Figure 2
ATG7 isoform 2 is not able to bind LC3. (A) Amino acid sequence alignment of ATG7 from S. cerevisae, D. melanogaster, C. elegans, H. sapiens and M. musculus. The exon lacking in human ATG7(2) is underlined (I626-K652). (B) Representation of the yeast Atg7 dimer showing the protomer surfaces colored in faint green or gray. Other ATG proteins, in ribbon representation, are shown to indicate their location in the respective protein complexes. The region lacking in human isoform 2 is shown in dark blue. (C) HEK293T cells were transfected with the two isoforms of Myc-tagged ATG7 or empty vector (EV) and protein lysates were prepared using differential detergent extraction prior to Western blot analysis. Membranes were probed with ATG7 and Actin antibodies. Quantification of band intensities was performed using ImageJ software. Error bars represent SEM of four independent experiments. Two-way Anova with Sidak’s multiple comparisons test was performed revealing no significant statistical difference. (D) Co-immunoprecipitation experiments in HEK293T overexpressing Myc-tagged empty vector (EV), ATG7(1) or ATG7(2), together with 3xFLAG-LC3. Constructs were detected using ATG7 and FLAG antibodies, respectively.
