Journal: Cell reports

Article Title: Global Analysis of Intercellular Homeodomain Protein Transfer

doi: 10.1016/j.celrep.2019.06.056

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Mouse monoclonal anti-VSX2 (E-12) antibody , Santa Cruz Biotechnology , RRID: AB_10842442.

Techniques: Recombinant, In Vitro, Transfection, Protease Inhibitor, Control, Plasmid Preparation, Software, Expressing, Fluorescence

Journal: Nature Communications

Article Title: Inflammation-induced Id2 promotes plasticity in regulatory T cells

doi: 10.1038/s41467-018-07254-2

Figure Lengend Snippet: Id2 expression inhibits enrichment and transcriptional activity of E2A on Foxp3 promoter. a RT - qPCR analysis of Foxp3 , E2A and Id2 mRNA in in vitro generated T H 0, T H 17, ex-Foxp3 T H 17, and iT reg cells. b ChIP-qPCR analysis for E2A occupancy at three putative E-box sites on Foxp3 promoter regions (−1593 to −1584, −1295 to −1286, and −837 to −829) and negative control (−411 to −244; No E-box site) in iT reg and ex-Foxp3 T H 17 cells. Enrichments are calculated relative to the input chromatin for corresponding sites. c Jurkat cells were transfected with either Foxp3 promoter reporter construct alone or along with expression plasmids encoding E2A and Id2 as indicated, followed by luciferase assay. d Effect of mutagenesis on Foxp3 promoter reporter activity. Jurkat cells were transiently co-transfected with the indicated plasmids for luciferase assay as described. N = Normal E box, X = Mutated E box. NS, not significant, *P < 0.05, **P < 0.005, ***P < 0.001 (Student’s t -test). All data are representative of two or three independent experiments (error bars, s.d.)

Article Snippet: 0.5 μg of the Id2 promoter-luciferase constructs or pGL4.17 empty vector was co-transfected with STAT3 (MR227265; Origene), IRF4 (MR226642; Origene) and BATF (MR222114; Origene) expression plasmids (HEK-293 T) and 1 μg of the Foxp3 promoter-luciferase construct was co-transfected with E2A (MG209745; Origene) and Id2 (MR200792; Origene) expression plasmids (Jurkat).

Techniques: Expressing, Activity Assay, Quantitative RT-PCR, In Vitro, Generated, Negative Control, Transfection, Construct, Luciferase, Mutagenesis

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Loss of RIG-I leads to a functional replacement with MDA5 in the Chinese tree shrew

doi: 10.1073/pnas.1604939113

Figure Lengend Snippet: Antiviral responses in tree shrew cells overexpressing tMDA5 and its mutants. (A) Different stimulatory effects of tMDA5, tMDA5 K188Q, and tMDA5 A402K on the activation of the IFN-β-Luc, NF-κB-Luc, and ISRE-Luc reporters in a dose-dependent manner. The procedures are same to Fig. 4A, except for SeV infection for 12 h. (B) Effects of overexpression of tMDA5, tMDA5 K188Q, and tMDA5 A402K on IFN-β-Luc, NF-κB-Luc, and ISRE-Luc reporters on NDV and VSV-GFP infections. Cells were transfected as A, except with 400 ng expression vector and different virus infections (NDV, MOI = 10; VSV-GFP, MOI = 0.01). (C) Different inhibition effects of tMDA5 and its mutants on VSV-GFP replication. (D and E) Effects of tMDA5 and its mutants on activation of phospho-IRF3, phospho-p65, phospho-IKKα/β, and phospho-IκBα. (F) Quantification of viral RNA bound by the Flag-tagged tMDA5 and its mutants from SeV-infected TSPRCs. The procedure and labels are the same as Fig. 3I. Data are representative of three independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001, Student t test. Bars represent mean ± SEM.

Article Snippet: The following antibodies were used in this study: mouse monoclonal anti-Flag (Abmart; M20008), mouse anti-c-Myc (9E11) (Life Technologies; MA1-16637), mouse monoclonal anti-GFP (EnoGene; E12-152), rabbit monoclonal anti-IRF3 (Cell Signal Technology; 4302), rabbit monoclonal anti-phospho-IRF3 (Ser396) (Cell Signal Technology; 4947), rabbit polyclonal anti-MDA5 (Merck Millipore; ABF210), rabbit polyclonal anti-LGP2 (IBL; 29030), rabbit polyclonal anti-MITA (LifeSpan BioSciences; LS-B7237), and mouse monoclonal anti β-actin (EnoGene; E1C602-2).

Techniques: Activation Assay, Infection, Over Expression, Transfection, Expressing, Plasmid Preparation, Inhibition