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hela hpv18  (ATCC)


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    Structured Review

    ATCC hela hpv18
    Hela Hpv18, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 10460 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/hpv18/HeLa/pm41599644-88-4-17
    Average 99 stars, based on 10460 article reviews
    hela hpv18 - by Bioz Stars, 2026-09
    99/100 stars

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    Related Articles

    Cell Culture:

    Article Title: Activation of p53-regulated pro-survival signals and hypoxia-independent mitochondrial targeting of TIGAR by human papillomavirus E6 oncoproteins.
    Article Snippet: The high-risk subtype human papillomaviruses (hrHPVs) infect and oncogenically transform basal epidermal stem cells associated with the development of squamous-cell epithelial cancers.. The viral E6 oncoprotein destabilizes the p53 tumor suppressor, inhibits p53 K120-acetylation by the Tat-interacting protein of 60 kDa (TIP60, or Kat5), and prevents p53-dependent apoptosis.. Intriguingly, the p53 gene is infrequently mutated in HPV + cervical cancer clinical isolates which suggests a possible paradoxical role for this gatekeeper in viral carcinogenesis.

    Control:

    Article Title: A fluorometric hybridization assay for detecting and genotyping high-risk human papillomavirus 16 and 18 in archival tissues of cervical specimens
    Article Snippet: .. Source of plasmids DNA DNA plasmids containing the entire genomes of HPV16 (ATCC 45113D) and HPV18 (ATCC 45152D) were purchased from American Type Culture Collection (ATCC, Manassas, USA) and used as the control to develop efficient hybridization methods. .. Source of plasmids DNA DNA plasmids containing the entire genomes of HPV16 (ATCC 45113D) and HPV18 (ATCC 45152D) were purchased from American Type Culture Collection (ATCC, Manassas, USA) and used as the control to develop efficient hybridization methods.

    Hybridization:

    Article Title: A fluorometric hybridization assay for detecting and genotyping high-risk human papillomavirus 16 and 18 in archival tissues of cervical specimens
    Article Snippet: .. Source of plasmids DNA DNA plasmids containing the entire genomes of HPV16 (ATCC 45113D) and HPV18 (ATCC 45152D) were purchased from American Type Culture Collection (ATCC, Manassas, USA) and used as the control to develop efficient hybridization methods. .. Source of plasmids DNA DNA plasmids containing the entire genomes of HPV16 (ATCC 45113D) and HPV18 (ATCC 45152D) were purchased from American Type Culture Collection (ATCC, Manassas, USA) and used as the control to develop efficient hybridization methods.

    Transfection:

    Article Title: WAPL-Dependent Repair of Damaged DNA Replication Forks Underlies Oncogene-Induced Loss of Sister Chromatid Cohesion.
    Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER RNAimax Life Technologies Cat# 13778 Lipofectamine3000 Invitrogen Cat# L3000001 Demecolcin Sigma Cat# D1925 siR-DNA Spirochrome Cat# SC007 Cell Player 96-well kinetic caspase-3/7 reagent Essen Bioscience Cat# 4440 GFP-Trap agarose beads Chromotek Cat# gta-20 Control agarose beads Chromotek Cat# bab-20 GST–Raf1 RBD coupled to glutathione– agarose beads (Wolthuis et al., 1998) N/A Propidium Iodide Thermo Fisher Scientific Cat# P3566 Puromycin Sigma Cat# P7255 Blasticidin Fisher Scientific Cat# A1113903 G418 Merck Cat# 345810 Doxycycline Sigma Cat# D9891 Fetal Calf Serum Life Technologies Cat# 3100738 Nonessential amino acids Gibco Cat# 11140035 Sodium pyruvate Gibco Cat# 11360039 Penicillin/Streptomycin Gibco/ Life technologies Cat# 15140122 Polybrene Sigma Cat# H9268 Fluor-gel with TES buffer Electron Microscopy Science Cat# 17985–30 Critical Commercial Assays High pure RNA isolation kit Roche Cat# 11828665001 Vysis D7S486/CEP 7 FISH Probe Kit Abbot Molecular Cat# 04N78–020 Experimental Models: Cell Lines TKO-Bcl2-p53KO (TBP) MEFs (Benedict et al., 2018) N/A TBP-WaplKD MEFs This paper N/A TBP-WaplKD #1 MEFs This paper N/A TBP-WaplKD #2 MEFs This paper N/A TBP-Pds5bKD #1 MEFs This paper N/A TBP-Pds5bKD #2 MEFs This paper N/A TBP-NipblKD This paper N/A BJ-hTERT ATCC N/A WaplKD BJ-hTERT This paper N/A HAP1 (Haarhuis et al., 2017) N/A HAP1 WaplKO (Haarhuis et al., 2017) N/A HAP1 Mau2KO (Haarhuis et al., 2017) N/A HAP1 WaplKO Mau2KO (Haarhuis et al., 2017) N/A RPE1-hTERT ATCC N/A RPE1-hTERT-TetOn-Cas9 This paper N/A RPE1-hTERT-TetOn-Cas9_p53KO This paper N/A RPE1-hTERT-TetOn-Cas9-STAG2KO or TP53KO or ESCO2 KO This paper N/A RPE1-hTERT-TetOn-Cas9_p53KO KRAS G12D#1 This paper N/A RPE1-hTERT-TetOn-Cas9_p53KO KRAS G12D#2 This paper N/A RPE1-hTERT-TetOn-Cas9_TetOn-Cas9_SMC3Venus_p53KO This paper N/A Human fibroblast VU-HF1 This paper N/A Human fibroblast VU-HF2 (previously named Fen5280) (Xia et al., 2007) N/A Human fibroblast VU-HF3 This paper N/A (Continued on next page) e2 Developmental Cell 52, 1–16.e1–e7, March 23, 2020 .. REAGENT or RESOURCE SOURCE IDENTIFIER Human fibroblast VU-HF4 This paper N/A LN9SV (Van der Lelij et al., 2009) N/A ARPE-19, CRL1790, RPTEC and HEK293T ATCC N/A HNSCC cells (Stoepker et al., 2015) N/A Lymphoblast HSC93 (Ishida and Buchwald, 1982) N/A Primary foreskin keratinocytes EK94-2 transfected with HPV16 and HPV18 (Steenbergen et al., 1996) N/A Colorectal cancer cell lines SW620, HCT116, COLO320, WiDr, Caco2 ATCC N/A Retinoblastoma cell lines Y79 and Weri (McFall et al., 1977; Reid et al., 1974) N/A Breast cancer cell lines Gift from J. Martens (Erasmus MC, Rotterdam) N/A VU-SCC78 + TetOn-STAG2 This paper N/A OCUB-M + TetOn-STAG2 This paper N/A Oligonucleotides For QPCR primer sequences, see section ‘Quantitative PCR’ This paper N/A For siRNA sequences, see section ‘siRNA experiments’ This paper N/A For other primers, see ‘Method Details’ This paper N/A ssODN template for the introduction of the KRASG12D mutation, see ‘CRISPR-Cas9 gene editing’ section This paper N/A Recombinant DNA pLVX-TRE3G Clontech Cat# 631187 pLVX-Tet3G Clontech Cat# 631187 Lenti-X HT packaging system Clontech Cat# 631276 pLenti-GFP-Muc18 Gift from P. Heutink N/A CSII-EF-MCS-mKO-hCdt1 (30/120) and CSII-EFMCS-mAG-hGem (1/110) Gift from A. Miyawaki (Sakaue-Sawano et al., 2008) N/A pBABE-blasticidin-H2B-YFP Gift from A. Koch N/A pLenti Gift from P. Heutink N/A pLKO.1 shRNA vectors from the Mission mouse TRC v1.0 collection Sigma See Method Details for shRNA sequences N/A Software and Algorithms Casplab Casplab http://casplab.com/download Fuij ImageJ https://imagej.nih.gov/ij/download.html Image Studio Lite Ver. ..

    Real-time Polymerase Chain Reaction:

    Article Title: WAPL-Dependent Repair of Damaged DNA Replication Forks Underlies Oncogene-Induced Loss of Sister Chromatid Cohesion.
    Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER RNAimax Life Technologies Cat# 13778 Lipofectamine3000 Invitrogen Cat# L3000001 Demecolcin Sigma Cat# D1925 siR-DNA Spirochrome Cat# SC007 Cell Player 96-well kinetic caspase-3/7 reagent Essen Bioscience Cat# 4440 GFP-Trap agarose beads Chromotek Cat# gta-20 Control agarose beads Chromotek Cat# bab-20 GST–Raf1 RBD coupled to glutathione– agarose beads (Wolthuis et al., 1998) N/A Propidium Iodide Thermo Fisher Scientific Cat# P3566 Puromycin Sigma Cat# P7255 Blasticidin Fisher Scientific Cat# A1113903 G418 Merck Cat# 345810 Doxycycline Sigma Cat# D9891 Fetal Calf Serum Life Technologies Cat# 3100738 Nonessential amino acids Gibco Cat# 11140035 Sodium pyruvate Gibco Cat# 11360039 Penicillin/Streptomycin Gibco/ Life technologies Cat# 15140122 Polybrene Sigma Cat# H9268 Fluor-gel with TES buffer Electron Microscopy Science Cat# 17985–30 Critical Commercial Assays High pure RNA isolation kit Roche Cat# 11828665001 Vysis D7S486/CEP 7 FISH Probe Kit Abbot Molecular Cat# 04N78–020 Experimental Models: Cell Lines TKO-Bcl2-p53KO (TBP) MEFs (Benedict et al., 2018) N/A TBP-WaplKD MEFs This paper N/A TBP-WaplKD #1 MEFs This paper N/A TBP-WaplKD #2 MEFs This paper N/A TBP-Pds5bKD #1 MEFs This paper N/A TBP-Pds5bKD #2 MEFs This paper N/A TBP-NipblKD This paper N/A BJ-hTERT ATCC N/A WaplKD BJ-hTERT This paper N/A HAP1 (Haarhuis et al., 2017) N/A HAP1 WaplKO (Haarhuis et al., 2017) N/A HAP1 Mau2KO (Haarhuis et al., 2017) N/A HAP1 WaplKO Mau2KO (Haarhuis et al., 2017) N/A RPE1-hTERT ATCC N/A RPE1-hTERT-TetOn-Cas9 This paper N/A RPE1-hTERT-TetOn-Cas9_p53KO This paper N/A RPE1-hTERT-TetOn-Cas9-STAG2KO or TP53KO or ESCO2 KO This paper N/A RPE1-hTERT-TetOn-Cas9_p53KO KRAS G12D#1 This paper N/A RPE1-hTERT-TetOn-Cas9_p53KO KRAS G12D#2 This paper N/A RPE1-hTERT-TetOn-Cas9_TetOn-Cas9_SMC3Venus_p53KO This paper N/A Human fibroblast VU-HF1 This paper N/A Human fibroblast VU-HF2 (previously named Fen5280) (Xia et al., 2007) N/A Human fibroblast VU-HF3 This paper N/A (Continued on next page) e2 Developmental Cell 52, 1–16.e1–e7, March 23, 2020 .. REAGENT or RESOURCE SOURCE IDENTIFIER Human fibroblast VU-HF4 This paper N/A LN9SV (Van der Lelij et al., 2009) N/A ARPE-19, CRL1790, RPTEC and HEK293T ATCC N/A HNSCC cells (Stoepker et al., 2015) N/A Lymphoblast HSC93 (Ishida and Buchwald, 1982) N/A Primary foreskin keratinocytes EK94-2 transfected with HPV16 and HPV18 (Steenbergen et al., 1996) N/A Colorectal cancer cell lines SW620, HCT116, COLO320, WiDr, Caco2 ATCC N/A Retinoblastoma cell lines Y79 and Weri (McFall et al., 1977; Reid et al., 1974) N/A Breast cancer cell lines Gift from J. Martens (Erasmus MC, Rotterdam) N/A VU-SCC78 + TetOn-STAG2 This paper N/A OCUB-M + TetOn-STAG2 This paper N/A Oligonucleotides For QPCR primer sequences, see section ‘Quantitative PCR’ This paper N/A For siRNA sequences, see section ‘siRNA experiments’ This paper N/A For other primers, see ‘Method Details’ This paper N/A ssODN template for the introduction of the KRASG12D mutation, see ‘CRISPR-Cas9 gene editing’ section This paper N/A Recombinant DNA pLVX-TRE3G Clontech Cat# 631187 pLVX-Tet3G Clontech Cat# 631187 Lenti-X HT packaging system Clontech Cat# 631276 pLenti-GFP-Muc18 Gift from P. Heutink N/A CSII-EF-MCS-mKO-hCdt1 (30/120) and CSII-EFMCS-mAG-hGem (1/110) Gift from A. Miyawaki (Sakaue-Sawano et al., 2008) N/A pBABE-blasticidin-H2B-YFP Gift from A. Koch N/A pLenti Gift from P. Heutink N/A pLKO.1 shRNA vectors from the Mission mouse TRC v1.0 collection Sigma See Method Details for shRNA sequences N/A Software and Algorithms Casplab Casplab http://casplab.com/download Fuij ImageJ https://imagej.nih.gov/ij/download.html Image Studio Lite Ver. ..

    Mutagenesis:

    Article Title: WAPL-Dependent Repair of Damaged DNA Replication Forks Underlies Oncogene-Induced Loss of Sister Chromatid Cohesion.
    Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER RNAimax Life Technologies Cat# 13778 Lipofectamine3000 Invitrogen Cat# L3000001 Demecolcin Sigma Cat# D1925 siR-DNA Spirochrome Cat# SC007 Cell Player 96-well kinetic caspase-3/7 reagent Essen Bioscience Cat# 4440 GFP-Trap agarose beads Chromotek Cat# gta-20 Control agarose beads Chromotek Cat# bab-20 GST–Raf1 RBD coupled to glutathione– agarose beads (Wolthuis et al., 1998) N/A Propidium Iodide Thermo Fisher Scientific Cat# P3566 Puromycin Sigma Cat# P7255 Blasticidin Fisher Scientific Cat# A1113903 G418 Merck Cat# 345810 Doxycycline Sigma Cat# D9891 Fetal Calf Serum Life Technologies Cat# 3100738 Nonessential amino acids Gibco Cat# 11140035 Sodium pyruvate Gibco Cat# 11360039 Penicillin/Streptomycin Gibco/ Life technologies Cat# 15140122 Polybrene Sigma Cat# H9268 Fluor-gel with TES buffer Electron Microscopy Science Cat# 17985–30 Critical Commercial Assays High pure RNA isolation kit Roche Cat# 11828665001 Vysis D7S486/CEP 7 FISH Probe Kit Abbot Molecular Cat# 04N78–020 Experimental Models: Cell Lines TKO-Bcl2-p53KO (TBP) MEFs (Benedict et al., 2018) N/A TBP-WaplKD MEFs This paper N/A TBP-WaplKD #1 MEFs This paper N/A TBP-WaplKD #2 MEFs This paper N/A TBP-Pds5bKD #1 MEFs This paper N/A TBP-Pds5bKD #2 MEFs This paper N/A TBP-NipblKD This paper N/A BJ-hTERT ATCC N/A WaplKD BJ-hTERT This paper N/A HAP1 (Haarhuis et al., 2017) N/A HAP1 WaplKO (Haarhuis et al., 2017) N/A HAP1 Mau2KO (Haarhuis et al., 2017) N/A HAP1 WaplKO Mau2KO (Haarhuis et al., 2017) N/A RPE1-hTERT ATCC N/A RPE1-hTERT-TetOn-Cas9 This paper N/A RPE1-hTERT-TetOn-Cas9_p53KO This paper N/A RPE1-hTERT-TetOn-Cas9-STAG2KO or TP53KO or ESCO2 KO This paper N/A RPE1-hTERT-TetOn-Cas9_p53KO KRAS G12D#1 This paper N/A RPE1-hTERT-TetOn-Cas9_p53KO KRAS G12D#2 This paper N/A RPE1-hTERT-TetOn-Cas9_TetOn-Cas9_SMC3Venus_p53KO This paper N/A Human fibroblast VU-HF1 This paper N/A Human fibroblast VU-HF2 (previously named Fen5280) (Xia et al., 2007) N/A Human fibroblast VU-HF3 This paper N/A (Continued on next page) e2 Developmental Cell 52, 1–16.e1–e7, March 23, 2020 .. REAGENT or RESOURCE SOURCE IDENTIFIER Human fibroblast VU-HF4 This paper N/A LN9SV (Van der Lelij et al., 2009) N/A ARPE-19, CRL1790, RPTEC and HEK293T ATCC N/A HNSCC cells (Stoepker et al., 2015) N/A Lymphoblast HSC93 (Ishida and Buchwald, 1982) N/A Primary foreskin keratinocytes EK94-2 transfected with HPV16 and HPV18 (Steenbergen et al., 1996) N/A Colorectal cancer cell lines SW620, HCT116, COLO320, WiDr, Caco2 ATCC N/A Retinoblastoma cell lines Y79 and Weri (McFall et al., 1977; Reid et al., 1974) N/A Breast cancer cell lines Gift from J. Martens (Erasmus MC, Rotterdam) N/A VU-SCC78 + TetOn-STAG2 This paper N/A OCUB-M + TetOn-STAG2 This paper N/A Oligonucleotides For QPCR primer sequences, see section ‘Quantitative PCR’ This paper N/A For siRNA sequences, see section ‘siRNA experiments’ This paper N/A For other primers, see ‘Method Details’ This paper N/A ssODN template for the introduction of the KRASG12D mutation, see ‘CRISPR-Cas9 gene editing’ section This paper N/A Recombinant DNA pLVX-TRE3G Clontech Cat# 631187 pLVX-Tet3G Clontech Cat# 631187 Lenti-X HT packaging system Clontech Cat# 631276 pLenti-GFP-Muc18 Gift from P. Heutink N/A CSII-EF-MCS-mKO-hCdt1 (30/120) and CSII-EFMCS-mAG-hGem (1/110) Gift from A. Miyawaki (Sakaue-Sawano et al., 2008) N/A pBABE-blasticidin-H2B-YFP Gift from A. Koch N/A pLenti Gift from P. Heutink N/A pLKO.1 shRNA vectors from the Mission mouse TRC v1.0 collection Sigma See Method Details for shRNA sequences N/A Software and Algorithms Casplab Casplab http://casplab.com/download Fuij ImageJ https://imagej.nih.gov/ij/download.html Image Studio Lite Ver. ..

    CRISPR:

    Article Title: WAPL-Dependent Repair of Damaged DNA Replication Forks Underlies Oncogene-Induced Loss of Sister Chromatid Cohesion.
    Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER RNAimax Life Technologies Cat# 13778 Lipofectamine3000 Invitrogen Cat# L3000001 Demecolcin Sigma Cat# D1925 siR-DNA Spirochrome Cat# SC007 Cell Player 96-well kinetic caspase-3/7 reagent Essen Bioscience Cat# 4440 GFP-Trap agarose beads Chromotek Cat# gta-20 Control agarose beads Chromotek Cat# bab-20 GST–Raf1 RBD coupled to glutathione– agarose beads (Wolthuis et al., 1998) N/A Propidium Iodide Thermo Fisher Scientific Cat# P3566 Puromycin Sigma Cat# P7255 Blasticidin Fisher Scientific Cat# A1113903 G418 Merck Cat# 345810 Doxycycline Sigma Cat# D9891 Fetal Calf Serum Life Technologies Cat# 3100738 Nonessential amino acids Gibco Cat# 11140035 Sodium pyruvate Gibco Cat# 11360039 Penicillin/Streptomycin Gibco/ Life technologies Cat# 15140122 Polybrene Sigma Cat# H9268 Fluor-gel with TES buffer Electron Microscopy Science Cat# 17985–30 Critical Commercial Assays High pure RNA isolation kit Roche Cat# 11828665001 Vysis D7S486/CEP 7 FISH Probe Kit Abbot Molecular Cat# 04N78–020 Experimental Models: Cell Lines TKO-Bcl2-p53KO (TBP) MEFs (Benedict et al., 2018) N/A TBP-WaplKD MEFs This paper N/A TBP-WaplKD #1 MEFs This paper N/A TBP-WaplKD #2 MEFs This paper N/A TBP-Pds5bKD #1 MEFs This paper N/A TBP-Pds5bKD #2 MEFs This paper N/A TBP-NipblKD This paper N/A BJ-hTERT ATCC N/A WaplKD BJ-hTERT This paper N/A HAP1 (Haarhuis et al., 2017) N/A HAP1 WaplKO (Haarhuis et al., 2017) N/A HAP1 Mau2KO (Haarhuis et al., 2017) N/A HAP1 WaplKO Mau2KO (Haarhuis et al., 2017) N/A RPE1-hTERT ATCC N/A RPE1-hTERT-TetOn-Cas9 This paper N/A RPE1-hTERT-TetOn-Cas9_p53KO This paper N/A RPE1-hTERT-TetOn-Cas9-STAG2KO or TP53KO or ESCO2 KO This paper N/A RPE1-hTERT-TetOn-Cas9_p53KO KRAS G12D#1 This paper N/A RPE1-hTERT-TetOn-Cas9_p53KO KRAS G12D#2 This paper N/A RPE1-hTERT-TetOn-Cas9_TetOn-Cas9_SMC3Venus_p53KO This paper N/A Human fibroblast VU-HF1 This paper N/A Human fibroblast VU-HF2 (previously named Fen5280) (Xia et al., 2007) N/A Human fibroblast VU-HF3 This paper N/A (Continued on next page) e2 Developmental Cell 52, 1–16.e1–e7, March 23, 2020 .. REAGENT or RESOURCE SOURCE IDENTIFIER Human fibroblast VU-HF4 This paper N/A LN9SV (Van der Lelij et al., 2009) N/A ARPE-19, CRL1790, RPTEC and HEK293T ATCC N/A HNSCC cells (Stoepker et al., 2015) N/A Lymphoblast HSC93 (Ishida and Buchwald, 1982) N/A Primary foreskin keratinocytes EK94-2 transfected with HPV16 and HPV18 (Steenbergen et al., 1996) N/A Colorectal cancer cell lines SW620, HCT116, COLO320, WiDr, Caco2 ATCC N/A Retinoblastoma cell lines Y79 and Weri (McFall et al., 1977; Reid et al., 1974) N/A Breast cancer cell lines Gift from J. Martens (Erasmus MC, Rotterdam) N/A VU-SCC78 + TetOn-STAG2 This paper N/A OCUB-M + TetOn-STAG2 This paper N/A Oligonucleotides For QPCR primer sequences, see section ‘Quantitative PCR’ This paper N/A For siRNA sequences, see section ‘siRNA experiments’ This paper N/A For other primers, see ‘Method Details’ This paper N/A ssODN template for the introduction of the KRASG12D mutation, see ‘CRISPR-Cas9 gene editing’ section This paper N/A Recombinant DNA pLVX-TRE3G Clontech Cat# 631187 pLVX-Tet3G Clontech Cat# 631187 Lenti-X HT packaging system Clontech Cat# 631276 pLenti-GFP-Muc18 Gift from P. Heutink N/A CSII-EF-MCS-mKO-hCdt1 (30/120) and CSII-EFMCS-mAG-hGem (1/110) Gift from A. Miyawaki (Sakaue-Sawano et al., 2008) N/A pBABE-blasticidin-H2B-YFP Gift from A. Koch N/A pLenti Gift from P. Heutink N/A pLKO.1 shRNA vectors from the Mission mouse TRC v1.0 collection Sigma See Method Details for shRNA sequences N/A Software and Algorithms Casplab Casplab http://casplab.com/download Fuij ImageJ https://imagej.nih.gov/ij/download.html Image Studio Lite Ver. ..

    Recombinant:

    Article Title: WAPL-Dependent Repair of Damaged DNA Replication Forks Underlies Oncogene-Induced Loss of Sister Chromatid Cohesion.
    Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER RNAimax Life Technologies Cat# 13778 Lipofectamine3000 Invitrogen Cat# L3000001 Demecolcin Sigma Cat# D1925 siR-DNA Spirochrome Cat# SC007 Cell Player 96-well kinetic caspase-3/7 reagent Essen Bioscience Cat# 4440 GFP-Trap agarose beads Chromotek Cat# gta-20 Control agarose beads Chromotek Cat# bab-20 GST–Raf1 RBD coupled to glutathione– agarose beads (Wolthuis et al., 1998) N/A Propidium Iodide Thermo Fisher Scientific Cat# P3566 Puromycin Sigma Cat# P7255 Blasticidin Fisher Scientific Cat# A1113903 G418 Merck Cat# 345810 Doxycycline Sigma Cat# D9891 Fetal Calf Serum Life Technologies Cat# 3100738 Nonessential amino acids Gibco Cat# 11140035 Sodium pyruvate Gibco Cat# 11360039 Penicillin/Streptomycin Gibco/ Life technologies Cat# 15140122 Polybrene Sigma Cat# H9268 Fluor-gel with TES buffer Electron Microscopy Science Cat# 17985–30 Critical Commercial Assays High pure RNA isolation kit Roche Cat# 11828665001 Vysis D7S486/CEP 7 FISH Probe Kit Abbot Molecular Cat# 04N78–020 Experimental Models: Cell Lines TKO-Bcl2-p53KO (TBP) MEFs (Benedict et al., 2018) N/A TBP-WaplKD MEFs This paper N/A TBP-WaplKD #1 MEFs This paper N/A TBP-WaplKD #2 MEFs This paper N/A TBP-Pds5bKD #1 MEFs This paper N/A TBP-Pds5bKD #2 MEFs This paper N/A TBP-NipblKD This paper N/A BJ-hTERT ATCC N/A WaplKD BJ-hTERT This paper N/A HAP1 (Haarhuis et al., 2017) N/A HAP1 WaplKO (Haarhuis et al., 2017) N/A HAP1 Mau2KO (Haarhuis et al., 2017) N/A HAP1 WaplKO Mau2KO (Haarhuis et al., 2017) N/A RPE1-hTERT ATCC N/A RPE1-hTERT-TetOn-Cas9 This paper N/A RPE1-hTERT-TetOn-Cas9_p53KO This paper N/A RPE1-hTERT-TetOn-Cas9-STAG2KO or TP53KO or ESCO2 KO This paper N/A RPE1-hTERT-TetOn-Cas9_p53KO KRAS G12D#1 This paper N/A RPE1-hTERT-TetOn-Cas9_p53KO KRAS G12D#2 This paper N/A RPE1-hTERT-TetOn-Cas9_TetOn-Cas9_SMC3Venus_p53KO This paper N/A Human fibroblast VU-HF1 This paper N/A Human fibroblast VU-HF2 (previously named Fen5280) (Xia et al., 2007) N/A Human fibroblast VU-HF3 This paper N/A (Continued on next page) e2 Developmental Cell 52, 1–16.e1–e7, March 23, 2020 .. REAGENT or RESOURCE SOURCE IDENTIFIER Human fibroblast VU-HF4 This paper N/A LN9SV (Van der Lelij et al., 2009) N/A ARPE-19, CRL1790, RPTEC and HEK293T ATCC N/A HNSCC cells (Stoepker et al., 2015) N/A Lymphoblast HSC93 (Ishida and Buchwald, 1982) N/A Primary foreskin keratinocytes EK94-2 transfected with HPV16 and HPV18 (Steenbergen et al., 1996) N/A Colorectal cancer cell lines SW620, HCT116, COLO320, WiDr, Caco2 ATCC N/A Retinoblastoma cell lines Y79 and Weri (McFall et al., 1977; Reid et al., 1974) N/A Breast cancer cell lines Gift from J. Martens (Erasmus MC, Rotterdam) N/A VU-SCC78 + TetOn-STAG2 This paper N/A OCUB-M + TetOn-STAG2 This paper N/A Oligonucleotides For QPCR primer sequences, see section ‘Quantitative PCR’ This paper N/A For siRNA sequences, see section ‘siRNA experiments’ This paper N/A For other primers, see ‘Method Details’ This paper N/A ssODN template for the introduction of the KRASG12D mutation, see ‘CRISPR-Cas9 gene editing’ section This paper N/A Recombinant DNA pLVX-TRE3G Clontech Cat# 631187 pLVX-Tet3G Clontech Cat# 631187 Lenti-X HT packaging system Clontech Cat# 631276 pLenti-GFP-Muc18 Gift from P. Heutink N/A CSII-EF-MCS-mKO-hCdt1 (30/120) and CSII-EFMCS-mAG-hGem (1/110) Gift from A. Miyawaki (Sakaue-Sawano et al., 2008) N/A pBABE-blasticidin-H2B-YFP Gift from A. Koch N/A pLenti Gift from P. Heutink N/A pLKO.1 shRNA vectors from the Mission mouse TRC v1.0 collection Sigma See Method Details for shRNA sequences N/A Software and Algorithms Casplab Casplab http://casplab.com/download Fuij ImageJ https://imagej.nih.gov/ij/download.html Image Studio Lite Ver. ..

    shRNA:

    Article Title: WAPL-Dependent Repair of Damaged DNA Replication Forks Underlies Oncogene-Induced Loss of Sister Chromatid Cohesion.
    Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER RNAimax Life Technologies Cat# 13778 Lipofectamine3000 Invitrogen Cat# L3000001 Demecolcin Sigma Cat# D1925 siR-DNA Spirochrome Cat# SC007 Cell Player 96-well kinetic caspase-3/7 reagent Essen Bioscience Cat# 4440 GFP-Trap agarose beads Chromotek Cat# gta-20 Control agarose beads Chromotek Cat# bab-20 GST–Raf1 RBD coupled to glutathione– agarose beads (Wolthuis et al., 1998) N/A Propidium Iodide Thermo Fisher Scientific Cat# P3566 Puromycin Sigma Cat# P7255 Blasticidin Fisher Scientific Cat# A1113903 G418 Merck Cat# 345810 Doxycycline Sigma Cat# D9891 Fetal Calf Serum Life Technologies Cat# 3100738 Nonessential amino acids Gibco Cat# 11140035 Sodium pyruvate Gibco Cat# 11360039 Penicillin/Streptomycin Gibco/ Life technologies Cat# 15140122 Polybrene Sigma Cat# H9268 Fluor-gel with TES buffer Electron Microscopy Science Cat# 17985–30 Critical Commercial Assays High pure RNA isolation kit Roche Cat# 11828665001 Vysis D7S486/CEP 7 FISH Probe Kit Abbot Molecular Cat# 04N78–020 Experimental Models: Cell Lines TKO-Bcl2-p53KO (TBP) MEFs (Benedict et al., 2018) N/A TBP-WaplKD MEFs This paper N/A TBP-WaplKD #1 MEFs This paper N/A TBP-WaplKD #2 MEFs This paper N/A TBP-Pds5bKD #1 MEFs This paper N/A TBP-Pds5bKD #2 MEFs This paper N/A TBP-NipblKD This paper N/A BJ-hTERT ATCC N/A WaplKD BJ-hTERT This paper N/A HAP1 (Haarhuis et al., 2017) N/A HAP1 WaplKO (Haarhuis et al., 2017) N/A HAP1 Mau2KO (Haarhuis et al., 2017) N/A HAP1 WaplKO Mau2KO (Haarhuis et al., 2017) N/A RPE1-hTERT ATCC N/A RPE1-hTERT-TetOn-Cas9 This paper N/A RPE1-hTERT-TetOn-Cas9_p53KO This paper N/A RPE1-hTERT-TetOn-Cas9-STAG2KO or TP53KO or ESCO2 KO This paper N/A RPE1-hTERT-TetOn-Cas9_p53KO KRAS G12D#1 This paper N/A RPE1-hTERT-TetOn-Cas9_p53KO KRAS G12D#2 This paper N/A RPE1-hTERT-TetOn-Cas9_TetOn-Cas9_SMC3Venus_p53KO This paper N/A Human fibroblast VU-HF1 This paper N/A Human fibroblast VU-HF2 (previously named Fen5280) (Xia et al., 2007) N/A Human fibroblast VU-HF3 This paper N/A (Continued on next page) e2 Developmental Cell 52, 1–16.e1–e7, March 23, 2020 .. REAGENT or RESOURCE SOURCE IDENTIFIER Human fibroblast VU-HF4 This paper N/A LN9SV (Van der Lelij et al., 2009) N/A ARPE-19, CRL1790, RPTEC and HEK293T ATCC N/A HNSCC cells (Stoepker et al., 2015) N/A Lymphoblast HSC93 (Ishida and Buchwald, 1982) N/A Primary foreskin keratinocytes EK94-2 transfected with HPV16 and HPV18 (Steenbergen et al., 1996) N/A Colorectal cancer cell lines SW620, HCT116, COLO320, WiDr, Caco2 ATCC N/A Retinoblastoma cell lines Y79 and Weri (McFall et al., 1977; Reid et al., 1974) N/A Breast cancer cell lines Gift from J. Martens (Erasmus MC, Rotterdam) N/A VU-SCC78 + TetOn-STAG2 This paper N/A OCUB-M + TetOn-STAG2 This paper N/A Oligonucleotides For QPCR primer sequences, see section ‘Quantitative PCR’ This paper N/A For siRNA sequences, see section ‘siRNA experiments’ This paper N/A For other primers, see ‘Method Details’ This paper N/A ssODN template for the introduction of the KRASG12D mutation, see ‘CRISPR-Cas9 gene editing’ section This paper N/A Recombinant DNA pLVX-TRE3G Clontech Cat# 631187 pLVX-Tet3G Clontech Cat# 631187 Lenti-X HT packaging system Clontech Cat# 631276 pLenti-GFP-Muc18 Gift from P. Heutink N/A CSII-EF-MCS-mKO-hCdt1 (30/120) and CSII-EFMCS-mAG-hGem (1/110) Gift from A. Miyawaki (Sakaue-Sawano et al., 2008) N/A pBABE-blasticidin-H2B-YFP Gift from A. Koch N/A pLenti Gift from P. Heutink N/A pLKO.1 shRNA vectors from the Mission mouse TRC v1.0 collection Sigma See Method Details for shRNA sequences N/A Software and Algorithms Casplab Casplab http://casplab.com/download Fuij ImageJ https://imagej.nih.gov/ij/download.html Image Studio Lite Ver. ..

    Software:

    Article Title: WAPL-Dependent Repair of Damaged DNA Replication Forks Underlies Oncogene-Induced Loss of Sister Chromatid Cohesion.
    Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER RNAimax Life Technologies Cat# 13778 Lipofectamine3000 Invitrogen Cat# L3000001 Demecolcin Sigma Cat# D1925 siR-DNA Spirochrome Cat# SC007 Cell Player 96-well kinetic caspase-3/7 reagent Essen Bioscience Cat# 4440 GFP-Trap agarose beads Chromotek Cat# gta-20 Control agarose beads Chromotek Cat# bab-20 GST–Raf1 RBD coupled to glutathione– agarose beads (Wolthuis et al., 1998) N/A Propidium Iodide Thermo Fisher Scientific Cat# P3566 Puromycin Sigma Cat# P7255 Blasticidin Fisher Scientific Cat# A1113903 G418 Merck Cat# 345810 Doxycycline Sigma Cat# D9891 Fetal Calf Serum Life Technologies Cat# 3100738 Nonessential amino acids Gibco Cat# 11140035 Sodium pyruvate Gibco Cat# 11360039 Penicillin/Streptomycin Gibco/ Life technologies Cat# 15140122 Polybrene Sigma Cat# H9268 Fluor-gel with TES buffer Electron Microscopy Science Cat# 17985–30 Critical Commercial Assays High pure RNA isolation kit Roche Cat# 11828665001 Vysis D7S486/CEP 7 FISH Probe Kit Abbot Molecular Cat# 04N78–020 Experimental Models: Cell Lines TKO-Bcl2-p53KO (TBP) MEFs (Benedict et al., 2018) N/A TBP-WaplKD MEFs This paper N/A TBP-WaplKD #1 MEFs This paper N/A TBP-WaplKD #2 MEFs This paper N/A TBP-Pds5bKD #1 MEFs This paper N/A TBP-Pds5bKD #2 MEFs This paper N/A TBP-NipblKD This paper N/A BJ-hTERT ATCC N/A WaplKD BJ-hTERT This paper N/A HAP1 (Haarhuis et al., 2017) N/A HAP1 WaplKO (Haarhuis et al., 2017) N/A HAP1 Mau2KO (Haarhuis et al., 2017) N/A HAP1 WaplKO Mau2KO (Haarhuis et al., 2017) N/A RPE1-hTERT ATCC N/A RPE1-hTERT-TetOn-Cas9 This paper N/A RPE1-hTERT-TetOn-Cas9_p53KO This paper N/A RPE1-hTERT-TetOn-Cas9-STAG2KO or TP53KO or ESCO2 KO This paper N/A RPE1-hTERT-TetOn-Cas9_p53KO KRAS G12D#1 This paper N/A RPE1-hTERT-TetOn-Cas9_p53KO KRAS G12D#2 This paper N/A RPE1-hTERT-TetOn-Cas9_TetOn-Cas9_SMC3Venus_p53KO This paper N/A Human fibroblast VU-HF1 This paper N/A Human fibroblast VU-HF2 (previously named Fen5280) (Xia et al., 2007) N/A Human fibroblast VU-HF3 This paper N/A (Continued on next page) e2 Developmental Cell 52, 1–16.e1–e7, March 23, 2020 .. REAGENT or RESOURCE SOURCE IDENTIFIER Human fibroblast VU-HF4 This paper N/A LN9SV (Van der Lelij et al., 2009) N/A ARPE-19, CRL1790, RPTEC and HEK293T ATCC N/A HNSCC cells (Stoepker et al., 2015) N/A Lymphoblast HSC93 (Ishida and Buchwald, 1982) N/A Primary foreskin keratinocytes EK94-2 transfected with HPV16 and HPV18 (Steenbergen et al., 1996) N/A Colorectal cancer cell lines SW620, HCT116, COLO320, WiDr, Caco2 ATCC N/A Retinoblastoma cell lines Y79 and Weri (McFall et al., 1977; Reid et al., 1974) N/A Breast cancer cell lines Gift from J. Martens (Erasmus MC, Rotterdam) N/A VU-SCC78 + TetOn-STAG2 This paper N/A OCUB-M + TetOn-STAG2 This paper N/A Oligonucleotides For QPCR primer sequences, see section ‘Quantitative PCR’ This paper N/A For siRNA sequences, see section ‘siRNA experiments’ This paper N/A For other primers, see ‘Method Details’ This paper N/A ssODN template for the introduction of the KRASG12D mutation, see ‘CRISPR-Cas9 gene editing’ section This paper N/A Recombinant DNA pLVX-TRE3G Clontech Cat# 631187 pLVX-Tet3G Clontech Cat# 631187 Lenti-X HT packaging system Clontech Cat# 631276 pLenti-GFP-Muc18 Gift from P. Heutink N/A CSII-EF-MCS-mKO-hCdt1 (30/120) and CSII-EFMCS-mAG-hGem (1/110) Gift from A. Miyawaki (Sakaue-Sawano et al., 2008) N/A pBABE-blasticidin-H2B-YFP Gift from A. Koch N/A pLenti Gift from P. Heutink N/A pLKO.1 shRNA vectors from the Mission mouse TRC v1.0 collection Sigma See Method Details for shRNA sequences N/A Software and Algorithms Casplab Casplab http://casplab.com/download Fuij ImageJ https://imagej.nih.gov/ij/download.html Image Studio Lite Ver. ..

    Plasmid Preparation:

    Article Title: An Isothermal, Multiplex Amplification Assay for Detection and Genotyping of Human Papillomaviruses in Formalin-Fixed, Paraffin-Embedded Tissues
    Article Snippet: Human placental DNA (Sigma-Aldrich, St. Louis, MO) was included as HPV-negative controls. .. HPV16 (ATCC 45113D), HPV18 (ATCC45152D), HPV35 (ATCC 40331), HPV52 (ATCC VRMC-29), and HPV56 (ATCC 40549) plasmid constructs were purchased from ATCC. .. HPV genotype-specific target fragments for HPV31, 33, 39, 45, 51, 53, 58, 59, 66, and 68 were synthesized and cloned into pBlueScript II KS(+) vector (Sangon Biotech, Shanghai, China), and the resulting plasmids were amplified in Escherichia coli .

    Construct:

    Article Title: An Isothermal, Multiplex Amplification Assay for Detection and Genotyping of Human Papillomaviruses in Formalin-Fixed, Paraffin-Embedded Tissues
    Article Snippet: Human placental DNA (Sigma-Aldrich, St. Louis, MO) was included as HPV-negative controls. .. HPV16 (ATCC 45113D), HPV18 (ATCC45152D), HPV35 (ATCC 40331), HPV52 (ATCC VRMC-29), and HPV56 (ATCC 40549) plasmid constructs were purchased from ATCC. .. HPV genotype-specific target fragments for HPV31, 33, 39, 45, 51, 53, 58, 59, 66, and 68 were synthesized and cloned into pBlueScript II KS(+) vector (Sangon Biotech, Shanghai, China), and the resulting plasmids were amplified in Escherichia coli .

    other:

    Article Title: An integrated isothermal nucleic acid amplification test to detect HPV16 and HPV18 DNA in resource-limited settings
    Article Snippet: Quantitative synthetic DNA standards were acquired from ATCC for HPV16 (VR-3240SD) and HPV18 (VR-3241SD). gBlocks Gene Fragment (gBlock) DNA for the full E7 genes of HPV6, 11, 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 68, 72, and 82 based on the sequences published in the PapillomaVirus Episteme (PaVE, available: pave.niaid.nih.gov ) ( 47 , 48 ), were purchased from Integrated DNA Technologies, Inc. (IDT, Coralville, IA).



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    Journal: mBio

    Article Title: An mRNA-encoded scFv antibody targeting the helix-α3 of HPV18 E7 oncoprotein as a novel antiviral strategy

    doi: 10.1128/mbio.02627-25

    Figure Lengend Snippet: Characterization of EC₅₀ and binding properties of six anti-HPV18E7 mAbs. ( A ) Antibody against HPV18E7 binding ability analysis by ELISA. Recombinant HPV18 E7 protein (1 μg/mL) was coated onto plates. Commercial antibody F-7 as a positive control. ( B ) Competition matrix of six anti-E7 mAbs. Yellow box and black lines indicate mAbs that compete with one another. The value contained in each box is 1 − a given mAb/control across duplicate; a value <0.5 was taken to be negative binding of the detection mAb. Non-competing pairs are shown in green. ( C ) Kinetic parameters for the binding of each antibody to HPV18E7, as determined by surface plasmon resonance analysis. ( D ) Six Anti-E7 mAbs could recognize HPV18 E7 protein in the HPV18-positive HeLa cells by immunofluorescence. Scale bar = 20 um. Commercial antibody F-7 as a positive control. No primary antibody group as a negative control. The rectangular artifacts on the right side of F-7, 14E5, and 17F2 are the result of a technical issue that occurred during file conversion. ( E ) Quantitative analysis of panel D . The signal targeting E7 (the green signal in panel D) was quantitatively analyzed using ImageJ and normalized. Y normalization = mean density (experimental group)/average of mean density (negative control). A pairwise difference analysis was performed between the experimental and control groups using t -test, n = 3, * P < 0.05, ** P < 0.01, and *** P < 0.001. ( F ) Western blot showed that Anti-E7 mAbs could recognize endogenously expressed HPV18 E7 protein in the HPV18-positive HeLa cells. β-actin as the control. This figure represents spliced gel segments combined for imaging.

    Article Snippet: The primary antibodies were monoclonal antibodies (2E8, 5C5, 8A7, 9A5, 14E5, and 17F2) and anti-HPV18E7 (Santa Cruz, no. sc-365035; working concentration, 4 μg/mL), anti-HA-HRP (Abcam, no. ab128131, 1:1,000), anti-β-actin-HRP (ProteinTech, HRP-60008, 1:5,000), anti-Rb (CST, no.9309S, 1:100), anti- phospho-Rb (CST, no.8516S, 1:100).

    Techniques: Binding Assay, Enzyme-linked Immunosorbent Assay, Recombinant, Positive Control, Control, SPR Assay, Immunofluorescence, Negative Control, Western Blot, Imaging

    Th17 cells induce CAIX, hexokinase II and SLC2A1 mRNA expression in an oxygen‐dependent manner. SiHa (left), SW756 (middle) and Hela (right) were stimulated with medium (black bars), rhIL‐17 (100 ng/mL, red bars) and conditioned media of in vitro generated Th17 cells (CMTH17, 20%, blue bars) and incubated under normoxic (21% O 2 , striped bars) or hypoxic oxygen conditions (1% O 2 ). After 24 h, the cells were analyzed for (A) CAIX, (B) hexokinase II and (C) SLC2A1 expression by qRT‐PCR analysis and normalized to RPL13A housekeeping gene expression. The quotient of the gen of interest/RPL13A of medium‐ stimulated cells incubated by 21% oxygen was set at 1. Shown are the results (mean + SD) from six independent stimulations. Asterisks (* p < .05, ** p < .01, *** p < .001, **** p < .0001) represent statistical significances. The p ‐value according to the nonparametric Mann–Whitney U ‐test.

    Journal: International Journal of Cancer

    Article Title: Th17 cells favor migration and invasiveness of cervical cancer cells under hypoxia in an IGF2BP2 ‐dependent manner

    doi: 10.1002/ijc.70340

    Figure Lengend Snippet: Th17 cells induce CAIX, hexokinase II and SLC2A1 mRNA expression in an oxygen‐dependent manner. SiHa (left), SW756 (middle) and Hela (right) were stimulated with medium (black bars), rhIL‐17 (100 ng/mL, red bars) and conditioned media of in vitro generated Th17 cells (CMTH17, 20%, blue bars) and incubated under normoxic (21% O 2 , striped bars) or hypoxic oxygen conditions (1% O 2 ). After 24 h, the cells were analyzed for (A) CAIX, (B) hexokinase II and (C) SLC2A1 expression by qRT‐PCR analysis and normalized to RPL13A housekeeping gene expression. The quotient of the gen of interest/RPL13A of medium‐ stimulated cells incubated by 21% oxygen was set at 1. Shown are the results (mean + SD) from six independent stimulations. Asterisks (* p < .05, ** p < .01, *** p < .001, **** p < .0001) represent statistical significances. The p ‐value according to the nonparametric Mann–Whitney U ‐test.

    Article Snippet: HPV18‐positive cervical carcinoma cell lines SW756 (RRID:CVCL_1727), HeLa (RRID:CVCL0030) and HPV16‐positive SiHa cells (RRID:CVCL_0032) were received from ATCC (SW756, SiHa) or DSMZ (HeLa).

    Techniques: Expressing, In Vitro, Generated, Incubation, Quantitative RT-PCR, Gene Expression, MANN-WHITNEY

    Th17‐induced expression of CAIX, SLC2A1 and hexokinase II on protein level in 2D cultures and 3D spheroids. (A, B) 2D monolayers of SiHa, SW756 and HeLa were stimulated with medium or CMTH17 and incubated under hypoxic conditions. Twenty‐four hours later CAIX (A, blue bars) and SLC2A1 (B, purple bars) expression were investigated by IF. Bars represent quantification of relative fluorescence/cell of 20 independent pictures (scale bar: 20 μm) From n = 2 independent experiments. The values of medium‐stimulated cells by 21% oxygen was set at 1%. (C) SiHa (left), SW756 (middle) and HeLa cells (right) were stimulated with 100 ng/mL rhIL‐17 (red bars) or 20% CMTH17 (blue bars) for 24 h by hypoxic conditions. Whole cell extracts were analyzed for hexokinase II expression by Western blot analysis. The relative hexokinase II expression (hexokinase/β‐Actin) in cells incubated by 1% oxygen was set at 1. β‐Actin was used as a loading control. Shown are the results (mean + SD) from four independent stimulations. (D–F) 3D spheroids of SW756 and HeLa were generated over 11 days in the presence of medium, rhIL‐17 (red bars) or CMTH17 (blue bars). 5 μm sections of fixed paraffin‐embedded spheroids were validated by HE stainings and analyzed for (D) CAIX, (E) SLC2A1 and (F) hexokinase II expression by IF (scale bar: 200 μm). Sections from CMTH17‐stimulated SW756 spheroids were used for CAIX (D) and hexokinase II (F) stainings, sections from medium‐stimulated HeLa spheroids were used for CAIX (D) and SLC2A1 (E) stainings, sections from IL‐17‐stimulated HeLa spheroids were used for SLC2A1 (E) and hexokinase II (F) stainings and sections from CMTH17‐stimulated HeLa spheroids were used for SLC2A1 (E) and hexokinase II (F) stainings. Bars represent quantification of relative fluorescence/spheroid of n = 6 independent spheroids, respectively (mean + SD). Asterisks (* p < .05, ** p < .01, **** p < .0001) represent statistical significances. The p ‐value according to the nonparametric Mann–Whitney U ‐test.

    Journal: International Journal of Cancer

    Article Title: Th17 cells favor migration and invasiveness of cervical cancer cells under hypoxia in an IGF2BP2 ‐dependent manner

    doi: 10.1002/ijc.70340

    Figure Lengend Snippet: Th17‐induced expression of CAIX, SLC2A1 and hexokinase II on protein level in 2D cultures and 3D spheroids. (A, B) 2D monolayers of SiHa, SW756 and HeLa were stimulated with medium or CMTH17 and incubated under hypoxic conditions. Twenty‐four hours later CAIX (A, blue bars) and SLC2A1 (B, purple bars) expression were investigated by IF. Bars represent quantification of relative fluorescence/cell of 20 independent pictures (scale bar: 20 μm) From n = 2 independent experiments. The values of medium‐stimulated cells by 21% oxygen was set at 1%. (C) SiHa (left), SW756 (middle) and HeLa cells (right) were stimulated with 100 ng/mL rhIL‐17 (red bars) or 20% CMTH17 (blue bars) for 24 h by hypoxic conditions. Whole cell extracts were analyzed for hexokinase II expression by Western blot analysis. The relative hexokinase II expression (hexokinase/β‐Actin) in cells incubated by 1% oxygen was set at 1. β‐Actin was used as a loading control. Shown are the results (mean + SD) from four independent stimulations. (D–F) 3D spheroids of SW756 and HeLa were generated over 11 days in the presence of medium, rhIL‐17 (red bars) or CMTH17 (blue bars). 5 μm sections of fixed paraffin‐embedded spheroids were validated by HE stainings and analyzed for (D) CAIX, (E) SLC2A1 and (F) hexokinase II expression by IF (scale bar: 200 μm). Sections from CMTH17‐stimulated SW756 spheroids were used for CAIX (D) and hexokinase II (F) stainings, sections from medium‐stimulated HeLa spheroids were used for CAIX (D) and SLC2A1 (E) stainings, sections from IL‐17‐stimulated HeLa spheroids were used for SLC2A1 (E) and hexokinase II (F) stainings and sections from CMTH17‐stimulated HeLa spheroids were used for SLC2A1 (E) and hexokinase II (F) stainings. Bars represent quantification of relative fluorescence/spheroid of n = 6 independent spheroids, respectively (mean + SD). Asterisks (* p < .05, ** p < .01, **** p < .0001) represent statistical significances. The p ‐value according to the nonparametric Mann–Whitney U ‐test.

    Article Snippet: HPV18‐positive cervical carcinoma cell lines SW756 (RRID:CVCL_1727), HeLa (RRID:CVCL0030) and HPV16‐positive SiHa cells (RRID:CVCL_0032) were received from ATCC (SW756, SiHa) or DSMZ (HeLa).

    Techniques: Expressing, Incubation, Fluorescence, Western Blot, Control, Generated, MANN-WHITNEY

    Th17 cells mediate enhanced glucose uptake as well as proliferation and migration of cervical cancer cells under hypoxic conditions. (A) HeLa, SiHa and SW756 cells were stimulated with rhIL‐17 (100 ng) or CMTH17 and cultured in normoxic (21% oxygen) or hypoxic (1% oxygen) conditions. After 24 h, the cells were incubated with the fluorescent glucose analogon NBDG (10 mg/mL diluted in PBS) for 1 h. Glucose uptake was investigated by IF. Bars represent quantification of relative fluorescence/cell of 150 cells in total from five independent pictures (scale bar: 200 μm) of independent stimulation experiments, respectively. The values of medium‐stimulated cells incubated by 21% oxygen was set at 1. (B, C) Monolayers of SiHa cells stimulated with medium (black lines), rhIL‐17 (red lines), CMTH17 (blue lines) were scratched and (B) stimulated with 300 μM cobalt chloride every 24 h to create and retain hypoxic conditions or (C) incubated by hypoxic conditions (1% O 2 ). Pictures of the scratches were taken after 0, 24, 48, and 72 h (scale bar: 200 μm). The relative loss of area was calculated for (B) 3 days or (C) 2 days, respectively, indicated by lines (left picture), in relation to time point 0 h (middle, line graphics). The relative loss of the area after 72 or 48 h of the cells stimulated with medium was set at 1 (right, bar chart). The line graphics show one representative experiment performed in doubles (mean ± SD). Bars represent data (mean + SD) of n = 3 independent experiments performed in doubles. (D) SiHa cells were stimulated with medium (black bars), rhIL‐17 (red bars) or CMTH17 (blue bars) and cultured in normoxic (21% oxygen) or hypoxic (1% oxygen) conditions. After 24 h, cells were used in transwell migration assays. Transmigrated cells were calculated after 24 h. Representative pictures (left panel; scale bar: 100 μm); Quantification of n = 3 experiments with six independent pictures, respectively (mean ± SD), lower panel. The number of medium stimulated normoxic cells was set at 1. Asterisks (* p < .05, ** p < .01, **** p < .0001) represent statistical significances. The p ‐value according to the nonparametric Mann–Whitney U ‐test.

    Journal: International Journal of Cancer

    Article Title: Th17 cells favor migration and invasiveness of cervical cancer cells under hypoxia in an IGF2BP2 ‐dependent manner

    doi: 10.1002/ijc.70340

    Figure Lengend Snippet: Th17 cells mediate enhanced glucose uptake as well as proliferation and migration of cervical cancer cells under hypoxic conditions. (A) HeLa, SiHa and SW756 cells were stimulated with rhIL‐17 (100 ng) or CMTH17 and cultured in normoxic (21% oxygen) or hypoxic (1% oxygen) conditions. After 24 h, the cells were incubated with the fluorescent glucose analogon NBDG (10 mg/mL diluted in PBS) for 1 h. Glucose uptake was investigated by IF. Bars represent quantification of relative fluorescence/cell of 150 cells in total from five independent pictures (scale bar: 200 μm) of independent stimulation experiments, respectively. The values of medium‐stimulated cells incubated by 21% oxygen was set at 1. (B, C) Monolayers of SiHa cells stimulated with medium (black lines), rhIL‐17 (red lines), CMTH17 (blue lines) were scratched and (B) stimulated with 300 μM cobalt chloride every 24 h to create and retain hypoxic conditions or (C) incubated by hypoxic conditions (1% O 2 ). Pictures of the scratches were taken after 0, 24, 48, and 72 h (scale bar: 200 μm). The relative loss of area was calculated for (B) 3 days or (C) 2 days, respectively, indicated by lines (left picture), in relation to time point 0 h (middle, line graphics). The relative loss of the area after 72 or 48 h of the cells stimulated with medium was set at 1 (right, bar chart). The line graphics show one representative experiment performed in doubles (mean ± SD). Bars represent data (mean + SD) of n = 3 independent experiments performed in doubles. (D) SiHa cells were stimulated with medium (black bars), rhIL‐17 (red bars) or CMTH17 (blue bars) and cultured in normoxic (21% oxygen) or hypoxic (1% oxygen) conditions. After 24 h, cells were used in transwell migration assays. Transmigrated cells were calculated after 24 h. Representative pictures (left panel; scale bar: 100 μm); Quantification of n = 3 experiments with six independent pictures, respectively (mean ± SD), lower panel. The number of medium stimulated normoxic cells was set at 1. Asterisks (* p < .05, ** p < .01, **** p < .0001) represent statistical significances. The p ‐value according to the nonparametric Mann–Whitney U ‐test.

    Article Snippet: HPV18‐positive cervical carcinoma cell lines SW756 (RRID:CVCL_1727), HeLa (RRID:CVCL0030) and HPV16‐positive SiHa cells (RRID:CVCL_0032) were received from ATCC (SW756, SiHa) or DSMZ (HeLa).

    Techniques: Migration, Cell Culture, Incubation, Fluorescence, MANN-WHITNEY

    Th17 cells increase IGF2BP2 expression on mRNA and protein expression levels in hypoxic cervical cancer cells. (A) SiHa, SW756 and HeLa cells were incubated by normoxic (21%) or hypoxic oxygen conditions (1% O 2 ). After 24 h, the cells were analyzed for IGF2BP2 expression by qRT‐PCR analysis and normalized to RPL13A housekeeping gene expression. The quotient of IFG2BP2/RPL13A of normoxic cells was set at 1. Shown are the results (mean + SD) from four independent experiments. (B, C) SiHa (left), SW756 (middle) and Hela (right) were stimulated with medium (black bars), rhIL‐17 (100 ng/mL, red bars) and conditioned media of in vitro generated Th17 cells (CMTH17, 20%, blue bars) and incubated by hypoxic oxygen conditions (1% O 2 ). (B) After 24 h, the cells were analyzed for IGF2BP2 expression by qRT‐PCR analysis and normalized to RPL13A housekeeping gene expression. The quotient of IFG2BP2/RPL13A of medium stimulated cells was set at 1. Shown are the results (mean + SD) from six independent stimulations. (C) In neutralization experiments, CM were prestimulated with neutralizing anti‐IL‐17 or respective isotype control antibodies for 2 h (light blue bars) and IGF2BP2 expression was analyzed in relation to RPL13A. (D) After 24 h, whole cell extracts were analyzed for IGF2BP2 expression by Western blot analysis. β‐Actin was used as a loading control. The relative IGF2BP2 expression (IGF2BP2/ β‐Actin) of medium stimulated cells was set at 1. Bars represent results (mean + SD) from four independent stimulations. (E) 3D spheroids of HeLa and SW756 were generated over 11 days in the presence of medium, rhIL‐17 (red bars) or CM of Th17 cells (blue bars). The 5‐μm sections of fixed paraffin‐embedded spheroids were validated by HE stainings and analyzed for IGF2BP2 expression by IF (scale bar: 200 μm). Sections from medium‐stimulated SW756 spheroids from Figure were used for IGF2BP2 stainings, sections from medium‐stimulated HeLa spheroids from Figure were used for IGF2BP2 stainings and sections from IL‐17‐stimulated HeLa spheroids from Figure were used for IGF2BP2 stainings. Bars represent quantification of relative fluorescence/spheroid of n = 6 independent spheroids, respectively (mean + SD). Asterisks (* p < .05, ** p < .01, *** p < .001) represent statistical significances. The p ‐value according to the nonparametric Mann–Whitney U ‐test.

    Journal: International Journal of Cancer

    Article Title: Th17 cells favor migration and invasiveness of cervical cancer cells under hypoxia in an IGF2BP2 ‐dependent manner

    doi: 10.1002/ijc.70340

    Figure Lengend Snippet: Th17 cells increase IGF2BP2 expression on mRNA and protein expression levels in hypoxic cervical cancer cells. (A) SiHa, SW756 and HeLa cells were incubated by normoxic (21%) or hypoxic oxygen conditions (1% O 2 ). After 24 h, the cells were analyzed for IGF2BP2 expression by qRT‐PCR analysis and normalized to RPL13A housekeeping gene expression. The quotient of IFG2BP2/RPL13A of normoxic cells was set at 1. Shown are the results (mean + SD) from four independent experiments. (B, C) SiHa (left), SW756 (middle) and Hela (right) were stimulated with medium (black bars), rhIL‐17 (100 ng/mL, red bars) and conditioned media of in vitro generated Th17 cells (CMTH17, 20%, blue bars) and incubated by hypoxic oxygen conditions (1% O 2 ). (B) After 24 h, the cells were analyzed for IGF2BP2 expression by qRT‐PCR analysis and normalized to RPL13A housekeeping gene expression. The quotient of IFG2BP2/RPL13A of medium stimulated cells was set at 1. Shown are the results (mean + SD) from six independent stimulations. (C) In neutralization experiments, CM were prestimulated with neutralizing anti‐IL‐17 or respective isotype control antibodies for 2 h (light blue bars) and IGF2BP2 expression was analyzed in relation to RPL13A. (D) After 24 h, whole cell extracts were analyzed for IGF2BP2 expression by Western blot analysis. β‐Actin was used as a loading control. The relative IGF2BP2 expression (IGF2BP2/ β‐Actin) of medium stimulated cells was set at 1. Bars represent results (mean + SD) from four independent stimulations. (E) 3D spheroids of HeLa and SW756 were generated over 11 days in the presence of medium, rhIL‐17 (red bars) or CM of Th17 cells (blue bars). The 5‐μm sections of fixed paraffin‐embedded spheroids were validated by HE stainings and analyzed for IGF2BP2 expression by IF (scale bar: 200 μm). Sections from medium‐stimulated SW756 spheroids from Figure were used for IGF2BP2 stainings, sections from medium‐stimulated HeLa spheroids from Figure were used for IGF2BP2 stainings and sections from IL‐17‐stimulated HeLa spheroids from Figure were used for IGF2BP2 stainings. Bars represent quantification of relative fluorescence/spheroid of n = 6 independent spheroids, respectively (mean + SD). Asterisks (* p < .05, ** p < .01, *** p < .001) represent statistical significances. The p ‐value according to the nonparametric Mann–Whitney U ‐test.

    Article Snippet: HPV18‐positive cervical carcinoma cell lines SW756 (RRID:CVCL_1727), HeLa (RRID:CVCL0030) and HPV16‐positive SiHa cells (RRID:CVCL_0032) were received from ATCC (SW756, SiHa) or DSMZ (HeLa).

    Techniques: Expressing, Incubation, Quantitative RT-PCR, Gene Expression, In Vitro, Generated, Neutralization, Control, Western Blot, Fluorescence, MANN-WHITNEY

    Th17‐hypoxia‐induced enhanced migration and invasion is dependent on IGF2BP2. (A) SiHa cells were transfected with IGF2BP2‐specific siRNA (siIGF2BP2 #1, #2) or siControl and stimulated with 100 ng/mL rhIL‐17 or CMTH17 for 24 h under hypoxic conditions. Whole cell extracts were analyzed for IGF2BP2 expression by Western blot analysis. β‐Actin was used as a loading control. The relative IGF2BP2 expression (IGF2BP2/β‐Actin) of medium stimulated cells was set at 1. (B, C) SiIGF2BP2 (#1, #2) transfected SiHa cells were scratched and stimulated with medium, rhIL‐17 or CMTH17 24 h post‐transfection. (B) Pictures of the scratches were taken after 0, 24, 48, and 72 h (scale bar: 200 μm). The relative loss of area was calculated for 3 days, indicated by lines, in relation to time point 0 h. The relative loss of the area after 72 h of the cells stimulated with medium was set at 1. (C) Bars represent data (mean + SD) of n = 3 independent experiments performed in doubles. (D, E) SiHa cells were stimulated with IGF2BP2 inhibitors (IGF2BP2 inihibitors#1, #2, 50 μM) for 1 h. Cells were scratched followed by stimulation with medium, rhIL‐17 or CMTH17 in the presence of IGF2BP2 inhibitors (IGF2BP2 inihibitors#1, #2, 25 μM for 72 h). Pictures of the scratches were taken after 0, 24, and 48 h (scale bar: 200 μm). The relative loss of area was calculated for 2 days, indicated by lines, in relation to time point 0 h. The relative loss of the area after 48 h of the cells stimulated with medium was set at 1. (E) Bars represent data (mean + SD) of n = 3 independent experiments performed in doubles. (F–H) Spheroids of SW756 cells and (I–K) spheroids of HeLa cells were generated in the absence or presence of rhIL‐17, CMTH17 cells, medium or CM of naive CD4 + cells. On day 5, spheroids were incubated with inhibitors (#1/#2, 25 μM) or DMSO as a control for 24 h. On day 6, spheroids were embedded into Matrigel. Pictures were taken for 8 days and spheroid invasion was calculated. Shown are the pictures of one representative experiment which depicts the invasiveness of the spheroids over the time (F–J, scale bar: 200 μm). The relative invasiveness after 8 days was determined in relation to medium or CM of naive CD4 + T cells, respectively and DMSO stimulated cells which was set at 1 (H, K, gray stripped bars). Due to the invasiveness of spheroids from HeLa cells on day 8, pictures were merged with the Microsoft Image Composite Editor program. Shown are the results (mean + SD) from six independent spheroids. Asterisks (* p < .05, ** p < .01, *** p < .001, **** p < .0001, n.s. = not significant) represent statistical significances. The p ‐value according to the nonparametric Mann–Whitney U ‐test.

    Journal: International Journal of Cancer

    Article Title: Th17 cells favor migration and invasiveness of cervical cancer cells under hypoxia in an IGF2BP2 ‐dependent manner

    doi: 10.1002/ijc.70340

    Figure Lengend Snippet: Th17‐hypoxia‐induced enhanced migration and invasion is dependent on IGF2BP2. (A) SiHa cells were transfected with IGF2BP2‐specific siRNA (siIGF2BP2 #1, #2) or siControl and stimulated with 100 ng/mL rhIL‐17 or CMTH17 for 24 h under hypoxic conditions. Whole cell extracts were analyzed for IGF2BP2 expression by Western blot analysis. β‐Actin was used as a loading control. The relative IGF2BP2 expression (IGF2BP2/β‐Actin) of medium stimulated cells was set at 1. (B, C) SiIGF2BP2 (#1, #2) transfected SiHa cells were scratched and stimulated with medium, rhIL‐17 or CMTH17 24 h post‐transfection. (B) Pictures of the scratches were taken after 0, 24, 48, and 72 h (scale bar: 200 μm). The relative loss of area was calculated for 3 days, indicated by lines, in relation to time point 0 h. The relative loss of the area after 72 h of the cells stimulated with medium was set at 1. (C) Bars represent data (mean + SD) of n = 3 independent experiments performed in doubles. (D, E) SiHa cells were stimulated with IGF2BP2 inhibitors (IGF2BP2 inihibitors#1, #2, 50 μM) for 1 h. Cells were scratched followed by stimulation with medium, rhIL‐17 or CMTH17 in the presence of IGF2BP2 inhibitors (IGF2BP2 inihibitors#1, #2, 25 μM for 72 h). Pictures of the scratches were taken after 0, 24, and 48 h (scale bar: 200 μm). The relative loss of area was calculated for 2 days, indicated by lines, in relation to time point 0 h. The relative loss of the area after 48 h of the cells stimulated with medium was set at 1. (E) Bars represent data (mean + SD) of n = 3 independent experiments performed in doubles. (F–H) Spheroids of SW756 cells and (I–K) spheroids of HeLa cells were generated in the absence or presence of rhIL‐17, CMTH17 cells, medium or CM of naive CD4 + cells. On day 5, spheroids were incubated with inhibitors (#1/#2, 25 μM) or DMSO as a control for 24 h. On day 6, spheroids were embedded into Matrigel. Pictures were taken for 8 days and spheroid invasion was calculated. Shown are the pictures of one representative experiment which depicts the invasiveness of the spheroids over the time (F–J, scale bar: 200 μm). The relative invasiveness after 8 days was determined in relation to medium or CM of naive CD4 + T cells, respectively and DMSO stimulated cells which was set at 1 (H, K, gray stripped bars). Due to the invasiveness of spheroids from HeLa cells on day 8, pictures were merged with the Microsoft Image Composite Editor program. Shown are the results (mean + SD) from six independent spheroids. Asterisks (* p < .05, ** p < .01, *** p < .001, **** p < .0001, n.s. = not significant) represent statistical significances. The p ‐value according to the nonparametric Mann–Whitney U ‐test.

    Article Snippet: HPV18‐positive cervical carcinoma cell lines SW756 (RRID:CVCL_1727), HeLa (RRID:CVCL0030) and HPV16‐positive SiHa cells (RRID:CVCL_0032) were received from ATCC (SW756, SiHa) or DSMZ (HeLa).

    Techniques: Migration, Transfection, Expressing, Western Blot, Control, Generated, Incubation, MANN-WHITNEY