Figure 3

Macroautophagy markers in SH-SY5Y cells overexpressing wild-type (WT) or A53T mutant α-synuclein. A. α-Synuclein-immunopositive dots and homogenous staining of α-synuclein in SH-SY5Y cells over-expressing WT α-synuclein plasmid transfection. B. Larger α-synuclein-immunopositive aggregates in SH-SY5Y cells with A53T α-synuclein plasmid transfection. C. Truncated small α-synuclein fractions were detected in both WT (lane 1) and A53T mutant (lane 2) α-synuclein transfected cells (20~49kDa) (non-transfected cells in lane 3). The ATG5-ATG12 complex (55kDa) was significantly increased in cells overexpressing α- synuclein protein (GFP-fused major band at ~49kDa, lanes 1 and 2). Repeat experiments showed there were no observable differences between cells transfected with WT (lane 1) or mutant (lane 2) α-synuclein in the levels of ATG5 monomer (band at ~33kDa), the active form of ATG8/LC3-II (~16kDa), or inATG6/Beclin1 (~60kDa), under the condition of equal protein loading (internal control of β-tublin-III at ~50kDa).