Journal: Cancer cell

Article Title: Binary pan-cancer classes with distinct vulnerabilities defined by pro- or anti-cancer YAP/TEAD activity

doi: 10.1016/j.ccell.2021.06.016

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Recombinant DNA psPax2 Didier Trono Addgene plasmid #12260; RRID: Addgene_12260 pMD2.G Didier Trono Addgene plasmid #12259; RRID: Addgene_12259 LentiCRISPR v2 Sanjana et al., 2014 Addgene plasmid #52961; RRID: Addgene_52961 pLKO.1-puro Stewart et al., 2003 Addgene plasmid #8453; RRID: Addgene_8453 Software and algorithms Kaluza, Flow Cytometry analysis software Beckman Coulter https://www.mybeckman.ca/flow-cytometry/software/kaluza Prism GraphPad https://www.graphpad.com/scientific-software/prism/ RSEM Li and Dewey, 2011 https://deweylab.github.io/RSEM/ DESeq2 Love et al., 2014 https://bioconductor.org/packages/release/bioc/html/DESeq2.html RRHO Plaisier et al., 2010 https://bioconductor.org/packages/release/bioc/html/RRHO.html RRHO2 Cahill et al., 2018 https://github.com/RRHO2/RRHO2 DeepTools Ramírez et al., 2016 https://github.com/deeptools/deeptools Homer package Heinz et al., 2010 http://homer.ucsd.edu/homer/ NMF Gaujoux and Seoighe, 2010 https://cran.r-project.org/web/packages/NMF/index.html fastQC N/A https://www.bioinformatics.babraham.ac.uk/projects/fastqc Bowtie2 Langmead and Salzberg, 2012 http://bowtie-bio.sourceforge.net/bowtie2 SAMtools Li et al., 2009 http://samtools.sourceforge.net MACS2 Zhang et al., 2008 https://github.com/macs3-project/MACS Integrative Genomics Viewer (IGV) Robinson et al., 2011 https://software.broadinstitute.org/software/igv/ UCSC tool bedGraphToBigWig Kent WJ., 2010 https://anaconda.org/bioconda/ucsc-bedgraphtobigwig ReMap Chèneby et al., 2018 http://remap.univ-amu.fr/ GSEA Subramanian, et al., 2005 https://www.gsea-msigdb.org/gsea/index.jsp R package ‘‘ggplot2’’ Wickham, 2016 https://cran.r-project.org/web/packages/ggplot2 R package ‘‘scales’’ N/A https://scales.r-lib.org Open in a separate window KEY RESOURCES TABLE YAP/TAZ silencing in multiple cancers reflects their tumor suppressor activity Cancers can be parsed into binary YAP on and YAP off classes Binary transcriptomes drive distinct adhesive, metabolic, genetic, and drug profiles Alternate YAP enhancers explain anti-cancer integrin-aVb5 program in YAP off cancers

Techniques: Plasmid Preparation, Derivative Assay, Recombinant, Electron Microscopy, Staining, SYBR Green Assay, Sample Prep, Microarray, Expressing, CRISPR, Software, Flow Cytometry

Journal: Current biology : CB

Article Title: Profilin-mediated actin allocation regulates the growth of epithelial microvilli

doi: 10.1016/j.cub.2019.08.051

Figure Lengend Snippet: Key Resources Table

Article Snippet: ​ REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies anti-β-actin Sigma Aldrich A5316 anti-pan-actin Cytoskeleton Inc. AAN01 anti-α-tubulin Santa Cruz Biotechnology B-5-1-2 anti-GAPDH Cell Signaling Inc. 14C10 anti-profilin-1 Sigma Aldrich P7624 anti-G-actin DSHB JLA20 Donkey anti-mouse 800 IRdye LI-COR Biosciences 926–32212 Donkey anti-rabbit 800 IRdye LI-COR Biosciences 926–32213 Goat anti-rabbit Alexa Fluor 488 Thermo Fisher Scientific A-11008 Goat anti-rabbit Alexa Fluor 568 Thermo Fisher Scientific A-11011 Chemicals CK-666 Calbiochem 182515 Verapamil Sigma Aldrich V1711202 Latrunculin A Thermo Fisher Scientific {"type":"entrez-nucleotide","attrs":{"text":"L12370","term_id":"290229","term_text":"L12370"}} L12370 Blasticidin Invivogen Ant-bl Phleomycin Invivogen Ant-ph Doxycycline RPI {"type":"entrez-nucleotide","attrs":{"text":"D43020","term_id":"3107280","term_text":"D43020"}} D43020 G418 RPI {"type":"entrez-nucleotide","attrs":{"text":"G64000","term_id":"6782527","term_text":"G64000"}} G64000 Isoflurane Piramal Healthcare 66794-017-25 TET-free FBS Atlanta Biological {"type":"entrez-protein","attrs":{"text":"S10350","term_id":"107801","term_text":"pir||S10350"}} S10350 FuGENE 6 Promega E2691 Polybrene Sigma Aldrich TR-1003-G Probes Alexa Fluor 488 Phalloidin Thermo Fisher Scientific A12379 Alexa Fluor 568 Phalloidin Thermo Fisher Scientific A12380 Deoxyribonuclease I, Alexa Fluor 488 conjugate Thermo Fisher Scientific {"type":"entrez-nucleotide","attrs":{"text":"D12371","term_id":"2148597","term_text":"D12371"}} D12371 Experimental Models: Cell Lines LS174T-W4 Dr. Hans Clevers [ 29 ] Experimental Models: Organisms/Strains Mus Musculus (C57BL/6J) Jackson Laboratory 000664 Recombinant DNA pLKO.1 (profilin-1 KD) Sigma Aldrich TRCN0000062803 pLKO.1 (profilin-1 KD) Sigma Aldrich TRCN0000062805 pLKO.1 (scramble) Dr. David Sabatini Addgene #1864 psPAX2 packaging plasmid Dr. Didier Trono Addgene #12260 pMD2.G Dr. Didier Trono Addgene #12259 Software and Algorithms FIJI https://fiji.sc N/A NIS AR Elements Analysis Nikon N/A Prism 8.1.2 www.graphpad.com N/A Open in a separate window Key Resources Table Inhibition of Arp2/3 accelerates growth of microvilli in vivo and in vitro Inhibition of Arp2/3 redistributes F-actin from the cortex into microvilli Profilin-1 is required for normal brush border assembly Acceleration of microvilli growth upon inhibition of Arp2/3 requires profilin-1

Techniques: Recombinant, Plasmid Preparation, Software

Journal: Nature Communications

Article Title: Different hotspot p53 mutants exert distinct phenotypes and predict outcome of colorectal cancer patients

doi: 10.1038/s41467-022-30481-7

Figure Lengend Snippet: a SW480 cells in which the endogenous TP53 genes (harboring R273H and P309S mutations) had been knocked out, were stably transduced with p53 R175H or p53 R273H . b Western blot analysis of p53 in SW480 knockout (KO) cells before and after transduction of p53 R175H or p53 R273H . n = 3. c SW480 TP53 KO cells and their derivatives expressing p53 R175H or p53 R273H were subjected to RNA-seq analysis. Shown is a heatmap of genes differentially expressed (fold change > 1.5, pAdj < 0.05) in p53 R273H overexpressing cells relative to p53 KO and p53 R175H overexpressing cells, n = 3. d Venn diagram of upregulated genes (fold change > 1.5, pAdj < 0.1) in p53 R273H overexpressors relative to p53 KO cells (blue circle) or p53 R175H overexpressors (green circle). The 140 overlapping genes were defined as the ‘R273 signature’. e Western blot analysis of p53 in SW480 cells stably transduced with shRNA directed against the 3’ UTR of the TP53 gene (shp53), followed by stable overexpression of shRNA-resistant p53 R175H or p53 R273H . shc = SW480 cells transduced with control shRNA, to visualize the endogenous p53. n = 3. f , g Gene Set Enrichment Analysis (GSEA) of differentially expressed genes in shp53 cells reconstituted with p53 R273H vs control shp53 cells or shp53 cells reconstituted with p53 R175H (ranked by fold change), using the R273 signature as the tested gene set. ES = Enrichment score. Source data is provided as a Source Data file.

Article Snippet: HT-29 and COGA-5 were infected with recombinant lentiviruses (pLKO.1-puro-shp53, (addgene, 19199)) to produce shRNA directed against the endogenous mutant p53 mRNA. p53 knockdown and mutant protein expression were verified by RT-qPCR and Western blot analysis.

Techniques: Stable Transfection, Transduction, Western Blot, Knock-Out, Expressing, RNA Sequencing, shRNA, Over Expression, Control

Journal: Nature Communications

Article Title: Different hotspot p53 mutants exert distinct phenotypes and predict outcome of colorectal cancer patients

doi: 10.1038/s41467-022-30481-7

Figure Lengend Snippet: a HCT116 cells were subjected to CRISPR/Cas 9 gene editing using RNP and ssODN to knock-in either the p53 R175H or the p53 R273H mutation. Cells which underwent the same process but did not end up with an edited genome, and thus retained wtp53 expression, served as CRISPR control. b RT-qPCR analysis of p21 mRNA in the cells in a . For the CRISPR/Cas9 knock-in clones, values in each experiment were determined separately for each individual clone, normalized to GAPDH mRNA, and then averaged. Values in the figure are displayed relative to the control parental cells, defined as 1.0. Mean ± SEM from four independent experiments. one-way ANOVA and Tukey’s post hoc test. c RT-qPCR analysis of representative R273 signature genes in the cells in a . Values were calculated as in b . Three biological repeats (ITGA7,CDC42EP5), four biological repeats (APOE) or five biological repeats (MRC2, ECM1). d Western blot analysis of HT-29 cells transduced with p53-specific shRNA (Shp53) or control shRNA (Shc). n = 2. e Western blot analysis of COGA-5 cells transduced with p53-specific shRNA (Shp53) or control shRNA (Shc). n = 2. f RT-qPCR analysis of representative R273 signature genes in the cells in d . Values were normalized to GAPDH mRNA and are shown relative to the Shc cells. Mean ± SEM from three independent experiments. Unpaired two-tailed t test. g RT-qPCR analysis of representative R273 signature genes in the cells in e . Values were normalized to GAPDH mRNA and are shown relative to the Shc cells. Mean ± SEM from four independent experiments. Unpaired two-tailed t test. Source data is provided as a Source Data file.

Article Snippet: HT-29 and COGA-5 were infected with recombinant lentiviruses (pLKO.1-puro-shp53, (addgene, 19199)) to produce shRNA directed against the endogenous mutant p53 mRNA. p53 knockdown and mutant protein expression were verified by RT-qPCR and Western blot analysis.

Techniques: CRISPR, Knock-In, Mutagenesis, Expressing, Control, Quantitative RT-PCR, Clone Assay, Western Blot, Transduction, shRNA, Two Tailed Test