independent experiments). Statistical analysis was performed using two-tailed Student’s t test. g Mouse study design. Mice were given a leucine-deficient diet (Leu-) or leucine-supplemented drinking water (1.5% leucine in drinking water, Leu+). Liver tumor was induced by hydrodynamic tail vein injection (HTVi) of c-Met/β-catenin plasmids or AKT/β-catenin plasmids. h Mice were euthanized when they developed a high burden of liver tumors. The log-rank test was used to compare overall survival between groups (Gray, control; Blue, Leu+; Orange, Leu-). ns, non-statistically significant. i , j Representative gross liver images, as well as H&E (hematoxylin-eosin) and Ki67 IHC staining images. Images were representative shown out of 6 independent mice per group. Scale bar, 200 μm. Source data are provided as a Source Data file.

Article Snippet: HepG2-sh NC and -sh SLC13A3 cells were seeded into 10 cm dishes and then cultured for 24 h. MeDIP assays were performed using a commercial methylated DNA immunoprecipitation (MeDIP) ChIP kit (ab117135, Abcam, Cambridge, MA).

Techniques: Incubation, Labeling, Two Tailed Test, Western Blot, Stable Transfection, Control, Knockdown, Infection, shRNA, Quantitative RT-PCR, Expressing, Immunoprecipitation, Methylated DNA Immunoprecipitation, Methylation, Purification, Positive Control, Amplification, Nucleic Acid Electrophoresis, Injection, Immunohistochemistry

Journal: Chinese Medical Journal Pulmonary and Critical Care Medicine

Article Title: Omics approaches in asthma research: Challenges and opportunities

doi: 10.1016/j.pccm.2024.02.002

Figure Lengend Snippet: Public available genomics datasets of asthma.

Article Snippet: DNA methylation is commonly assessed using DNA methylation chips, such as the Infinium Human Methylation 450K beadchip, which covers around 450,000 CpG sites, or the Infinium Methylation EPIC Beadchip, which extends the coverage to approximately 850,000 CpG sites across the entire genome.

Techniques: Gene Expression, Microarray, Infection, DNA Methylation Assay