pgg  (MedChemExpress)

Journal: Ultrasonics Sonochemistry

Article Title: Process optimization and antioxidant activity of white peony root components extracted via ultrasound-assisted deep eutectic solvents

doi: 10.1016/j.ultsonch.2025.107685

Figure Lengend Snippet: A) HPLC chromatogram of white peony root. The numbers represented the following compounds: 1. Catechin (Cat), 2. Albiflorin (Alb), 3. Paeoniflorin (PF), and 4. 1,2,3,4,6-penta- O -galloyl -β -ᴅ-glucose (PGg). B) Effect of different DESs and conventional solvents on the extraction yield of WPR via UAE. The error bars indicate the mean (±) SD ( n = 3).

Article Snippet: The reference compounds, which comprised catechins (Cat), paeoniflorin (PF), albiflorin (Alb), and 1,2,3,4,6-penta-O-galloyl -β - d -glucose (PGg), were purchased from Shanghai Yuanye Bio-Technology Co., Ltd. (Shanghai, China).

Techniques: Extraction

Journal: Cancer Communications

Article Title: RNA m 1 A methyltransferase TRMT61A promotes colorectal tumorigenesis by enhancing ONECUT2 mRNA stability and is a potential therapeutic target

doi: 10.1002/cac2.70070

Figure Lengend Snippet: RNA m 1 A modifications played essential roles in CRC. (A) RNA m 1 A modification level was upregulated in CRC compared to paired adjacent normal tissues. The Y‐axis represents the ratio of m 1 A signal level to adenosine (A) signal level, and a two‐tailed Wilcoxon matched‐pairs signed rank test was performed for comparison. The black dotted line represents the median value. (B) CRISPR‐Cas9 screening in CRC cell lines (HCT116 and POP92) identifies TRMT61A as an essential gene for CRC survival. (C) Overexpression of TRMT61A increased global m 1 A levels, and depleted TRMT61A decreased global m 1 A levels in CRC cell lines HCT116 and RKO by dot blotting. MB staining was conducted as the loading control. (D) Correlation between TRMT61A mRNA expression and global mRNA m 1 A contents in Beijing CRC tissues. (E) The TRMT61A and TRMT6 mRNA expression in 150 pairs of primary CRCs and adjacent normal tissues from the Beijing cohort and in 50 pairs of primary CRCs and adjacent normal tissues from the TCGA cohort. (F) WB analysis of TRMT6 and TRMT61A from paired CRC and adjacent normal tissues from the Beijing cohort ( n = 10). (G) Representative IHC images of TRMT61A protein expression in a tissue microarray containing 28 pairs of CRC and adjacent normal tissues from the Hong Kong cohort. The staining score of TRMT61A was quantified accordingly. (H) OS curves of the 179 CRC patients with complete survival data from the Hong Kong cohort (left panel), and multivariable analysis of risk factors for OS (right panel). (I) High TRMT61A mRNA expression predicted worse OS in GSE29623 datasets (left panel), multivariable analysis of risk factors for OS (right panel). Abbreviations: CRC, colorectal cancer; TRMT61A, TRNA methyltransferase 61A; m 1 A, N1‐methyladenosine; LC‐MS/MS, Liquid Chromatography‐Tandem Mass Spectrometry; sgRNA, single‐guide RNA; MB, Methyl blue; IHC, Immunohistochemistry; TCGA, The Cancer Genome Atlas; OS, Overall survival; HR, Hazard Ratio; CI, Confidence Interval.

Article Snippet: Cell viability was measured at different time points using CellTiter 96 AQueousOne Solution Cell Proliferation Assay (#G3581, Promega) after transfection with sgRNA, siRNA or plasmid DNA, or after treatment with the TRMT61A protein inhibitor pentagalloylglucose (PGG) (5 μmol/L, #HY‐N0527, MedChemExpress).

Techniques: Modification, Two Tailed Test, Comparison, CRISPR, Over Expression, Staining, Control, Expressing, Microarray, Liquid Chromatography with Mass Spectroscopy, Liquid Chromatography, Mass Spectrometry, Immunohistochemistry

Journal: Cancer Communications

Article Title: RNA m 1 A methyltransferase TRMT61A promotes colorectal tumorigenesis by enhancing ONECUT2 mRNA stability and is a potential therapeutic target

doi: 10.1002/cac2.70070

Figure Lengend Snippet: TRMT61A played an oncogenic role in CRC. (A) Schematic illustration of generating Trmt61a conditional knockout mice ( Trmt61a flox/flox ) using CRISPR/Cas9‐mediated gnome engineering strategy. (B) A colon‐specific Trmt61a conditional knockout ( Trmt61a cko ) mouse model was generated by crossing Trmt61a flox/flox mice with Cdx2‐Cre Ert2 mice. Subsequently, Trmt61a cko mice was further crossbred with Apc Min/+ mice to facilitate CRC formation. The knockout of TRMT61A in colon tissues was confirmed by WB, n = 16 mice per group. (C) Representative images of colon tissues (distal to the ileocecal junction, excluding the cecum) of Apc Min/+ and Apc Min/+ Trmt61a cko mice. n = 16 mice per group. Colon tumor number and tumor volume were calculated. (D and E) H&E, Ki‐67, and TUNEL staining of colon tumors derived from Apc Min/+ and Apc Min/+ Trmt61a cko mice. n = 5 mice per group. (F and G) Knockout of TRMT61A significantly suppressed cell proliferation and colony formation, whereas overexpression of WT but not D181A TRMT61A exerted the opposite effect. (H) The efficiency of tumor colony formation in CRC patient‐derived organoids was calculated after 7 days of culture. n = 3 per group. Scattered dots represent individual samples, while horizontal bars indicate mean values.Abbreviations: CRC, colorectal cancer; TRMT61A, TRNA methyltransferase 61A; CRISPR, clustered regularly interspaced short palindromic repeats; TUNEL, Terminal deoxynucleotidyl transferase dUTP nick end labeling; WB, Western blot; WT, wild‐type; sgRNA, single guide RNA; EV, empty vector; CTRL, control.

Article Snippet: Cell viability was measured at different time points using CellTiter 96 AQueousOne Solution Cell Proliferation Assay (#G3581, Promega) after transfection with sgRNA, siRNA or plasmid DNA, or after treatment with the TRMT61A protein inhibitor pentagalloylglucose (PGG) (5 μmol/L, #HY‐N0527, MedChemExpress).

Techniques: Knock-Out, CRISPR, Generated, TUNEL Assay, Staining, Derivative Assay, Over Expression, Western Blot, Plasmid Preparation, Control

Journal: Cancer Communications

Article Title: RNA m 1 A methyltransferase TRMT61A promotes colorectal tumorigenesis by enhancing ONECUT2 mRNA stability and is a potential therapeutic target

doi: 10.1002/cac2.70070

Figure Lengend Snippet: Depletion of TRMT61A expression inhibited CRC growth in vivo. (A) HCT116 ( n = 10) and RKO ( n = 8) xenografts expressing sgTRMT61A showed significantly stunted growth compared to controls. (B) Representative images of Ki‐67‐positive cells and quantitative data analysis. (C) Representative images of TUNEL‐positive cells and quantitative data analysis. (D) Schematic showing the construction VNP‐siRNA. (E) VNP‐ siTRMT61A treatment significantly suppressed the growth of HCT116 (VNP‐ siCTRL, n = 10; VNP‐ siTRMT61A, n = 12) and RKO (VNP‐ siCTRL, n = 12; VNP‐ siTRMT61A, n = 10) xenografts. Two samples from the VNP‐ siCTRL group of HCT116 xenografts and the VNP‐ siTRMT61A group of RKO xenografts did not develop tumors prior to the VNP treatment and were therefore excluded from the analysis. (F) Representative images of Ki‐67‐positive cells and quantitative data analysis. (G) Representative images of TUNEL‐positive cells and quantitative data analysis. The red arrows indicate TUNEL‐positive apoptotic cells. To quantify IHC staining, at least 5 fields per slide and 5 slides per animal were counted at 200× magnification. Scattered dots represent individual samples, while horizontal bars indicate mean values. Abbreviations: CRC, colorectal cancer; TRMT61A, TRNA methyltransferase 61A; VNP, Vesicle‐like poly (lactic‐co‐glycolic acid)‐based nanoparticle; TUNEL, Terminal deoxynucleotidyl transferase dUTP nick end labeling; IHC, immunohistochemistry; sgRNA, single guide RNA; EV, empty vector; CTRL, control.

Article Snippet: Cell viability was measured at different time points using CellTiter 96 AQueousOne Solution Cell Proliferation Assay (#G3581, Promega) after transfection with sgRNA, siRNA or plasmid DNA, or after treatment with the TRMT61A protein inhibitor pentagalloylglucose (PGG) (5 μmol/L, #HY‐N0527, MedChemExpress).

Techniques: Expressing, In Vivo, TUNEL Assay, Immunohistochemistry, Plasmid Preparation, Control

Journal: Cancer Communications

Article Title: RNA m 1 A methyltransferase TRMT61A promotes colorectal tumorigenesis by enhancing ONECUT2 mRNA stability and is a potential therapeutic target

doi: 10.1002/cac2.70070

Figure Lengend Snippet: ONECUT2 is a critical downstream target of TRMT61A. (A) Flow chart of m 1 A‐seq, RNA‐seq, and Ribo‐seq. (B) The normalized distribution of m 1 A peaks and identified m 1 A motifs in HCT116 cells. (C) A heat map of the m 1 A levels of potential TRMT61A downstream targets identified by m 1 A‐seq. Lower m 1 A levels are depicted in blue, while higher m 1 A levels are shown in red. Each row represents a single transcript, and each column corresponds to a sample. n = 434. (D) Top panel: Differential RNA expression of TRMT61A‐regulated targets ( n = 434) versus all other genes after TRMT61A depletion. Bottom panel: Statistically significant expression changes ( P adj < 0.01) in 55 TRMT61A target transcripts after TRMT61A depletion. A two‐tailed Mann‐Whitney U test was performed to compare the groups. (E) The expression levels of the top 10 TRMT61A target genes (showing downregulated mRNA expression following TRMT61A depletion) were compared between primary CRCs and adjacent normal tissues using the TCGA‐CRC dataset. (F) m 1 A‐seq data were displayed as read alignment tracks and coverage plots, showing read distribution across the genome and enrichment in IP versus input samples. (G) MeRIP‐qPCR analysis of m 1 A level in the select mRNAs from HCT116 and RKO cells with or without TRMT61A knockout. (H) qPCR analysis of ONECUT2 mRNA expression in HCT116 and RKO cells with or without TRMT61A knockout (left), and with or without TRMT61A overexpression (right). (I) Protein expressions of TRMT61A and ONECUT2 were evaluated by WB. (J) Evaluation of ONECUT2 mRNA stability in HCT116 and RKO cells with or without TRMT61A knockout. (K) Evaluation of ONECUT2 mRNA stability in HCT116 and RKO cells with or without TRMT61A overexpression. (L) The RIP‐qPCR analysis confirmed that TRMT61A‐WT, but not TRMT61A‐bMUT (which failed to bind to TRMT6), was binding to the ONECUT2 mRNA. (M) The relative luciferase activity of the ONECUT2 m 1 A‐WT or m 1 A‐MUT reporter in HCT116 and RKO cells was measured upon overexpression of TRMT61A‐WT or TRMT61A D181A . (N) Cell proliferation assays of HCT116 and RKO. (O) ONECUT2 mRNA expression in 150 pairs of primary CRCs and adjacent normal tissues from the Beijing cohort by qPCR. (P) Correlation between TRMT61A and ONECUT2 mRNA expressions in primary CRCs from the Beijing cohort. Abbreviations: m 1 A‐seq, m 1 A‐sequencing; RNA‐seq, RNA sequencing; CRC, colorectal cancer; TRMT61A, TRNA methyltransferase 61A; WB, Western blot; WT, wild‐type; MUT, mutant; ONECUT2, one cut homeobox 2; RIP‐qPCR, RNA immunoprecipitation‐qPCR; sgRNA, single guide RNA; EV, empty vector; CTRL, control.

Article Snippet: Cell viability was measured at different time points using CellTiter 96 AQueousOne Solution Cell Proliferation Assay (#G3581, Promega) after transfection with sgRNA, siRNA or plasmid DNA, or after treatment with the TRMT61A protein inhibitor pentagalloylglucose (PGG) (5 μmol/L, #HY‐N0527, MedChemExpress).

Techniques: RNA Sequencing, RNA Expression, Expressing, Two Tailed Test, MANN-WHITNEY, Knock-Out, Over Expression, Binding Assay, Luciferase, Activity Assay, Sequencing, Western Blot, Mutagenesis, RNA Immunoprecipitation, Plasmid Preparation, Control

Journal: Cancer Communications

Article Title: RNA m 1 A methyltransferase TRMT61A promotes colorectal tumorigenesis by enhancing ONECUT2 mRNA stability and is a potential therapeutic target

doi: 10.1002/cac2.70070

Figure Lengend Snippet: TRMT61A promotes the m 1 A‐ONECUT2‐MAPK/ERK axis. (A) Venn diagram showing overlaps between gene sets analyzed by GSEA with RNA‐seq and Ribo‐seq data. (B) GSEA enrichment plots of MAPK/ERK signaling. (C) WB results showed depletion of TRMT61A inactivated MAPK/ERK signaling. (D) WB was performed in HCT116 and RKO cells with indicated treatments. (E) Cell viability assays were performed in CRC cells with indicated treatments. (F) Integrative analysis of RNA‐seq data from HCT116 cells with or without sgTRMT61A , RNA‐seq data from HCT116 cells with or without siONECUT2 , and genes involved in the MAPK/ERK pathway. (G) qPCR analysis of ITGB1 and SOS1 mRNA expressions in HCT116 and RKO cells with indicated treatments. (H) ONECUT2, SOS1 and ITGB1 protein expressions were evaluated by WB. (I) ChIP‐PCR confirmed the binding of ONECUT2 to the promoter region of SOS1 . PCR amplification was performed using total DNA fragments (input) and DNA fragments immunoprecipitated by anti‐ONECUT2 or anti‐IgG. (J) The luciferase activity of the reporter containing the wild‐type or mutant SOS1 promoter region was measured in cells with or without ONECUT2 depletion. (K) WB was performed in colon tumors of Apc Min/+ and Apc Min/+ Trmt61a cko mice. Scattered dots represent individual samples, while horizontal bars indicate mean values. Abbreviations: ChIP‐PCR, Chromatin immunoprecipitation‐PCR; CRC, colorectal cancer; TRMT61A, TRNA methyltransferase 61A; ONECUT2, one cut homeobox 2; MAPK/ERK, mitogen‐activated protein kinase/extracellular signal‐regulated kinase; SOS1, son of sevenless homolog 1; ITGB1, Integrin Subunit Beta 1; WB, Western blot; sgRNA, single guide RNA; EV, empty vector; CTRL, control.

Article Snippet: Cell viability was measured at different time points using CellTiter 96 AQueousOne Solution Cell Proliferation Assay (#G3581, Promega) after transfection with sgRNA, siRNA or plasmid DNA, or after treatment with the TRMT61A protein inhibitor pentagalloylglucose (PGG) (5 μmol/L, #HY‐N0527, MedChemExpress).

Techniques: RNA Sequencing, Binding Assay, Amplification, Immunoprecipitation, Luciferase, Activity Assay, Mutagenesis, Chromatin Immunoprecipitation, Western Blot, Plasmid Preparation, Control

Journal: Cancer Communications

Article Title: RNA m 1 A methyltransferase TRMT61A promotes colorectal tumorigenesis by enhancing ONECUT2 mRNA stability and is a potential therapeutic target

doi: 10.1002/cac2.70070

Figure Lengend Snippet: PGG is a potent inhibitor of TRMT61A. (A) Flow chart of structure‐based Virtual Screening. Docking models were developed based on the TRMT61A crystal structure. (B) Effects of top 30 compounds (5 µmol/L) on the viability of HCT116 and RKO cells. * P < 0.05, ** P < 0.01, *** P < 0.001. (C) 2D and 3D ligand interaction diagrams for PGG and TRMT61A. (D) CRC cells were treated with PGG at different dosages for 72 h. (E) Effects of PGG (5 µmol/L, 72 h) on CRC proliferation in HCT116 and RKO by cell viability assays. (F) Effects of PGG (5 µmol/L, 96 h) on CRC cell proliferation in HCT116 by cell counting assays. (G) RNA m 1 A dot blotting. MB serves as loading control. (H) ONECUT2 mRNA expression in CRC cells with or without PGG (5 µmol/L) treatment for 48 h. (I) ONECUT2 protein expression in CRC cells with or without PGG treatment for 48 h. (J) Plasmids expressing TRMT61A‐WT or TRMT61A‐7A were generated and confirmed by Sanger sequencing. (K) DARTS of cells treated with indicated PGG. (L) CETSA of cells treated with 5 µmol/L PGG for 30 min. (M) Recombinant human TRMT61A protein was generated in E. coli and confirmed by either Coomassie brilliant blue or WB staining as indicated by red arrows. MST analysis was conducted to measure the dissociation kinetics of PGG towards recombinant human TRMT61A protein (residues 2‐289). Scattered dots represent individual samples, while horizontal bars indicate mean values. Abbreviations: PGG, Pentagalloylglucose; MB, Methylene blue; DARTS, Drug affinity responsive target stability; MST, MicroScale thermophoresis; CETSA, Cellular thermal shift assay; CRC, colorectal cancer; TRMT61A, TRNA methyltransferase 61A; WB, Western blot; WT, wild‐type; sgRNA, single guide RNA; EV, empty vector; CTRL, control.

Article Snippet: Cell viability was measured at different time points using CellTiter 96 AQueousOne Solution Cell Proliferation Assay (#G3581, Promega) after transfection with sgRNA, siRNA or plasmid DNA, or after treatment with the TRMT61A protein inhibitor pentagalloylglucose (PGG) (5 μmol/L, #HY‐N0527, MedChemExpress).

Techniques: Cell Counting, Control, Expressing, Generated, Sequencing, Recombinant, Staining, Microscale Thermophoresis, Thermal Shift Assay, Western Blot, Plasmid Preparation

Journal: Cancer Communications

Article Title: RNA m 1 A methyltransferase TRMT61A promotes colorectal tumorigenesis by enhancing ONECUT2 mRNA stability and is a potential therapeutic target

doi: 10.1002/cac2.70070

Figure Lengend Snippet: PGG displays potent anti‐CRC efficacy. (A) HCT116 and RKO xenograft models with or without PGG treatment. One sample from the PBS group (HCT116 and RKO xenografts) and the PGG group (RKO xenografts) did not develop tumors prior to the PGG treatment and were therefore excluded from the analysis. (B) Schematic illustration of AOM/DSS‐induced CRC model with or without PGG treatment. (C) Representative macroscopic images of colons from mice and representative images of colon tissues of mice. Arrows indicate tumors. (D) Colon tumor number and tumor size were calculated. (E) H&E, Ki‐67, and TUNEL staining of colon tumors in AOM/DSS‐treated mice. The red arrows indicate TUNEL‐positive apoptotic cells. (F) Mice body weight of mice was monitored after PGG treatment. Mice colon length was determined upon sacrifice. (G) Liver or kidney function tests using blood samples of AOM/DSS‐treated mice. The pink box indicates the reference interval of each indicator. (H) WB analysis of tumor tissues. (I) Tumor colony formation efficiency of CRC patient‐derived organoids with or without PGG (5 or 10 µmol/L) treatment. (J) Schematic representations of the role of TRMT61A in CRC. Abbreviations: AOM/DSS, Azoxymethane/ Dextran sodium sulfate; H&E, Hematoxylin and Eosin; TUNEL, Terminal deoxynucleotidyl transferase dUTP nick end labeling; CRC, colorectal cancer; TRMT61A, TRNA methyltransferase 61A; WB, Western blot.

Article Snippet: Cell viability was measured at different time points using CellTiter 96 AQueousOne Solution Cell Proliferation Assay (#G3581, Promega) after transfection with sgRNA, siRNA or plasmid DNA, or after treatment with the TRMT61A protein inhibitor pentagalloylglucose (PGG) (5 μmol/L, #HY‐N0527, MedChemExpress).

Techniques: TUNEL Assay, Staining, Derivative Assay, Western Blot