Journal: Oncogene

Article Title: BRCA1 physically associates with p53 and stimulates its transcriptional activity.

doi: 10.1038/sj.onc.1201932

Figure Lengend Snippet: Figure 2 BRCA1 interacts with p53 both in vitro and in vivo. (a) GST or GST-p53 proteins were expressed in E. coli and mixed with in vitro-translated (IVT) 35S-methionine-labeled BRCA1 (lanes 2, 5 and 6) or luciferase (negative control; lanes 1, 3 and 4), precipitated with Glutathione-sepharose beads, and analysed by 7.5% SDS ± PAGE. In lanes 1 and 2, 1/10th of the IVT-labeled proteins used for binding assays were loaded directly. (b) GST-p53 interacts with endogenous BRCA1. GST-p53 expressed as above was mixed with COS-7 cell lysate, precipitated with Glutathione-sepharose beads, separated by 7.5% SDS ± PAGE, and analysed by immunoblotting with anti-BRCA1 (SG11) antibody. (c) Total cell lysate from COS-7 (lane 1), HeLa (lane 2) and U2OS (lane 3) cells were analysed by Western blotting for BRCA1 with anti-BRCA1 (SG11) antibody. (d) Total cell lysate from COS-7 (lane 1), HeLa (lane 2) and U2OS (lane 3) cells were immunoprecipitated with anti-BRCA1 (C-20) and the immunoprecipitates were analysed by anti-BRCA1 (SG11). (e) COS-7 extracts were immunoprecipitated with anti-p16 (lane 2; negative control), anti-BRCA1 antibodies, C20 (lane 3) and SG11 (lane 4), anti-MDM-2 (Ab1) antibody (lane 5: positive control) and anti-p53 (lane 6). (f) HeLa cell extracts were immunoprecipitated with anti-p16 (lane 1), anti-BRCA1 antibodies C20 (lane 2), SG11 (lane 3) and anti-p53 (lane 4). (g) U2OS cell extracts were immunoprecipitated with anti-p16 (lane 2; negative control), anti-BRCA1 antibodies, SG11 (lane 3), anti-MDM-2 antibody (lane 4; positive control) and anti-p53, Ab6 (lane 6). The co-precipitated p53 (Figure 2e, f and g) was detected by immunoblotting with HRP-conjugated anti-p53 antibody as described in Materials and methods. (h) SW480 cell extracts were immunoprecipitated with anti-BRCA1 antibody, Ab1 (lane 1) and IgG (lane 2; negative control). The co-precipitated p53 was detected by immunoblotting with anti-p53 (Ab7) antibody and anti-mouse HRP antibody

Article Snippet: SAOS-2, HeLa, COS7, HBL100, U2OS and 293 cells were obtained from ATCC.

Techniques: In Vitro, In Vivo, Labeling, Luciferase, Negative Control, SDS Page, Binding Assay, Western Blot, Immunoprecipitation, Positive Control

Journal: Virology

Article Title: Genetic analysis of the internal ribosome entry segment of bovine viral diarrhea virus.

doi: 10.1006/viro.1998.9425

Figure Lengend Snippet: FIG. 2. (A) Analysis of 59 end-dependent scanning and IRES-dependent reporter protein expression in vivo. CAT (solid bars) and LUC (striped bars) activity levels in lysates of CV-1 cells infected with vSP6 and transfected with DNA from each of the plasmids as indicated. (B) Effect of poliovirus infection on IRES-dependent translation. Catalytic activity of reporter proteins expressed in CV-1 cells infected with vSP6, transfected with pBi59BVDV (BVDV) or pBi59polio (polio), and superinfected with poliovirus strain Mahoney (cross-hatched bars) or mock-infected (bars with dotted pattern).

Article Snippet: Transfections were carried out at 37°C and 5% CO2 for 2 h. After transfection, the medium containing the DNA–lipid complexes was removed, and cells were supplemented with 1 ml of Dulbecco’s minimum essential medium and 10% fetal bovine serum for an additional 16 h. Poliovirus type I strain Mahoney (American Type Culture Collection) was propagated in Cos-7 cells.

Techniques: Expressing, In Vivo, Activity Assay, Infection, Transfection