proteostat® assay enz-51023-kp050  (Enzo Biochem)

Journal: eLife

Article Title: Targeting DNA topoisomerases or checkpoint kinases results in an overload of chaperone systems, triggering aggregation of a metastable subproteome

doi: 10.7554/eLife.70726

Figure Lengend Snippet:

Article Snippet: Chemical compound, drug , ProteoStat , Enzo Life Sciences , ENZ-51023-KP050 , .

Techniques: Cell Culture, Luciferase, Recombinant, Plasmid Preparation, CRISPR, Sequencing, Silver Staining, Isolation, Software

Journal: eLife

Article Title: Targeting DNA topoisomerases or checkpoint kinases results in an overload of chaperone systems, triggering aggregation of a metastable subproteome

doi: 10.7554/eLife.70726

Figure Lengend Snippet: See also and . ( A ) Western blot of whole-cell lysate (WCL) and aggregated proteins isolated from HEK293T cells treated transiently with DMSO or camptothecin (CPT), followed by treatment with the VER-155008 HSP70 inhibitor (10 μM), probed with the indicated antibodies. n = 3. ( B ) Aggregated (silver stain) and WCL (Coomassie) fractions of U2OS wild-type and ATM KO cells, with or without overexpression of HSPB5. ( C ) Quantification of ( B ). ( D ) Representative immunofluorescence pictures of U2OS wild-type and ATM KO cells stably overexpressing HSPB5 or not, stained with ProteoStat (red) and Hoechst (blue). ( E ) Quantification of aggresome signatures in ( D ). ( F ). Representative immunofluorescence pictures of U2OS wild-type and ATM KO cells stably overexpressing HSPB5 or not, stained with anti-FUS (green) and Hoechst (blue). ( G ) Quantification of extranuclear FUS inclusions in ( F ). ( H ) Aggregated (silver stain) and WCL (Coomassie) fractions of HEK293 cells stably overexpressing HSPB5 or not, treated transiently with DMSO or CPT. Three technical repeats are shown here. ( I ) Quantification of three independent biological repeats of ( H ). In ( C ), ( E ), ( G ), and ( I ), squares represent independent biological repeats, bars represent mean ± SEM. p-Values are obtained by two-tailed Student’s t -tests followed by a Bonferroni correction for multiple comparisons. Figure 6—source data 1. Data from . Raw Western blot images with each antibody annotated and cropped parts indicated in red. Figure 6—source data 2. Data from . Raw Coomassie- and silver-stained SDS-PAGE gels and Western blot images with cropped parts indicated in red. Figure 6—source data 3. Data from . Raw Coomassie- and silver-stained SDS-PAGE gels and Western blot images with cropped parts indicated in red.

Article Snippet: ProteoStat staining (ENZO, ENZ-51023-KP050) was performed according to the manufacturer’s instructions.

Techniques: Western Blot, Isolation, Silver Staining, Over Expression, Immunofluorescence, Stable Transfection, Staining, Two Tailed Test, SDS Page

Journal: eLife

Article Title: Targeting DNA topoisomerases or checkpoint kinases results in an overload of chaperone systems, triggering aggregation of a metastable subproteome

doi: 10.7554/eLife.70726

Figure Lengend Snippet:

Article Snippet: ProteoStat staining (ENZO, ENZ-51023-KP050) was performed according to the manufacturer’s instructions.

Techniques: Cell Culture, Luciferase, Recombinant, Plasmid Preparation, CRISPR, Sequencing, Silver Staining, Isolation, Software

Journal: bioRxiv

Article Title: Targeting DNA topoisomerases or checkpoint kinases results in an overload of chaperone systems, triggering aggregation of a metastable subproteome

doi: 10.1101/2021.06.04.447142

Figure Lengend Snippet: See also . (A) Aggregated (silver stain) and whole cell lysate (WCL; Coomassie) fractions of HEK293T cells treated transiently with CPT, followed by treatment with the VER-155008 HSP70 inhibitor. n=3. (B) Western blot of WCL and aggregated proteins isolated from HEK293T cells treated transiently with CPT, followed by treatment with the VER-155008 HSP70 inhibitor (10 μM), probed with the indicated antibodies. n=3. (C) Aggregated (silver stain) and whole cell lysate (WCL; Coomassie) fractions of U2OS wild-type and ATM KO cells, with or without overexpression of HSPB5. (D) Quantification of C. (E) Representative immunofluorescence pictures of U2OS wild-type and ATM KO cells stably overexpressing HSPB5 or not, stained with Proteostat® (red) and Hoechst (blue). (F) Quantification of aggresome signatures in E. (G). Representative immunofluorescence pictures of U2OS wild-type and ATM KO cells stably overexpressing HSPB5 or not, stained with anti-FUS (green) and Hoechst (blue). (H) Quantification of extranuclear FUS inclusions in G. In D, F and H, squares represent independent experiments, bars represent mean ± SEM. P-values are obtained by two-tailed Student’s t-tests followed by a Bonferroni correction for multiple comparisons.

Article Snippet: ProteoStat® staining (ENZO, ENZ-51023-KP050) was performed according to the manufacturer’s instructions.

Techniques: Silver Staining, Western Blot, Isolation, Over Expression, Immunofluorescence, Stable Transfection, Staining, Two Tailed Test

Journal: G3: Genes|Genomes|Genetics

Article Title: Loss of N-Glycanase 1 Alters Transcriptional and Translational Regulation in K562 Cell Lines

doi: 10.1534/g3.119.401031

Figure Lengend Snippet: Correction of NGLY1-deficient phenotypes in K562 cell lines by exogenous expression of a DYK-tagged NGLY1 protein. (A) Western blot analysis of NGLY1-deficient K562 cell line's expression of plasmid-based N- or C-terminally tagged NGLY1. Antibodies probed are labeled along the left side of the figure along with corresponding kDa mol. weight markers. The same protein lysates were loaded on two gels. (B) FACS analysis of NGLY1-deficient K562 cell line's expression of plasmid-based N- or C-terminally tagged NGLY1. Results were quantified, averaged, and graphed. (C) Immunofluorescent staining of NGLY1-deficient K562 cell lines with DAPI (blue) and Proteostat (red). (D) Quantification of Proteostat staining of NGLY1-deficient K562 cells by flow cytometry analysis. Rescued lines include both C- and N-terminally tagged NGLY1.

Article Snippet: The Proteostat Dye (Enzo ENZ-51023-KP050 ) was used according to manufacturer’s protocol.

Techniques: Expressing, Western Blot, Plasmid Preparation, Labeling, Staining, Flow Cytometry