Journal: Synthetic and Systems Biotechnology

Article Title: Metabolic reprogramming of abscisic acid-producing strain Botrytis cinerea TB-31 toward terpenoid biosynthesis using a CRISPR/Cas9 ribonucleoprotein system

doi: 10.1016/j.synbio.2025.12.002

Figure Lengend Snippet: Transcriptomic analysis of B. cinerea strains TB-31, △ bcaba3 , and △ bcaba1234 . (A)∼(B) Regression analysis between transcriptomic data (RNA-seq) and RT‒qPCR validation. △ bcaba3 vs. TB-31: R 2 = 0.94, P < 0.001 (A); △ bcaba1234 vs. TB-31: R 2 = 0.97, P < 0.001 (B). (C) Heatmap of the relative transcriptional levels of the genes in the MVA pathway in △ bcaba1234 compared with those in TB-31. Gene transcription levels were normalized using Z score transformation, calculated as (X-mean)/SD, to represent the number of standard deviations above or below the mean transcription level. Upregulated genes are highlighted in red, while downregulated genes are shown in blue. The data are derived from three independent biological replicates. (D) Heatmap of the relative transcriptional levels of DEGs involved in other secondary metabolism pathways in △ bcaba1234 compared with those in TB-31. (E) Heatmap of the relative transcriptional levels of genes involved in intracellular primary metabolic pathways in the △ bcaba1234 strain compared with those in the TB-31 strain. Metabolic pathways are marked with gray boxes. PPP: pentose phosphate pathway; TCA: tricarboxylic acid cycle; GYC: glyoxylate cycle.

Article Snippet: The sgRNAs were transcribed using the T7 High Yield RNA Transcription Kit (Nanjing Vazyme Biotech Co., Ltd., Nanjing, China), and then purified using the RNA Clean & Concentrator Kit (Zymo Research Corp., Irvine, California, USA).

Techniques: RNA Sequencing, Biomarker Discovery, Transformation Assay, Derivative Assay

Journal: Molecular Therapy Oncology

Article Title: Survivin/BIRC5-derived peptide disrupts survivin dimerization and cell division and induces multifaceted anti-cancer effects

doi: 10.1016/j.omton.2025.201123

Figure Lengend Snippet: Cell death induction by 1H13-survivin/BIRC5-derived peptides targeted to the cytoplasm, mitochondria, or nucleus (A) The sequences added to the 1H13-BIRC5 peptide to target it to the: (1) cytoplasm, (2) mitochondria, or (3) nucleus. The underlined sequence represents the 1H13 peptide; in red, amino acids indicate D-amino acid substitutions. (B) A549 cells were incubated for 90 min with 5 μM FITC-labeled, nucleus-targeted peptide IH13-Nuc, and were also IF-stained with anti--SMAC, anti-IP3R or anti-GM130 antibodies, and stained with DAPI to visualize the mitochondria, ER, Golgi, and nucleus, respectively. Confocal microscope images are shown, with white arrows indicating peptide co-localization with the mitochondria (SMAC). Orange and yellow arrows indicate peptide presence in the nucleus and cytosol, respectively. (C) A549 cells were incubated with the mitochondria- or nucleus-targeted 1H13-BIRC5-derived peptide for 24 h in serum-free medium, followed by a cell proliferation assay using the SRB method. (D and E) Apoptotic cell death as induced in A549 cells following incubation for 24 h with the nucleus-targeted peptide (2/3D-1H13-Nuc) in the presence or absence of the indicated concentrations of the peptides in serum-free medium and subjected to FITC–annexin V/PI staining, followed by a flow cytometry analysis. Representative histograms for control and selected peptide concentration (D) and analysis of early and late apoptotic stages are shown (E). (F and G) Cell death as induced by 2/3D-1H13-Nuc in different cell lines, A549, SH-SY5Y, U-87MG, PC-3, and HUV-EC-C (F) or Jurkat, K562, and KMH2-LC (G) were treated with the indicated concentrations of the peptide for 24 h, then subjected to cell death analysis using propidium iodide (PI) staining and flow cytometry. (H and I) A549 cells were seeded at a density of 2 × 10 5 cells per well in a 12-well plate. After 24 h, the cells were transfected with 2 μg of a pCMV3-survivin expression plasmid (HG10356-UT, Sino Biological, China) or with an empty pCMV3 plasmid (control) using JetPrime transfection reagent (Polyplus, France), following the manufacturer’s instructions. Twenty-four hours post-transfection, the cells were re-seeded at 1 × 10 5 cells per well in a 12-well plate. After another 24 h, the culture medium was replaced with serum-free medium, and the cells were treated with the indicated concentrations of the 2/3D-1H13-Nuc peptide. Survivin overexpression levels were assessed by immunoblotting (H), and cell death was analyzed by propidium iodide (PI) staining followed by FACS analysis (I). Results represent the means ± SEM ( n = 3).

Article Snippet: After 24 h, the cells were transfected with 2 μg of a pCMV3-survivin expression plasmid (HG10356-UT, Sino Biological, China) or with an empty pCMV3 plasmid (control) using JetPrime transfection reagent (Polyplus, France), following the manufacturer’s instructions.

Techniques: Derivative Assay, Sequencing, Incubation, Labeling, Staining, Microscopy, Proliferation Assay, Flow Cytometry, Control, Concentration Assay, Transfection, Expressing, Plasmid Preparation, Over Expression, Western Blot

Journal: NAR Cancer

Article Title: Ribosome biogenesis is increased in hepatocellular carcinoma and represents a potential therapeutic target

doi: 10.1093/narcan/zcaf058

Figure Lengend Snippet: Nucleolar-targeting compounds inhibit nucleolar activity and induce selective DNA damage. ( A ) Representative images of EU incorporation (green) in THLE-3, PLC/PRF/5, and SNU-449 cells 24 h after treatment with four anti-cancer drugs targeting the nucleolus: CX-5461, BMH-21, Oxaliplatin, and JP-1302, or targeted therapy Sorafenib. Cells were treated with DMSO as control and were stained with DAPI (blue) and Pol II (yellow). ( B ) Mean nucleolar EU intensity per nucleus as a readout for rDNA transcription in THLE-3, PLC/PRF/5, and SNU-449 cells 24 h after treatment with four anti-cancer drugs targeting the nucleolus: CX-5461, BMH-21, Oxaliplatin, and JP-1302, or targeted therapy Sorafenib. Cells were treated with DMSO as control. The graph depicts one representative biological replicate ( n = 3), with data points representing individual cells, and the median indicated by the red line. A Kruskal–Wallis test was applied to assess differences between groups for the replicates combined, and statistical significance is indicated by numerical P -values. ( C ) Representative images of nuclear 53BP1 and γH2AX foci accumulation in THLE-3, PLC/PRF/5, and SNU-449 cells 24 h after treatment with four anti-cancer drugs targeting the nucleolus: CX-5461, BMH-21, Oxaliplatin, and JP-1302, or targeted therapy Sorafenib. Cells were treated with DMSO as control, and stained with DAPI (blue), 53BP1 (yellow), and γH2AX (red). ( D ) Fold change of nuclear 53BP1 foci per cell relative to DMSO. Individual points in the graph represent biological replicates ( n = 3). The graph depicts the mean ± SD and statistical significance was assessed by one sample t test and indicated by numerical P -values. ( E ) Fold change of nuclear γH2AX foci per cell relative to DMSO otherwise as in panel (D).

Article Snippet: Reverse transcription polymerase chain reaction (RT-PCR) was performed using the complementary DNA (cDNA) transcription kit with random primers (Thermo Fisher #4368814). cDNA was subjected to quantitative PCR (qPCR) using the Applied Biosystems 7500 Fast Real-Time PCR system with Fast SYBR TM Green Master Mix (Applied Biosystems #4385612) for detection.

Techniques: Activity Assay, Control, Staining