Journal: Scientific Reports

Article Title: CDC25C downregulation suppresses HCC growth via mitochondrial stress-induced autophagy and apoptosis

doi: 10.1038/s41598-026-36351-2

Figure Lengend Snippet: The suppressed levels of CDC25C in cell line and xenograft model. ( A ) The CDC25C gene was detected via agarose gel electrophoresis, with β-actin serving as an internal control. The marker used was a 1000 bp Gene Ruler, and the expected product sizes were 970 bp for CDC25C and 709 bp for β-actin. The negative control prepared from a sample containing just the Master-Mix, in which there was no DNA contamination. ( B ) The relative expression of CDC25C mRNA was evaluated using qRT-PCR. ( C ) The expression level of CDC25C protein was assessed through Western blotting and quantified using ImageJ software, with GAPDH serving as the internal control. Data are presented as means ± SDs from three independent experiments. c p <0.001.

Article Snippet: CDC25C , Mouse , #66912-1-Ig , Proteintech , , 1:1000.

Techniques: Agarose Gel Electrophoresis, Control, Marker, Negative Control, Expressing, Quantitative RT-PCR, Western Blot, Software

Journal: Scientific Reports

Article Title: CDC25C downregulation suppresses HCC growth via mitochondrial stress-induced autophagy and apoptosis

doi: 10.1038/s41598-026-36351-2

Figure Lengend Snippet: Downregulation of CDC25C inhibited the malignant biological behaviors of Hepa1-6 cells. ( A ) The colony formation ability was assessed using a plate cloning assay at 100× magnification, with the colony formation rate calculated by counting the number of colonies containing more than 50 cells. ( B ) The lateral migration ability of cells was measured using a wound healing assay at 100× magnification, and the migration rate (scratch healing rate) was calculated by measuring the width of the scratch. ( C , D ) The longitudinal migration and invasion abilities of the cells were evaluated using Transwell assays, quantifying the number of cells that passed through the filter. The scale bar represents 20 μm. Data are presented as means ± SDs from three independent experiments. a p <0.05, b p <0.01, c p <0.001.

Article Snippet: CDC25C , Mouse , #66912-1-Ig , Proteintech , , 1:1000.

Techniques: Cloning, Migration, Wound Healing Assay

Journal: Scientific Reports

Article Title: CDC25C downregulation suppresses HCC growth via mitochondrial stress-induced autophagy and apoptosis

doi: 10.1038/s41598-026-36351-2

Figure Lengend Snippet: Downregulation of CDC25C altered the morphology of the subcellular structure of Hepa1-6 cells. The images of Hepa1-6 cells captured under transmission electron microscopy. The scale bar represents a measurement of 2 μm at lower magnification and 200 nm at higher magnification. White arrows indicate healthy mitochondria, orange arrows denote autophagosomes, blue arrow highlight cellular vacuolation, and red arrows signify swollen and dysfunctional mitochondria.

Article Snippet: CDC25C , Mouse , #66912-1-Ig , Proteintech , , 1:1000.

Techniques: Transmission Assay, Electron Microscopy

Journal: Scientific Reports

Article Title: CDC25C downregulation suppresses HCC growth via mitochondrial stress-induced autophagy and apoptosis

doi: 10.1038/s41598-026-36351-2

Figure Lengend Snippet: Downregulation of CDC25C induced a mitochondrial stress response. ( A , B ) Mitochondrial calcium concentrations were measured in Hepa1-6 and AML12 cells using the Rhod-2/AM fluorescent probe, with ImageJ software employed to quantify the red fluorescence intensity. ( C , D ) ROS levels were assessed in Hepa1-6 cells and AML12 using the MitoSOX fluorescent probe, and the red fluorescence intensity was quantified using ImageJ software. ( E ) The relative mRNA expression levels of CHOP, HSP60, ClpP, and LONP1 were evaluated via qRT-PCR. ( F , G ) The relative protein expression levels of CHOP, HSP60, ClpP, and LONP1 were determined through Western blotting and quantified using ImageJ software, with GAPDH serving as the internal control. The scale bar represents 10 μm. Data are presented as means ± SDs from three independent experiments. a p <0.05, b p <0.01, c p <0.001. ‘ns’ indicates no statistical significance.

Article Snippet: CDC25C , Mouse , #66912-1-Ig , Proteintech , , 1:1000.

Techniques: Software, Fluorescence, Expressing, Quantitative RT-PCR, Western Blot, Control

Journal: Scientific Reports

Article Title: CDC25C downregulation suppresses HCC growth via mitochondrial stress-induced autophagy and apoptosis

doi: 10.1038/s41598-026-36351-2

Figure Lengend Snippet: Downregulation of CDC25C induced an autophagic response. ( A ) The relative mRNA expression levels of LC3, p62, and Beclin1 were evaluated using qRT-PCR. ( B , C ) The relative protein expression levels of LC3, p62, and Beclin1 were determined through Western blotting and quantified using ImageJ software, with GAPDH serving as the internal control. Data are presented as means ± SDs from three independent experiments. a p < 0.05, b p < 0.01, c p <0.001.

Article Snippet: CDC25C , Mouse , #66912-1-Ig , Proteintech , , 1:1000.

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Software, Control

Journal: Scientific Reports

Article Title: CDC25C downregulation suppresses HCC growth via mitochondrial stress-induced autophagy and apoptosis

doi: 10.1038/s41598-026-36351-2

Figure Lengend Snippet: Downregulation of CDC25C induced a mitochondria-mediated apoptosis. ( A ) Hoechst 33258 staining was employed to morphologically identify apoptotic cells. ( B ) Annexin V/TMRE costaining was utilized for apoptosis detection via flow cytometry to investigate the apoptosis rate and quantified using ImageJ software. ( C ) The relative mRNA expression levels of Cyt c, Caspase-3 and Caspase-9 were evaluated using qRT-PCR. ( D , E ) The relative protein expression levels of Cyt c, Caspase-3 and Caspase-9 were determined through Western blotting and quantified using ImageJ software, with GAPDH serving as the internal control. The scale bar represents 50 μm. Data are presented as means ± SDs from three independent experiments. a p < 0.05, b p < 0.01, c p <0.001. ‘ns’ indicates no statistical significance.

Article Snippet: CDC25C , Mouse , #66912-1-Ig , Proteintech , , 1:1000.

Techniques: Staining, Flow Cytometry, Software, Expressing, Quantitative RT-PCR, Western Blot, Control

Journal: PLoS ONE

Article Title: Androgens Upregulate Cdc25C Protein by Inhibiting Its Proteasomal and Lysosomal Degradation Pathways

doi: 10.1371/journal.pone.0061934

Figure Lengend Snippet: LNCaP C-33, C-81 and PC-3 PCa cells were plated at a density of 8×10 3 , 6×10 3 and 4.8×10 3 cells/cm 2 , respectively, in duplicates for 3 days in regular medium. (Left FBS panel, lanes #1–3) Cells were replaced with fresh regular medium for 24 hr and then harvested. (Right SR panel, lanes #4–6) All three PCa cells were then steroid-starved for 48 hr in a steroid-reduced (SR) medium and then harvested. Total cell lysate proteins were analyzed for cPAcP, SHP1, SHP2, Cdc25A, Cdc25B, Cdc25C, Cyclin B1 and Cyclin D1. Cdc25C spliced forms were observed upon long exposure of films (lower panel of Cdc25C). β-actin was analyzed and used as a loading control.

Article Snippet: The Cdc25C cDNA plasmid was purchased from OriGene Technologies, Inc. (Rockville, MD, USA).

Techniques:

Journal: PLoS ONE

Article Title: Androgens Upregulate Cdc25C Protein by Inhibiting Its Proteasomal and Lysosomal Degradation Pathways

doi: 10.1371/journal.pone.0061934

Figure Lengend Snippet: (A) LNCaP C-33 and C-81 cells were seeded at a density of 8×10 3 and 6×10 3 cells/cm 2 , respectively, in duplicates for 3 days in regular medium. Cells were steroid-starved for 48 hr in SR medium and then treated with 10 nM DHT for 24 hr and 48 hr. Total cell lysate proteins were harvested and analyzed for Cdc25C, Cdc25B, cPAcP, Cyclin B1 and Cyclin D1 proteins. β-actin was detected for serving as a loading control. (B) AS LNCaP C-33 and (C) VCaP cells were plated at a density of 8×10 3 and 2×10 4 cells/cm 2 , respectively, for 3 days, and then steroid starved for 48 hr in SR medium. Cells were treated with 10 nM DHT with or without 10 µM Casodex for 48 hr. (B) For C-33 cells, total cell lysate proteins were analyzed for AR, Cdc25A, Cdc25B, Cdc25C, cPAcP, PSA, Cyclin D1 and Cyclin B1 protein levels. (C) For VCaP cells, total cell lysate proteins were analyzed for AR, Cdc25C, Cyclin D1, Cyclin B1 and PSA protein levels. β-actin was analyzed in each experiment and used as a loading control. (D) Effect of DHT vs . EGF on Cdc25C protein level in C-33 cells. Steroid-starved C-33 cells were treated with 10 nM DHT or 10 ng/ml EGF for 24 hr, and control cells were treated with solvent alone. All cells were harvested for analyzing Cdc25C protein level. As controls, cPAcP, ErbB-2 and its tyrosine phosphorylation at Y877, AR and its phosphorylation at S81, and Cyclin B1 were also analyzed. Tubulin was detected for serving as a loading control.

Article Snippet: The Cdc25C cDNA plasmid was purchased from OriGene Technologies, Inc. (Rockville, MD, USA).

Techniques:

Journal: PLoS ONE

Article Title: Androgens Upregulate Cdc25C Protein by Inhibiting Its Proteasomal and Lysosomal Degradation Pathways

doi: 10.1371/journal.pone.0061934

Figure Lengend Snippet: (A,B) LNCaP C-33 and (C) PC-3 cells were plated at a density of 1.2×10 4 or 1×10 4 cells/cm 2 for 72 hr and then transfected with Cdc25C cDNA (A) or shRNA (B,C) plasmids. Control cells were transfected with the respective vector alone. (A) Cdc25C cDNA transfected C-33 cells were cultured in regular medium and the cell number was then counted after 2 days. (B) Cdc25C shRNA-transfected C-33 cells were steroid starved for 48 hr in SR medium, and then treated with or without 10 nM DHT for 2 days. (C) Cdc25C shRNA transfected PC-3 cells were cultured in regular medium for 2 days, and the cell number was counted. The ratio of cell proliferation was calculated by normalizing the experimental cell number to that of control cells transfected with vector alone, respectively. Total cell lysate proteins were analyzed for Cdc25A, Cdc25B, Cdc25C, Cyclin B1 and/or Cyclin D1 proteins. β-actin was used as a loading control. The ratios of Cdc25C, Cyclin B1 and CyclinD1 protein levels to β-actin were calculated after semi-quantification by densitometric analyses on films with different exposure time periods. * p <0.05, n = 2×3; Bar, standard deviation.

Article Snippet: The Cdc25C cDNA plasmid was purchased from OriGene Technologies, Inc. (Rockville, MD, USA).

Techniques: Transfection, shRNA, Plasmid Preparation, Cell Culture, Standard Deviation

Journal: PLoS ONE

Article Title: Androgens Upregulate Cdc25C Protein by Inhibiting Its Proteasomal and Lysosomal Degradation Pathways

doi: 10.1371/journal.pone.0061934

Figure Lengend Snippet: (A) Effects of de novo biosynthesis inhibitors on Cdc25C protein levels by androgens. LNCaP C-33 cells were plated at a density of 1.6×10 4 cells/cm 2 for 3 days in regular medium. Cells were steroid starved for 48 hr in a SR medium, and then treated with 5 µg/mL Actinomycin D (Act D) or 10 µg/mL cycloheximide (CHX) for 24 hr alone or 10 nM DHT was added 30 min post treatment with Act D and CHX. Total cell lysate proteins were analyzed for Cdc25C, AR and PSA protein level. β-actin was analyzed as a loading control. (B & C) Effects of inhibitors of proteasomal degradation pathway on Cdc25C protein level. LNCaP C-33 cells were seeded in regular medium for 72 hr, steroid starved for 48 hr and the cells were treated with or without 10 nM DHT or different concentrations of proteasomal inhibitors (B) MG132 (0.01, 0.05 and 0.1 µM) and (C) Epoximicin (0.1 and 1.0 M). Cells were harvested and analyzed for Cdc25C protein levels. β-actin was analyzed serving as a loading control. (D) Effect of androgens on the ubiquitination of Cdc25C protein. Steroid-starved LNCaP C-33 cells were treated with 10 nM DHT or solvent alone for 48 hr. An aliquot of total cellular lysate proteins was immunoprecipitated (IP) by reacting with anti-Cdc25C Ab or normal IgG and followed by Protein A-Sepharose beads. The immune complexes were analyzed by immunoblotting (IB) with anti-Cdc25C Ab (left panel) or anti-ubiquitin Ab (right panel). The positions of ubiquitinated Cdc25C protein (Ubi-Cdc25C) are indicated by arrows; while an additional band was detected by anti-ubiquitin Ab (Complex Ib). (E & F) Effects of inhibitors of lysosomal degradation pathway on Cdc25C protein levels. LNCaP C-33 cells were seeded as described in B & C, after steroid starvation, cells were treated with or without 10 nM DHT or different concentrations of lysosomal protease inhibitors. (E) Leupeptin (10 to 200 µM) and (F) E64D (0.01 to 1.0 µM) for 48 hr. Cells were harvested and analyzed for Cdc25C protein levels. β-actin was analyzed as a loading control.

Article Snippet: The Cdc25C cDNA plasmid was purchased from OriGene Technologies, Inc. (Rockville, MD, USA).

Techniques: Immunoprecipitation, Western Blot

Journal: Genes & Development

Article Title: Rapid embryonic cell cycles defer the establishment of heterochromatin by Eggless/SetDB1 in Drosophila

doi: 10.1101/gad.321646.118

Figure Lengend Snippet: Interphase duration limits the establishment of heterochromatin. ( A ) Selected stills from live imaging of Halo-egg (shown in green) embryos injected with Cy5-labeled Histone proteins (shown in magenta). Egg dissociates from the chromatin upon mitotic chromosome condensation (150 sec). Bar, 3 µm. See Supplemental Movie S6 . ( B ) Arresting the cell cycle in interphase 13 permits continued accumulation of Egg and HP1a at heterochromatin. Embryos were filmed by confocal microscopy after injection of buffer or dsRNA against cyclins A, B, and B3. Bar, 2 µm. The beginning of cycle 13 was set as 0:00. See Supplemental Movie S7 . ( C–E ) Shortening interphase duration reduces HP1a accumulation. ( C ) Stills from confocal microscopy tracking GFP-HP1a in embryos laid from mothers either heterozygous or homozygous for grp. Note that grp embryos enter mitosis 13 before completing S phase, leading to anaphase bridges (17:00). Bar, 4 µm. The start of interphase 13 was set as 0:00. ( D ) Plot of the average number of HP1a foci per nucleus over time in control embryos (orange) or grp embryos (black). Each line represents data from a different embryo, and foci of GFP-HP1a were counted in ∼15 nuclei per embryo. ( E ) Inducing an early mitosis 14 by expression of twine (cdc25) interrupts heterochromatin formation. Embryos expressing GFP-HP1a were filmed following injection of twine mRNA during cycle 13. Bar, 3 µm. The beginning of cycle 14 was set as 0:00.

Article Snippet: JabbaTrap and Cdc25 ( twine ) mRNAs were synthesized using the CellScript T7 mRNA production system and injected at a concentration of 600 ng/μL. dsRNA against the cyclins A, B, and B3 was prepared as described in .

Techniques: Imaging, Injection, Labeling, Confocal Microscopy, Control, Expressing

Journal: Lead-Seeking Approaches

Article Title: Lead Discovery Using Virtual Screening

doi: 10.1007/7355_2009_3

Figure Lengend Snippet: Virtual Screening examples using primarily SBVS

Article Snippet: CDC25 phosphatase , Chembridge 413 K (313 K after filtering) , FRED/Surflex , 99 hits of 1,500 tested , [ ] .

Techniques: Binding Assay