Journal: Turkish Journal of Hematology

Article Title: Oncolytic Myxoma virus Increases Autophagy in Multiple Myeloma

doi: 10.4274/tjh.galenos.2024.2023.0403

Figure Lengend Snippet: ELISA results of p62 protein concentrations in MYXV-infected U-266 and MOPC-315 cells according to hours of incubation: A) 24 h of incubation period, B) 48 h of incubation, C) 72 h of incubation. ELISA: Enzyme-linked immunosorbent assay; MYXV: Myxoma virus .

Article Snippet: In MM cell lines, the levels of apoptotic protein Bcl-2 (catalog no: E1832Hu, BT LAB, China) and the autophagic proteins Beclin-1 (catalog no: E2011Hu, BT LAB), human sequestosome-1 (p62) (catalog no: E6779 Hu, BT LAB), human LC3B (catalog no: MBS1603826, MyBioSource, USA), and human ATG-5 (catalog no: EH1729, FineTest, USA) were measured by ELISA according to the manufacturer’s instructions.

Techniques: Enzyme-linked Immunosorbent Assay, Infection, Incubation, Virus

Journal: Molecular and Cellular Biology

Article Title: Targeting SQSTM1/p62 Induces Cargo Loading Failure and Converts Autophagy to Apoptosis via NBK/Bik

doi: 10.1128/mcb.01383-13

Figure Lengend Snippet: FIG. 3. Cdk inhibition down-regulates SQSTM1/p62 but fails to affect LC3 processing during

Article Snippet: The SDS-insoluble aggregates trapped on the filter were probed with anti-ubiquitin (Cell Signaling, Beverly, MA), anti-p62 (Santa Cruz Biotech, Santa Cruz, CA), or anti-Bik (ProSci, Poway, CA), respectively.

Techniques: Inhibition

Journal: Molecular and Cellular Biology

Article Title: Targeting SQSTM1/p62 Induces Cargo Loading Failure and Converts Autophagy to Apoptosis via NBK/Bik

doi: 10.1128/mcb.01383-13

Figure Lengend Snippet: FIG. 4. SQSTM1/p62 down-regulation results in cargo loading failure and inefficient autophagy.

Article Snippet: The SDS-insoluble aggregates trapped on the filter were probed with anti-ubiquitin (Cell Signaling, Beverly, MA), anti-p62 (Santa Cruz Biotech, Santa Cruz, CA), or anti-Bik (ProSci, Poway, CA), respectively.

Techniques:

Journal: Molecular and Cellular Biology

Article Title: Targeting SQSTM1/p62 Induces Cargo Loading Failure and Converts Autophagy to Apoptosis via NBK/Bik

doi: 10.1128/mcb.01383-13

Figure Lengend Snippet: FIG. 5. Expression of SQSTM1/p62 diminishes the increased lethality of BH3-mimetics in p62-

Article Snippet: The SDS-insoluble aggregates trapped on the filter were probed with anti-ubiquitin (Cell Signaling, Beverly, MA), anti-p62 (Santa Cruz Biotech, Santa Cruz, CA), or anti-Bik (ProSci, Poway, CA), respectively.

Techniques: Expressing

Journal: Molecular and Cellular Biology

Article Title: Targeting SQSTM1/p62 Induces Cargo Loading Failure and Converts Autophagy to Apoptosis via NBK/Bik

doi: 10.1128/mcb.01383-13

Figure Lengend Snippet: FIG. 9. A mechanistic model of SQSTM1/p62 and NBK/Bik acting as novel molecular switches

Article Snippet: The SDS-insoluble aggregates trapped on the filter were probed with anti-ubiquitin (Cell Signaling, Beverly, MA), anti-p62 (Santa Cruz Biotech, Santa Cruz, CA), or anti-Bik (ProSci, Poway, CA), respectively.

Techniques:

Journal: Cancer & Metabolism

Article Title: Inhibition of 6-phosphofructo-2-kinase (PFKFB3) induces autophagy as a survival mechanism

doi: 10.1186/2049-3002-2-2

Figure Lengend Snippet: Transfection of HCT-116 cells with PFKFB3 siRNA stimulates autophagy. LC3-II and p62 protein levels were determined using Western blotting 48 hours after transfection with either control (ctrl) or a siRNA directed against PFKFB3 (PFKFB3) (A) . Treatment with 1 nM bafilomycin A1 (Baf A1) was used to determine if LC3-II levels were a result of increased autophagic flux or impaired degradation (A) . Quantitative densitometry was performed to assess relative protein levels (B, C) . LC3-II and p62 levels are expressed as the mean fold change ± SD from three experiments relative to LC3-I or β-actin and control. After 48 hours of transfection with either control (ctrl) or PFKFB3-specific siRNA, cells also were stained with acridine orange, observed by fluorescent microscopy and collected by flow cytometry to measure the relative content of acidic compartments (D) . Examination of the cells by electron microscopy demonstrated that PFKFB3 siRNA transfection resulted in cells containing intracellular structures consistent with autophagosomes (E; arrow) . Data are presented as the mean ± SD from three experiments (* P <0.05).

Article Snippet: Antibodies against LC3, p62, p-p70S6K, p70S6K, pS6, S6, phospho-AMPK, AMPK, phospho-ULK1, and ULK1 (Cell Signaling, Danvers, MA, USA), PFKFB3 (Proteintech, Chicago, IL, USA), and β-actin (Sigma) were diluted 1:1,000 and incubated overnight at 4°C, with the exception of p62 and β-actin Ab, which were incubated at room temperature for 1 hour.

Techniques: Transfection, Western Blot, Staining, Microscopy, Flow Cytometry, Electron Microscopy

Journal: Cancer & Metabolism

Article Title: Inhibition of 6-phosphofructo-2-kinase (PFKFB3) induces autophagy as a survival mechanism

doi: 10.1186/2049-3002-2-2

Figure Lengend Snippet: PFKFB3 inhibition with 3PO stimulates autophagy. HCT-116 cells were treated with either vehicle, or 7.5, 10, or 15 μM 3PO for 24 hours and LC3-II and p62 expression was measured by Western blot (A) and densitometry (B, C) . Addition of bafilomycin A1 (Baf A1) was used to determine if the changes in LC3-II were the result of increased synthesis or impaired degradation. LC3-II quantitation is relative to control + bafilomycin due to the absence of a visible band in the control sample. HCT-116 cells were also stained with 1 μg/mL acridine orange for 15 minutes, viewed using a fluorescent microscope, harvested for flow cytometry and gating was used to quantitate the number of cells with a high AO fluorescence and expressed relative to vehicle (D, E) . Using electron microscopy, autophagic structures were seen in cells exposed to 3PO (F; arrow) .

Article Snippet: Antibodies against LC3, p62, p-p70S6K, p70S6K, pS6, S6, phospho-AMPK, AMPK, phospho-ULK1, and ULK1 (Cell Signaling, Danvers, MA, USA), PFKFB3 (Proteintech, Chicago, IL, USA), and β-actin (Sigma) were diluted 1:1,000 and incubated overnight at 4°C, with the exception of p62 and β-actin Ab, which were incubated at room temperature for 1 hour.

Techniques: Inhibition, Expressing, Western Blot, Quantitation Assay, Staining, Microscopy, Flow Cytometry, Fluorescence, Electron Microscopy