Journal: EMBO Molecular Medicine

Article Title: Loss of CTLH component MAEA impairs DNA repair and replication and leads to developmental delay

doi: 10.1038/s44321-025-00352-x

Figure Lengend Snippet: ( A – C ) Clonogenic survival assays in WT and MAEA KO cells with HU ( A ), aphidicolin ( B ), and ATRi ( C ). ( D , E ) Quantification of chromatin-associated γH2AX and RPA signal in WT and MAEA KO cells upon HU ( D ) or ATRi ( E ) treatment for the indicated times. ( F ) Quantification of chromatin-associated γH2AX and RPA signal upon ATRi treatment in MAEA KO U2OS cells complemented with the indicated eGFP expression constructs. Representative images are shown in Fig. . ( G ) DNA fiber assay measuring replication fork progression in eGFP Only ( MAEA −/− ) and eGFP-MAEA WT cells following treatment with camptothecin. Clonogenic data ( A – C ) are from three independent experiments. Statistics were generated using an ordinary two-way ANOVA. Bars represent the mean ± SEM. DNA fiber data are the combined result of three independent experiments. P values were generated by performing a two-tailed Kruskal–Wallis test. γ is a measure of effect size. Bars represent the mean ± 95% CI. * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, **** P ≤ 0.0001; exact P values can be found in Appendix Table . Scatter plots represent two independent experiments. HU hydroxyurea, ATRi ATR inhibitor (AZD3768). .

Article Snippet: Hydroxyurea (HU) , Merck , H8627.

Techniques: Expressing, Construct, Generated, Two Tailed Test

Journal: EMBO Molecular Medicine

Article Title: Loss of CTLH component MAEA impairs DNA repair and replication and leads to developmental delay

doi: 10.1038/s44321-025-00352-x

Figure Lengend Snippet: ( A , B ) Quantification of replication tract lengths ( A ) and spontaneous replication fork stalling ( B ) in MAEA KO U2OS cells complemented with the indicated eGFP expression constructs. ( C , D ) Quantification of HU-induced replication fork stalling ( C ) and fork restart after HU wash-out ( D ) in MAEA KO U2OS cells complemented with the indicated eGFP expression constructs. ( E ) Quantification of replication fork degradation after HU treatment in MAEA KO U2OS cells complemented with the indicated eGFP expression constructs. ( F ) Quantification of replication fork degradation in primary fibroblasts from Patients 1, 2, and 4 versus a WT control fibroblast cell line following treatment with HU. The data were the combined result of three independent experiments. P values were generated by performing a two-tailed Kruskal–Wallis test on scatter plots and an ordinary one-way ANOVA on histograms. γ is a measure of effect size. Bars represent the mean ± 95% CI. * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, **** P ≤ 0.0001; exact P values can be found in Appendix Table . HU hydroxyurea, KO knockout. .

Article Snippet: Hydroxyurea (HU) , Merck , H8627.

Techniques: Expressing, Construct, Control, Generated, Two Tailed Test, Knock-Out

Journal: Nature Communications

Article Title: USP37 prevents unscheduled replisome unloading through MCM complex deubiquitination

doi: 10.1038/s41467-025-59770-7

Figure Lengend Snippet: a Expression-corrected CERES correlation scores were downloaded from the DepMap database for genes similar to USP37 knockout. Proteins involved in DNA replication and the DNA damage response are significantly enriched. Proteins are color coded similarly to Fig. (MCMs = blue, CDC45 = purple, polymerases = brown). Insert consists of ~ 750 of the > 17000 proteins analyzed, or ~ 4%. Source data are provided as a Source Data file. b Workflow. Cells were treated with siControl or siUSP37; hydroxyurea (HU) was added concurrently with the siRNA treatment to induce replication stress at 150 μM for 16 h. HU-treated cells were harvested at 16 h or released into fresh media without HU to recover for an additional 8 h, then harvested at 24 h. Cells were analyzed by flow cytometry for endogenous bound γH2AX and DNA content to distinguish G1 from G2 cells. c Bar graph for % γH2AX positive cells, acquired by chromatin flow cytometry from three biological replicates; mean with error bars ± SEM, unpaired two-tailed t test: siControl versus siUSP37 in HU or siControl versus siUSP37 8 h after release from HU; p = 0.1290, 0.6141, respectively. Source data are provided as a Source Data file. d Bar graph for % G2 cells from the same samples in ( c ). Mean with error bars ± SEM, unpaired two-tailed t test: siControl versus siUSP37 8 hours after release from HU; p = 0.0273. Source data are provided as a Source Data file. e Immunoblotting for USP37 or p21 from the same samples in ( c ). Representative of three biological replicates. f Box and whisker plots for chromatin-bound CDC45 per cell in S phase from the same samples in ( c ). Box represents 25 th −75 th percentile with line at median. The aggregate of three biological replicates was randomly down-sampled to 11,000 cells per sample. Relative fold-change of the means of bound CDC45 intensity from the three replicates was computed: siControl versus siUSP37 or siControl + HU versus siUSP37 + HU, unpaired two-tailed t test, p < 0.0001, p = 0.0422, respectively. Active CMG = ongoing forks under normal conditions; stalled CMG = stalled forks under stress conditions. Source data are provided as a Source Data file.

Article Snippet: The following chemicals/inhibitors were used in this study: Doxycycline (dox) (CalBiochem, cat. # 32485) was used at 2.5 or 5 ng/mL for the rescue experiments, and 100 or 25 ng/mL for the cyclin E1 or c-MYC overproduction experiments; ATR inhibitor (ATRi) AZD6738 (Selleck, cat. #S7693) was used at 5 μM; p97 inhibitor (Selleck, cat. #S8101) was used at 1.25 μM for the flow cytometry experiments and 5 μM for biochemical experiments; hydroxyurea (HU) drug (Selleck, cat. #S1896) was used at 150 μM for the HU block and release experiments.

Techniques: Expressing, Knock-Out, Flow Cytometry, Two Tailed Test, Western Blot, Whisker Assay