Journal: Cell reports

Article Title: Tissue-Specific Regulation of the Wnt/β-Catenin Pathway by PAGE4 Inhibition of Tankyrase.

doi: 10.1016/j.celrep.2020.107922

Figure Lengend Snippet: Figure 2. PAGE4 Inhibits PARylation, Ubiquitination, and Degradation of the TNKS1 Substrate AXIN1 (A) PAGE4 reduces AXIN1 PARylation. Western blot analysis of Myc-IP samples (in duplicate) from HEK293 cells expressing Myc-AXIN1 with FLAG-vector or FLAG-PAGE4. PARylated AXIN1 was detected using Fc-RNF146-WWE reagent. (B) PAGE4 decreases AXIN1 ubiquitination. Western blot analysis of Myc-ubiquitin-IP samples from HEK293 cells expressing Myc-ubiquitin, GFP-PAGE4, and FLAG-AXIN1, as indicated. Cells were treated with 10 mM MG132 6 h before harvesting. (C) PAGE4 stabilizes AXIN1 and TNKS1 proteins. Western blot analysis of endogenous TNKS1 and AXIN1 from HEK293 cells expressing PAGE4 or treated with 5 mM XAV939 (TNKS1 inhibitor) for 24 h. (D) Endogenous PAGE4 stabilizes AXIN1 and TNKS1 proteins. Western blot analysis of endogenous TNKS1 and AXIN1 in AGS parental or AGS PAGE4-knockout (KO) clones. Indicated samples were treated with 10 mM IWR-1 for 24 h. (E and F) PAGE4 regulates AXIN1 and TNKS1 abundance in an RNF146-dependent manner. HuTu80 cells were treated with the indicated siRNAs for 72 h before immunoblotting for the indicated proteins. (E) PAGE4 depletion decreases TNKS1 and AXIN1 levels. (F) Knockdown of RNF146 rescuees AXIN1 and TNKS1 levels upon depletion of PAGE4. (G) A model summarizing PAGE4 orchestration of AXIN1 stabilization via binding with TNKS1. See also Figure S2.

Article Snippet: RWPE-1 ATCC Cat# CRL-11609 HT-1376 ATCC Cat# CRL-1472 U2OS ATCC Cat# HTB-96 HuTu80 ATCC Cat# HTB-40 AGS ATCC Cat# CRL-1739 L-Wnt-3A ATCC Cat# CRL-2647 AGS PAGE4 KO This study N/A HuTu80 PAGE4 KO This study N/A BHPrS1 A gift from Simon Hayward, Vanderbilt University N/A Experimental Models: Organisms/Strains Loxp-stop-loxp-FLAG-PAGE4 Generated in this study using C57BL6 (from Jackson Laboratory) N/A Actb-Cre mice The Jackson Laboratory Cat#019099 TCF/Lef1:H2B-GFP/+ mice The Jackson Laboratory Cat#013752 Experimental Models: Tissues Normal Human Prostate slides US Biomax Cat# HuFPT141 Human Normal Placenta US Biomax Cat# HuFPT080 Normal Human Epididymis US Biomax Custom ordered Normal Human Colon US Biomax Cat# HuFPT036 Normal Human Gonad Array US Biomax Cat# TEN1201 Normal Human Ovary US Biomax Cat# HuFPT076 (Continued on next page) Cell Reports 32, 107922, July 21, 2020 e2

Techniques: Ubiquitin Proteomics, Western Blot, Expressing, Plasmid Preparation, Knock-Out, Clone Assay, Knockdown, Binding Assay

Journal: International journal of molecular sciences

Article Title: Cell Membrane-Integrated Neuroligin-1 Regulates the Anti-Inflammatory Effects of CRC Cell-Derived Exosomes.

doi: 10.3390/ijms26020503

Figure Lengend Snippet: Figure 6. M0 macrophages were treated with the exosomes of the NLG1 expressing (EXO) and NLG1 knock-downed (sh-EXO) CRC cells and the mRNA expression level (Log2 FC) of the CD206 and CD80 genes as phenotypical markers (a) and pro-inflammatory genes including IL8, IL6, IL1β, and TNFa as makers for the macrophage functionality (b), were analyzed using qRT-PCR. Data shown for qRT-PCR are the means ± standard deviation (SD).

Article Snippet: NLG1 silenced HuTu80 cell populations were generated by infection with lentiviral vectors targeting NLG1 from Origene: cat # TL3111634B (shNLGN1b) (pGFP-C-shLenti shRNA-29mer expression vector-ORIGENE, Rockville, MD, USA).

Techniques: Expressing, Quantitative RT-PCR, Standard Deviation

Journal: International journal of molecular sciences

Article Title: Cell Membrane-Integrated Neuroligin-1 Regulates the Anti-Inflammatory Effects of CRC Cell-Derived Exosomes.

doi: 10.3390/ijms26020503

Figure Lengend Snippet: Figure 5. M0 macrophages were treated with exosomes of NLG1-expressing CRC cells (HuTu80 cells), IL13+1L4 (M2-stimuli), LPS+INF-γ (M1-stimuli) for 48 h. The expression levels of CD206, and CD80 as makers for the macrophage phenotype (a), IL8, IL6, TNFa, and IL1β genes as makers for the macrophage functionality (b), were determined using qRT-PCR. Gene expression level (Log2 FC) was calculated relative to untreated M0 macrophages. Data shown are the means ± standard deviation (SD).

Article Snippet: NLG1 silenced HuTu80 cell populations were generated by infection with lentiviral vectors targeting NLG1 from Origene: cat # TL3111634B (shNLGN1b) (pGFP-C-shLenti shRNA-29mer expression vector-ORIGENE, Rockville, MD, USA).

Techniques: Expressing, Quantitative RT-PCR, Gene Expression, Standard Deviation