Fig. 2: PS1 overexpression decreases the levels of TDP43 and γ-secretase inhibition promotes TDP43-43KD accumulation.

A PS1 Cleavage of TDP43. HeLa cells were seeded in 35-mm dishes for 24 h. Then, one group was transfected with LacZ as a control, the second with 4 μg of PS1 plasmid, and the remaining three groups were pre-treated for 1 h with z-VAD, L685,458, and with their combination before transfection. Endogenous TDP43 levels were detected by western Blot analysis (Panel 1). The membranes in the top panels were reprobed with GAPDH antibodies to indicate the relative sample loading (bottom panels). C Effect of PS1 mutants on TDP43. HeLa cells were transfected with LacZ, PS1, PS1 D257A, and PS1 D385A for 24 h. Then, western blotting was performed to assess the levels of TDP43, and PS1-cas to verify that PS1 had been transfected into cells (panel 2). E Immunofluorescence detection of TDP43 levels after PS1 overexpression and inhibitor treatment. HeLa cells were transfected with LacZ as a blank control (Lane 1), PS1 alone (Lane 2), and pre-treated with z-VAD and L685,458 for 1 h before PS1 transfection (Lane 3). After 24 h, TDP43 levels were detected by immunofluorescence analysis. The TDP43 fluorescence intensity was decreased in the PS1 group, while, it increased in the inhibitor-treated group. DAPI was used to stain the nuclei, and TDP43 was labeled with FITC to emit green fluorescence. The merge image shows the integration of both channels. B–F Grayscale values of endogenous TDP43. The grayscale values in the graphs were calculated using Image J software analysis, and data are the mean ± SD of at least three independent experiments (*p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001, ns: not significant). The use of # indicates a comparison with the PS1+z-VAD + L685,458 group (# < 0.05, ## < 0.01, ### < 0.001, #### < 0.0001).