Journal: Molecular cell

Article Title: Telomere Loop Dynamics in Chromosome End Protection.

doi: 10.1016/j.molcel.2018.06.025

Figure Lengend Snippet: Figure 1. TRF2 Deletion in MEFs Induces a Transient DDR-Positive and NHEJ-Resistant State (A) Examples of interphase-TIF, metaphase-TIF, and cytogenetic chromosome assays in TRF2F/F CreER LgT MEFs following 4-OHT-induced TRF2 deletion. (B) Quantitation of DDR(+) telomeres by metaphase-TIF assay and telomere fusions by cytogenetic chromosome spreads in TRF2F/F CreER LgT MEFs following 4-OHT treatment (mean ± SD; n = 3 biological replicates of R30 metaphase spreads per replicate). Western blots of whole-cell extracts from the indicated times and ionizing radiation (IR) treated controls are shown. (C) Additional examples of interphase-TIF assays following 4-OHT treatment. (D) Quantitation of interphase-TIF in TRF2F/F CreER LgT MEFs cells following 4-OHT treatment (three biological replicates scoring R50 nuclei per replicate are compiled into a Tukey boxplot; **p < 0.01; ns, not significant; Mann-Whitney test).

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Experimental Models: Cell Lines TRF2Floxed/Floxed Rosa26-CreERT2 pBabeSV40LT murine embryonic fibroblasts Eros Lazzerini-Denchi (The Scripps Research Institute) Okamoto et al., 2013 293T Tracy Bryan (CMRI) N/A HT1080 6TG Eric Stanbridge (University of California at Irvine) N/A HeLa LT Jan Karlseder (Salk Institute) O’Sullivan et al., 2014 IMR90 E6E7 Jan Karlseder (Salk Institute) Cesare et al., 2013 U-2OS Jan Karlseder (Salk Institute) N/A Oligonucleotides (CCCTAA)3 Integrated DNA Technologies N/A Recombinant DNA pVPack-GP Vector Agilent Cat# 217566 pVPack-Ampho Vector Agilent Cat# 217568 pLPC-Myc-TRF2 Addgene Cat# 16066 pLPC-Myc-TRFcT Addgene Cat# 44573 pLKO.1 non-targeting control Addgene Cat# 1864 pLKO.1 TRF2 sh-F The Broad Institute Cat# TRCN0000004811 pLKO.1 TRF2 sh-G The Broad Institute Cat# TRCN0000018358 Software and Algorithms ZEISS Zen Black and Zen Blue ZEISS https://www.zeiss.com/microscopy/int/products/ microscope-software/zen.html Metafer 4 and ISIS Metasystems https://metasystems-international.com/ ImageJ Version 1.51h NIH, USA https://imagej.nih.gov/ij SoftWorX Version 6.5.2 GE Healthcare Life Sciences http://incelldownload.gehealthcare.com/bin/ download_data/SoftWoRx/6.5.2/SoftWoRx.htm IMARIS 3D/4D Version 9.1.2 Bitplane http://www.bitplane.com/imaris/imaris OMX Master Control Software - AcquireSR softWoRx reconstruction and analysis software GE Healthcare Life Sciences http://www.gelifesciences.com/en/au/shop/ cell-imaging-and-analysis/high-and-super- resolution-microscopes Leica Las X Leica Microsystems https://www.leica-microsystems.com/products/ microscope-software/details/product/leica-las-x-ls/ Huygens Professional Edition Scientific Volume Imaging https://svi.nl/Huygens-Professional FACS Diva Version 8 BD Biosciences https://www.bdbiosciences.com/in/instruments/ software/facsdiva/features/overview.jsp FlowJo Version 7.2.2 FlowJo https://www.flowjo.com/solutions/flowjo/downloads/ previous-versions ImageQuant Version TL 8.1 GE Healthcare Life Sciences http://www.gelifesciences.com/en/ir/shop/ protein-analysis/molecular-imaging-for-proteins/ imaging-software/imagequant-tl-8-1-p-00110?current= 29000605 GraphPad Prism Version 5 GraphPad Software https://www.graphpad.com

Techniques: Quantitation Assay, Western Blot, MANN-WHITNEY

Journal: Molecular cell

Article Title: Telomere Loop Dynamics in Chromosome End Protection.

doi: 10.1016/j.molcel.2018.06.025

Figure Lengend Snippet: Figure 3. T-Loop Formation Is Mediated by the TRFH Domain of TRF2 (A) Timeline of the experimentation in (C)–(I). (B) Graphical depiction of TRF1, TRF2, and TRFcT.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Experimental Models: Cell Lines TRF2Floxed/Floxed Rosa26-CreERT2 pBabeSV40LT murine embryonic fibroblasts Eros Lazzerini-Denchi (The Scripps Research Institute) Okamoto et al., 2013 293T Tracy Bryan (CMRI) N/A HT1080 6TG Eric Stanbridge (University of California at Irvine) N/A HeLa LT Jan Karlseder (Salk Institute) O’Sullivan et al., 2014 IMR90 E6E7 Jan Karlseder (Salk Institute) Cesare et al., 2013 U-2OS Jan Karlseder (Salk Institute) N/A Oligonucleotides (CCCTAA)3 Integrated DNA Technologies N/A Recombinant DNA pVPack-GP Vector Agilent Cat# 217566 pVPack-Ampho Vector Agilent Cat# 217568 pLPC-Myc-TRF2 Addgene Cat# 16066 pLPC-Myc-TRFcT Addgene Cat# 44573 pLKO.1 non-targeting control Addgene Cat# 1864 pLKO.1 TRF2 sh-F The Broad Institute Cat# TRCN0000004811 pLKO.1 TRF2 sh-G The Broad Institute Cat# TRCN0000018358 Software and Algorithms ZEISS Zen Black and Zen Blue ZEISS https://www.zeiss.com/microscopy/int/products/ microscope-software/zen.html Metafer 4 and ISIS Metasystems https://metasystems-international.com/ ImageJ Version 1.51h NIH, USA https://imagej.nih.gov/ij SoftWorX Version 6.5.2 GE Healthcare Life Sciences http://incelldownload.gehealthcare.com/bin/ download_data/SoftWoRx/6.5.2/SoftWoRx.htm IMARIS 3D/4D Version 9.1.2 Bitplane http://www.bitplane.com/imaris/imaris OMX Master Control Software - AcquireSR softWoRx reconstruction and analysis software GE Healthcare Life Sciences http://www.gelifesciences.com/en/au/shop/ cell-imaging-and-analysis/high-and-super- resolution-microscopes Leica Las X Leica Microsystems https://www.leica-microsystems.com/products/ microscope-software/details/product/leica-las-x-ls/ Huygens Professional Edition Scientific Volume Imaging https://svi.nl/Huygens-Professional FACS Diva Version 8 BD Biosciences https://www.bdbiosciences.com/in/instruments/ software/facsdiva/features/overview.jsp FlowJo Version 7.2.2 FlowJo https://www.flowjo.com/solutions/flowjo/downloads/ previous-versions ImageQuant Version TL 8.1 GE Healthcare Life Sciences http://www.gelifesciences.com/en/ir/shop/ protein-analysis/molecular-imaging-for-proteins/ imaging-software/imagequant-tl-8-1-p-00110?current= 29000605 GraphPad Prism Version 5 GraphPad Software https://www.graphpad.com

Techniques:

Journal: Molecular cell

Article Title: Telomere Loop Dynamics in Chromosome End Protection.

doi: 10.1016/j.molcel.2018.06.025

Figure Lengend Snippet: Figure 5. Mitotic Arrest Induces an Aurora-B-Dependent Telomere Conformational Change from Looped to Linear, Coinciding with Telomere DDR Activation (A) Timeline of the experimentation in (B)–(G). (B) Examples of metaphase-TIF assays in mitotically arrested HeLa LT TRF2 sh-G cells. (C) Quantitation of the metaphase-TIF assays shown in (B) (mean ± SEM; all data points from two biological replicates of R30 metaphases; **p < 0.01, Mann-Whitney test). (D) Examples of Airyscan microscopy of macromolecular telomere structure in mitotically arrested HeLa LT TRF2 sh-G cells. Scale bar, 1 mm. (E) Quantitation of looped telomeres in mitotically arrested HeLa LT cells. Calculations exclude ambiguous molecules (n = 2 biological replicates of R794 molecules per replicate; line depicts the mean). (F) Determination of trioxsalen and UV cross-linking efficiency in mitotically arrested HeLa LT cells treated with DMSO or Hesperadin (mean ± SEM; n = 3 biological replicates; *p < 0.05; Student’s t test). (G) Length distribution of the telomere molecules scored in (E) (mean ± range; n = two biological replicates scoring R794 molecules per replicate).

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Experimental Models: Cell Lines TRF2Floxed/Floxed Rosa26-CreERT2 pBabeSV40LT murine embryonic fibroblasts Eros Lazzerini-Denchi (The Scripps Research Institute) Okamoto et al., 2013 293T Tracy Bryan (CMRI) N/A HT1080 6TG Eric Stanbridge (University of California at Irvine) N/A HeLa LT Jan Karlseder (Salk Institute) O’Sullivan et al., 2014 IMR90 E6E7 Jan Karlseder (Salk Institute) Cesare et al., 2013 U-2OS Jan Karlseder (Salk Institute) N/A Oligonucleotides (CCCTAA)3 Integrated DNA Technologies N/A Recombinant DNA pVPack-GP Vector Agilent Cat# 217566 pVPack-Ampho Vector Agilent Cat# 217568 pLPC-Myc-TRF2 Addgene Cat# 16066 pLPC-Myc-TRFcT Addgene Cat# 44573 pLKO.1 non-targeting control Addgene Cat# 1864 pLKO.1 TRF2 sh-F The Broad Institute Cat# TRCN0000004811 pLKO.1 TRF2 sh-G The Broad Institute Cat# TRCN0000018358 Software and Algorithms ZEISS Zen Black and Zen Blue ZEISS https://www.zeiss.com/microscopy/int/products/ microscope-software/zen.html Metafer 4 and ISIS Metasystems https://metasystems-international.com/ ImageJ Version 1.51h NIH, USA https://imagej.nih.gov/ij SoftWorX Version 6.5.2 GE Healthcare Life Sciences http://incelldownload.gehealthcare.com/bin/ download_data/SoftWoRx/6.5.2/SoftWoRx.htm IMARIS 3D/4D Version 9.1.2 Bitplane http://www.bitplane.com/imaris/imaris OMX Master Control Software - AcquireSR softWoRx reconstruction and analysis software GE Healthcare Life Sciences http://www.gelifesciences.com/en/au/shop/ cell-imaging-and-analysis/high-and-super- resolution-microscopes Leica Las X Leica Microsystems https://www.leica-microsystems.com/products/ microscope-software/details/product/leica-las-x-ls/ Huygens Professional Edition Scientific Volume Imaging https://svi.nl/Huygens-Professional FACS Diva Version 8 BD Biosciences https://www.bdbiosciences.com/in/instruments/ software/facsdiva/features/overview.jsp FlowJo Version 7.2.2 FlowJo https://www.flowjo.com/solutions/flowjo/downloads/ previous-versions ImageQuant Version TL 8.1 GE Healthcare Life Sciences http://www.gelifesciences.com/en/ir/shop/ protein-analysis/molecular-imaging-for-proteins/ imaging-software/imagequant-tl-8-1-p-00110?current= 29000605 GraphPad Prism Version 5 GraphPad Software https://www.graphpad.com

Techniques: Activation Assay, Quantitation Assay, MANN-WHITNEY, Microscopy