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. 2014 May 12;111(21):7741–7746. doi: 10.1073/pnas.1407001111

Fig. 2.

Fig. 2.

ATG16L1 T300A is more susceptible to caspase 3- and caspase 7-mediated cleavage. (A) Immunoblot analysis of LC3-I/II and p62 in MEFs untreated or stimulated for 4 h with 10 µg/mL E64d plus pepstatin or 100 nM Torin 1 and 10 μg/mL E64d plus pepstatin. Blots were probed for LC3-I/II and p62. Actin served as a loading control. (B) Schematic of ATG16L1 domains with caspase cleavage consensus sequence and location of the T300A polymorphism. CC, coiled-coil domain. WD, WD40 domain. (C–E) ATG16L1 WT (isoform 1) or ATG16L1 T300A, with or without an additional D299E mutation, was in vitro-translated with [35S]methionine and incubated with recombinant caspases for 1 h at 37 °C. ATG16L1 cleavage was observed by autoradiography. (F and G) Western blots showing ATG16L1 cleavage from HeLa cells transfected with indicated constructs and treated for 4 h with 1 μM staurosporine or 20 μM zVAD. FL, full-length.