flag fbxo11  (Addgene inc)

Journal: Cell Reports

Article Title: Aryl Hydrocarbon Receptor Interacting Protein Maintains Germinal Center B Cells through Suppression of BCL6 Degradation

doi: 10.1016/j.celrep.2019.04.014

Figure Lengend Snippet: AIP Protects BCL6 from FBXO11-Mediated Proteasomal Degradation (A) HEK293T cells were transfected with either a FLAG-tagged FBXO11 or empty vector (EV-FLAG), together with MYC-tagged AIP. A majority of the whole-cell extracts (WCEs) were subjected to immunoprecipitation (IP) using FLAG antibody (rows 1 and 2), and the rest of the WCEs were used in immunoblotting (rows 3 and 4). (B) HEK293T cells were transfected with either FLAG-tagged BCL6 or EV-FLAG, together MYC-tagged AIP . A majority of the WCEs were subjected to IP using FLAG antibody (rows 1 and 2), the and the rest of the WCEs were used in immunoblotting (rows 3 and 4). (C) OCI-LY7 DLBCL cells were lysed and immunoprecipitated with antibodies to IgG, AIP, UCHL1 and immunoblottedfor AIP and UCHL1. (D) OCI-LY7 DLBCL cells were lysed and immunoprecipitated with antibodies to IgG, BCL6, UCHL1 and immunoblotted for BCL6 and UCHL1. (E) OCI-LY7 DLBCL cells were lysed and immunoprecipitated with antibodies to IgG, FBXO11 and BCL6 and immunoblotted for BCL6 and FBXO11. β-actin was used as a loading control. (F) HEK293T cells were transfected with MYC-tagged AIP or EV-MYC, FLAG-tagged FBXO11, HIS-tagged BCL6, and hemagglutinin (HA)-tagged ubiquitin. Where indicated, cells were treated with MG132 post-transfection for 2.5 h to inhibit proteasomal degradation. (G) HEK cells were transfected as in (F) and FLAG-tagged UCHL1. Cells were harvested and subjected to IP BCL6. Corresponding WCEs are shown. (H) OC1-LY7 cells were stained with AIP, BCL6, UCHL1, and FBXO11. DAPI was used as a nuclear stain. Arrowheads show areas of co-localization.

Article Snippet: FBXO11-FLAG pCMV3 , SinoBiological , Cat # BC130445.

Techniques: Transfection, Plasmid Preparation, Immunoprecipitation, Western Blot, Staining

Journal: Cell Reports

Article Title: Aryl Hydrocarbon Receptor Interacting Protein Maintains Germinal Center B Cells through Suppression of BCL6 Degradation

doi: 10.1016/j.celrep.2019.04.014

Figure Lengend Snippet: AIP Is Overexpressed in Human DLBCLs (A) AIP is expressed in many tumors, and AIP was found to be most highly expressed in DLBCLs. Data were obtained from http://www.ciobioportal.org . (B) AIP staining from reactive lymph nodes (n = 88 mean 13 ± 2.5) and DLBCL biopsies (n = 74 mean 20 ± 2.5) ∗∗∗ p < 0.0001 (two-tailed Mann-Whitney U test). (C) Survival analysis of DLBCL patients with high and low AIP expression. Data obtained from http://www.genomicscape.com/ . (D) BCL6 and AIP protein expression in DLBCL cell lines. (E) Ratio between AIP and BCL6 expression. EBV, Epstein-Barr-virus-infected B cells; PB, peripheral blood B cells. (F) Lentiviral delivery of scrambled or shRNAi against Aip to OC1-LY7 cells. Cells were examined 48 h after transfection, and the expression of AIP and BCL6 and cell viability were analyzed by flow cytometry. (G) Diagram showing the interaction between FBXO11, UCHL1, BCL6, and AIP. Under genotoxic environments found in GC B cells or DLBCL cells, AIP brings UCHL1 to deubiquitinate BCL6, thus maintaining its expression. In the absence of AIP, BCL6 gets ubiquitinated by FBXO11 and undergoes proteasomal degradation. ∗∗∗ p < 0.0001 (a Mann-Whitney U test).

Article Snippet: FBXO11-FLAG pCMV3 , SinoBiological , Cat # BC130445.

Techniques: Staining, Two Tailed Test, MANN-WHITNEY, Expressing, Infection, Transfection, Flow Cytometry

Journal: Cell Reports

Article Title: Aryl Hydrocarbon Receptor Interacting Protein Maintains Germinal Center B Cells through Suppression of BCL6 Degradation

doi: 10.1016/j.celrep.2019.04.014

Figure Lengend Snippet:

Article Snippet: FBXO11-FLAG pCMV3 , SinoBiological , Cat # BC130445.

Techniques: Plasmid Preparation, Microarray, Recombinant, shRNA, Software