Fig. 3: Eh-induced active caspase-6 mediated ATG16L1 protein degradation.

a THP-1 macrophages were pre-treated with pan-caspase inhibitor Z-VAD-fmk (50, 100 μM) for 1 h before incubation with Eh for 10 and 30 min. b THP-1 macrophages were pre-treated with caspase-6 inhibitor Z-VEID-fmk (20, 50, 100 μM) for 1 h before stimulation with Eh for 10 min. c THP-1 macrophages were stimulated with Eh for different time points to detect active caspase-6 and its known substrate lamin A/C. d Caspase-6 activity was measured in presence of Eh and Eh along with caspase-6 inhibitor Z-VEID-fmk (50 μM). e Bone marrow derived macrophages (BMDMs) were pre-treated with pan-caspase inhibitor Z-VAD-fmk and caspase-6 inhibitor Z-VEID-fmk for 1 h with 50 μM concentration prior to stimulation with Eh for 10 min. Restoration of ATG16L1 protein and ATG12-ATG5 conjugate were assessed by western blot along with lamin A/C restoration. f THP-1 macrophages were transfected with 50 nM caspase-6 siRNA, or scramble siRNA by nuclear factor technique. After 48 h, transfected cells were stimulated with Eh for 5 min and western blot was performed to detect indicated proteins. Equal amount of lysed cell lysates was loaded onto SDS-PAGE gel and immunoblotted with indicated antibodies. Cells without any treatment is symbolized as (−ve). Eh-macrophage 1:20 ratio used. Results are representative of three independent experiments (n = 3). *P < 0.05, **P < 0.01.