Journal: European Journal of Immunology

Article Title: Fever‐Induced Heat Shock Protein‐70 Regulates Macrophage IL‐1β and IL‐10 Secretion During Mycobacterium tuberculosis Infection

doi: 10.1002/eji.202551963

Figure Lengend Snippet: HSP70 antagonism decreases Mtb ‐stimulated IL‐1β and IL‐10 secretion and gene expression from MDM at 37°C and 40°C. (A) IL‐1β and (B) IL‐10 secretion from Mtb ‐stimulated MDM, either pretreated with vehicle (open bars) or with 25 µM HSP70 antagonist Ver155008 (black bars, n = 6 donors) at 37°C or 40°C for 24 h. Fold changes in secretion are relative to vehicle control pretreated Mtb ‐infected MDM at 37°C. (C) il1b and (D) il10 fold change in gene expression from Mtb ‐stimulated MDM, either pretreated with vehicle (open bars) or with 25 µM Ver155008 (black bars, n = 3 donors) at 37°C or 40°C for 24 h. Fold change in gene expression is relative to the vehicle control pretreated control MDM at 37°C. Mean ± SEM are shown. Two‐way ANOVA statistical testing was performed (* p < 0.05, *** p < 0.001, **** p < 0.0001).

Article Snippet: In later experiments, MDM were pretreated with 25 μM Ver155008, a HSP70 antagonist (Santa Cruz Biotechnology, USA), or 500 ng/mL recombinant human HSP70 (rhHSP70, Bio‐Techne, UK) 1 h before infection.

Techniques: Gene Expression, Control, Infection

Journal: European Journal of Immunology

Article Title: Fever‐Induced Heat Shock Protein‐70 Regulates Macrophage IL‐1β and IL‐10 Secretion During Mycobacterium tuberculosis Infection

doi: 10.1002/eji.202551963

Figure Lengend Snippet: Recombinant HSP70 increases IL‐1β secretion at 37°C, and inhibition of HSP70 activity increases Mtb ‐induced HSP70 secretion at 37°C, and to a greater extent at 40°C. (A) IL‐1β and (B) IL‐10 secretion from Mtb ‐stimulated MDM, either pretreated with vehicle (open bars) or with 500 ng/mL recombinant human HSP70 (rhHSP70, black bars, n = 4–6 donors) at 37°C or 40°C for 24 h. Fold changes in secretion are relative to vehicle control pretreated Mtb ‐infected MDM at 37°C. (C) HSP70 secretion from Mtb ‐stimulated MDM, either pretreated with vehicle (open bars) or with 25 µM Ver155008 (black bars, n = 4 donors) at 37°C or 40°C for 24, 48, or 72 h. Mean ± SEM are shown. Two‐way ANOVA statistical testing was performed (* p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001).

Article Snippet: In later experiments, MDM were pretreated with 25 μM Ver155008, a HSP70 antagonist (Santa Cruz Biotechnology, USA), or 500 ng/mL recombinant human HSP70 (rhHSP70, Bio‐Techne, UK) 1 h before infection.

Techniques: Recombinant, Inhibition, Activity Assay, Control, Infection

Journal: Science advances

Article Title: USP8 and Hsp70 regulate endoreplication by synergistically promoting Fzr deubiquitination and stabilization.

doi: 10.1126/sciadv.adq9111

Figure Lengend Snippet: Fig. 5. Hsp70 promotes the interaction between Fzr and USP8 by mediating Fzr folding. (A and B) Exogenously overexpressed Hsp70 promoted the interaction be- tween exogenously overexpressed USP8 and Fzr in BmE cells and S2 cells. (C and D) GST pull-down assay showed that supplementation with recombinant Hsp70 in- creased the USP8-Fzr interaction in vitro. (E to H) The interaction between endogenous USP8 and Fzr in the cultured glands and cells was attenuated following a treatment with the Hsp70 interacting activity inhibitor PES for 2 hours. (I and J) The treatment with the Hsp70 inhibitors PES or VER155008 decreased the luciferase enzyme activity of luciferase-fused Fzr (Fzr-Luc) in cultured cells, suggesting that Hsp70 promoted proper folding of Fzr. (K and L) Overexpression of intact Hsp70 increased the luciferase enzyme activity of Fzr-Luc in cultured cells, but truncated Hsp70 with the deletion of SBD domain or NBD domain lost this promotion. The data are presented as the mean ± SE (error bars) of three independent biological replicates. For the significance test: *P < 0.05 and **P < 0.01 versus the control.

Article Snippet: Several inhibitors, including PES (Selleck) that inhibits the interaction between Hsp70 and its partners, VER155008 (Selleck) that inhibits the enzymatic activity of Hsp70, and IN- 2 (Selleck) that blocks the enzymatic activity of USP8, were used in the present study.

Techniques: Pull Down Assay, Recombinant, In Vitro, Cell Culture, Activity Assay, Luciferase, Over Expression, Control