Journal: Molecular Pain

Article Title: Transcriptional regulation of metabotropic glutamate receptor 2/3 expression by the NF-κB pathway in primary dorsal root ganglia neurons: a possible mechanism for the analgesic effect of L-acetylcarnitine

doi: 10.1186/1744-8069-2-20

Figure Lengend Snippet: Acetylation of p65/Rel-A induced by L-acetylcarnitine treatment . p65/Rel-A was immunoprecipitated from DRG cultures after L-acetylcarnitine (A) or carnitine (B) treatment. Western blots show immunoreactivity for anti-acetyl-lysine (red fluorescence) and anti-p65 (green fluorescence). A single red band migrating at ~65 kDa was identified on Western blots. Western blots demonstrate that L-acetylcarnitine (A), but not carnitine (B) treatment induces acetylation of the p65/Rel-A protein. (C) Results are expressed as ratio of integrated intensity of acetyl-lysine and p65 bands. * p < 0.05 vs controls (Student's t test).

Article Snippet: The membrane was blocked in Odyssey blocker (Li-COR) for 1 hr and anti-acetyl-lysine (Upstate) and anti-p65/Rel-A (Santa Cruz) primary antibody were simultaneously used for immunoblotting overnight at 4°C.

Techniques: Immunoprecipitation, Western Blot, Fluorescence

Journal: Evidence-based Complementary and Alternative Medicine : eCAM

Article Title: The Anti-Inflammatory Effect of Zhibaidihuang Decoction on Recurrent Oral Ulcer with Sirt1 as the Key Regulatory Target

doi: 10.1155/2021/8886699

Figure Lengend Snippet: IHC analyses of expression of Sirt1, NF- κ B, and TNF- α in oral mucosa. The expressed Sirt1, NF- κ B p65, and TNF- α in oral mucosa of Ct, AZC, and ZBDHD-H rats were immunohistochemically (IHC) analyzed and observed under the microscope (x200). The positive cells were pointed out by arrows.

Article Snippet: The specific antibodies against Sirt1 (sc-74465), NF- κ B p65 (sc-372), Stat3 (sc-482), Foxp3 (sc-28705), acetyl lysine (sc-284922), and TNF- α (sc-4261) were all purchased from Santa Cruz Biotechnology (Texas, USA).

Techniques: Expressing, Microscopy

Journal: Molecular Cancer

Article Title: Tip60-mediated acetylation activates transcription independent apoptotic activity of Abl

doi: 10.1186/1476-4598-10-88

Figure Lengend Snippet: IR exposure induces c-Abl acetylation . (a) HeLa cells were exposed to 5 Gy of IR or 20 J/m2 of UV and collected at 0, 2 and 4 h later. Anti-Abl immunoprecipitates were prepared using K-12 antibody and resolved on a 5-15% SDSPAGE. The proteins were transferred onto Immobilon-P and immune-analyzed with pan-specific anti-acetyl lysine antibody ( top panel ). The membrane was stripped as described in methods and reprobed with anti-Abl antibody ( bottom panel ). (b) Abl -/- 3T3 cells reconstituted with Flag-tagged Abl were either mock, IR (5 Gy) or UV (20 J/m2) treated and collected 4 h later. Anti-Abl immunoprecipitates were prepared and immune-analyzed with acetyl lysine ( top panel ) or Flag antibody ( bottom panel ). (c) Abl -/- 3T3 cells reconstituted with Flag-tagged Abl were labeled with 14C acetyl CoA for 2 h and then exposed to IR (5 Gy) or UV (20 J/m2) and collected 4 h later. Anti-Abl immunoprecipitates were prepared, resolved on 8% SDS-PAGE, and subjected to fluorography followed by autoradiography ( top panel ). An aliquot of the immune complex was immune-analyzed with Flag antibody ( bottom panel ). (d) 293 cells were transfected with either vector alone or the indicated Abl constructs shown in panel e , using lipofectamine. At 36 h post-transfection, cells were exposed to 5 Gy of IR and anti-Flag immunoprecipitates were formed and immune-analyzed with anti-acetyl lysine ( top panel ) or anti-Flag antibody ( bottom panel ). (e) Schematic representation of the Abl constructs used in the experimentation described in panel d .

Article Snippet: Pan-specific acetyl-lysine (C4; sc-8663), monoclonal anti-p53 (D01; sc-126), anti-p300 (sc-585) and tubulin antibodies were obtained from Santa Cruz Biotech Company.

Techniques: Labeling, SDS Page, Autoradiography, Transfection, Plasmid Preparation, Construct

Journal: Molecular Cancer

Article Title: Tip60-mediated acetylation activates transcription independent apoptotic activity of Abl

doi: 10.1186/1476-4598-10-88

Figure Lengend Snippet: K921 of Abl is modified by acetylation following DNA strand breaks . (a) Structure of c-Abl with the consensus acetylation sequence (mouse and human) located in the DNA-binding domain (DBD) is shown. (b) Abl -/- 3T3 cells, reconstituted with Flag-tagged WT or K>R (921, 924) mutants of Abl, were either mock-treated or exposed to 5Gy of IR. Anti-Flag immunoprecipitates were prepared and immune-analyzed with acetyl-lysine ( top panel ) or Flag antibody ( bottom panel ). (c) Immunoblot, shown in panel A, was reprobed against K921 site-specific acetyl-Abl antibody ( top panel ). An aliquot of the lysate was also immune-analyzed with anti-Flag antibody ( bottom panel ). (d) Bleomycin induces K921 Abl acetylation. Abl -/- 3T3 cells reconstituted with vector or Flag-tagged Abl were exposed to bleomycin (10 μM; 1 h) and lysates, prepared 4 h later, were subjected to immunoblotting with K921 acetyl Abl ( top panel ) or anti-Flag ( bottom pane l) antibody.

Article Snippet: Pan-specific acetyl-lysine (C4; sc-8663), monoclonal anti-p53 (D01; sc-126), anti-p300 (sc-585) and tubulin antibodies were obtained from Santa Cruz Biotech Company.

Techniques: Modification, Sequencing, Binding Assay, Western Blot, Plasmid Preparation

Journal: Molecular Cancer

Article Title: Tip60-mediated acetylation activates transcription independent apoptotic activity of Abl

doi: 10.1186/1476-4598-10-88

Figure Lengend Snippet: K921 Abl acetylation is mediated by Tip60 acetyl-transferase . (a) Human colorectal cancer cells (HCT116+Ch3;H3) were transfected with siRNA specific for Luciferase, Tip60 or hMOF and then mock treated or exposed to IR (5 Gy) at 36 h post-transfection. Lysates were formed at 4 post-treatment, normalized for equal protein content, and immune-analyzed with K921 Ac Abl or Abl antibody. An aliquot of the lysate was also subjected to immunoblotting with anti-Tip60 or hMOF antibody. (b) 293T cells were transfected with plasmids carrying WT or HAT-defective Tip60mt cDNA, and at 36 h post-transfection, cells were mock-treated or exposed to 5 Gy of IR. Lysates were formed 4 h later and subjected to immunoblotting with K921 Ac Abl ( top panel ), anti-Flag ( middle panel ) and anti-Tip60 antibodies, respectively ( bottom panel ). (c) Structural domains of GST-Abl fusion products (Abl-ABC, Abl-XS and Abl-SE). (d) Tip60 acetylates c-Abl in vitro . GST-fusion proteins, shown in panel C, were expressed in bacteria purified, and incubated with Tip60 immunoprecipitates prepared from IR (5 Gy)-treated HeLa cells in the presence of 14 C-acetyl CoA in HAT buffer. The reaction products were resolved in 5-15% SDS-PAGE and the gel was dried and exposed to autoradiography ( top panel ). Coomassie blue staining of the GST-fusion products ( middle panel ). Anti-Tip60 immunoblotting ( bottom panel ).

Article Snippet: Pan-specific acetyl-lysine (C4; sc-8663), monoclonal anti-p53 (D01; sc-126), anti-p300 (sc-585) and tubulin antibodies were obtained from Santa Cruz Biotech Company.

Techniques: Transfection, Luciferase, Western Blot, In Vitro, Purification, Incubation, SDS Page, Autoradiography, Staining