Journal: Chembiochem

Article Title: Monoclonal Antibodies Accessing the Cytosol of Living Cells and Binding to Polo‐Like Kinase 1 Interdomain Linker Affect Mitotic Behavior

doi: 10.1002/cbic.202500858

Figure Lengend Snippet: Anti‐PLK1 mAbs characterization. (A) Clones 35‐206, 3F8, and 13E8 were raised against full‐length PLK1, fragment 300–600, or fragment 300–400, respectively. KD: Kinase Domain; IDL: Interdomain Linker; PBD: Polo Box Domain. (B) Epitope mapping by Western blot. Lane 1: protein ladder; Lanes 2‐4: U2OS cells expressing 68 kDa PLK1, 14 kDa IDL (300–400), or 35 kDa IDL‐PBD (300–603), respectively.

Article Snippet: Human cervical carcinoma cells HeLa (ATCC Cat# CCL‐2, RRID:CVCL_0030), histone‐green fluorescent protein expressing HeLa cells H2BGFP‐HeLa (SCC117, Merck Millipore, RRID:CVCL_ZM02), human osteosarcoma cells U2OS (ATCC HTB‐96) U2OS (RRID:CVCL_0042) and human osteosarcoma cells expressing both luciferase (LUC) and green fluorescent protein (EGFP) (EGFPLuc‐U2OS, produced in our laboratory) were cultured adherently on plastic substrates (75 cm 2 Falcon tissue culture flasks) in high‐glucose Dulbecco’s Modified Eagle Medium (DMEM) supplemented with 10% fetal bovine serum (FBS) (Perbio, Brebières, France), 2 mM L‐glutamine, 100 U/mL penicillin G, and 100 μg/mL streptomycin.

Techniques: Clone Assay, Western Blot, Expressing

Journal: PLOS Biology

Article Title: Specificity and recognition of the ADP-ribosyl-ubiquitin modification in the DNA damage response

doi: 10.1371/journal.pbio.3003747

Figure Lengend Snippet: (A) Representative confocal images and (B) recruitment kinetics of full-length YFP-RNF114 recruitment to sites of DNA damage induced by 405 nm irradiation in U2OS WT (Black), DTX2KO (Red), or DTX3LKO (Blue) cells. (C) Representative confocal images and (D) recruitment kinetics of YFP-RNF114 ZnF2-3 + UIM recruitment to sites of DNA damage induced by 405 nm irradiation in U2OS WT (Black), DTX2KO (Red), or DTX3LKO (Blue) cells. (E) Representative confocal images and (F) recruitment kinetics of full-length YFP-RNF114 recruitment to sites of DNA damage induced by 405 nm irradiation in U2OS WT (Black) or DTX2KO (Red) treated or not with 1 μM Olaparib. (G) Representative confocal images and (H) recruitment kinetics of full-length YFP-RNF114 recruitment to sites of DNA damage induced by 405 nm irradiation in U2OS WT (Black) or HPF1KO (Purple) cells. (I) Representative confocal images and (J) recruitment kinetics of full length YFP-RNF138 recruitment to sites of DNA damage induced by 405 nm irradiation in U2OS WT (Black) or DTX2KO (Red) cells. (K) Representative confocal images and (L) recruitment kinetics of full-length YFP-RNF166 recruitment to sites of DNA damage induced by 405 nm irradiation in U2OS WT (Black) or DTX2KO (Red) cells. (M) Representative confocal images and (N) recruitment kinetics of full-length mCherry-RNF114 recruitment to sites of DNA damage induced by 405 nm irradiation in U2OS WT (Black), DTX2KO (Red) cells, and DTX2KO cells complemented with YFP-DTX2 WT (Blue) or E608R (Green). For all images, Scale bar, 5 µm. Data is representative of at least 3 independent replicates. Curves show mean ± s.e.m. of between 8 and 20 cells per condition. P values were obtained using an unpaired Student t test with Bonferroni correction (ns, not significant, ** P < 0.01,**** P < 0.001). The data for this figure can be found at https://www.ebi.ac.uk/biostudies/studies/ S-BIAD2514.

Article Snippet: Briefly, human U2OS osteosarcoma (American Type Culture Collection (ATCC HTB-96) WT, DTX2KO, DTX3LKO, [ ] or HPF1KO [ ] cells were cultured in Dulbecco’s modified Eagle’s medium (Sigma-Aldrich) supplemented with 10% fetal bovine serum (FBS; Gibco) and penicillin-streptomycin (100 U/ml; Gibco) in a humidified atmosphere at 37°C with 5% CO 2 .

Techniques: Irradiation