HSP110 stabilizes MyD88. Immunoblot analysis of HSP110-GFP, total HSP110, (A) MyD88 L265P, or (B) MyD88 WT 48 hours after transfection with plasmids coding for MyD88 L265P (0.7 µg) together with control GFP, HSP110 DE9GFP, or increasing concentrations of HSP110-GFP in HEK293 cells (n = 3). HSC70 served as a loading control. (C) Immunoblot analysis of HSP110-GFP and MyD88 L265P at 48 hours after transfection with plasmids coding for MyD88 L265P (0.5 μg) together with control GFP, increasing concentrations of the cytoplasmic-only mutant of HSP110 (HSP110 deficient in the nuclear localization signal [HSP110-NLS]) in HEK293 cells (n = 3). (D) Immunoblot analysis of HSP110-GFP, total HSP110, MyD88 L265P, or MyD88 WT in HEK293 cells treated with cycloheximide (CHX; 100 μg/mL) during the indicated times, 24 hours after transfection with control-GFP or HSP110-GFP plasmids. HSC70 served as a loading control. Lower panel shows the MyD88 immunoblot intensity from the above experiment. (E) Immunoblot analysis of HSP110 and MyD88 in HBL1, OCI-Ly3, and U2932 60 hours after transfection by a universal control siRNA or with an siRNA that targets HSP110 (n = 3). conc, concentration. (F) Immunoblot analysis of total HSP110 and MyD88 in HBL1 transfected as in (E) and treated or not with MG132 (10 μM) during the last 3 or 5 hours of the culture (n = 3).