Hsp60

T cells were stained with primary antibodies against HSP90α (2G5.G3, Abcam, USA), HSP90β (EPR16621, Abcam, USA), HSP70 (C92F3A-5, Enzo Life Sciences, USA), HSC70 (1B5, Stressgen, USA), HSP40/DNAJB1 (3B9.E6, Abcam, USA), GRP78 (StressMarq Biosciences, USA), and HSP60 (StressMarq Biosciences, USA). Secondary antibodies were goat anti-rabbit IgG H&L (PE) preadsorbed (Abcam), anti-mouse IgG1 (PE) (Miltenyi Biotec, Germany), and anti-rat F(ab’)2 (PE) (Sony Biotechnology, USA). T cells were also stained with anti-human CD279/PD-1 (REA1165, Miltenyi Biotec, Germany) (PE), CD152/CTLA-4 (REA1003, Miltenyi Biotec, Germany) (PE), primary antibody against STAT3 (4G4B45, Sony Biotechnology, USA), and secondary antibody anti-mouse IgG1 (PE) (Miltenyi Biotec, Germany). The FcR blocking reagent (Miltenyi Biotec, Germany) and isotype controls were used to exclude non-specific binding. For intracellular staining, cells were fixed and permeabilized using Cytofix/Cytoperm. Cells were analyzed by flow cytometry.

The following reagents were purchased from the indicated manufacturers: SMI-16a and mouse monoclonal antibody against Noxa from Calbiochem (Darmstadt, Germany); rabbit polyclonal anti-antibodies against human IRF4, bortezomib and mouse monoclonal antibody against C/EBP-homologous protein (CHOP), phosphorylated eukaryotic Initiation Factor 2α (p-eIF2α) and activating transcription factor 4 (ATF4) from from Cell Signaling Technology (Beverly, MA); Pim-2 and c-Myc from Abcam (Cambridge, UK); HSP70 from Biolegend (San Diego, CA); HSP60 from StressMarq Biosciences Inc.(Victoria, Canada); Mcl-1 from Medical and Biotechnological Laboratories (Nagoya, Japan); tunicamycin, puromycin and β-actin from Sigma (Saint Louis, MO); anti-puromycin antibody from Kerafast, Inc (Boston, MA); and anti-ubiquitin antibody was obtained from Millipore (Darmstadt, Germany).