1. AS aggregates interact with SERCA in SH‐SY5Y cells
AS aggregates interact with SERCA in SH‐SY5Y cells Non‐mitotic SH‐SY5Y cells were grown as depicted in Fig 1B for 5 days after which they were fixed and subjected to proximity ligation assay using Syn211 to target AS and anti‐SERCA2. Representative images are from one experiment of three. AS‐expressing SH‐SY5Y cells were analysed by proximity ligation assay to demonstrate an intracellular interaction between SERCA and AS using SERCA2 and syn211 as antibody pair. Positive PLA signals are shown as red staining merged with the phase contrast image and DAPI‐stained nuclei (blue). Scale bar is 20 μm.Proximity ligation assays as in (A), but with cells treated with AS aggregation inhibitor, ASI‐1d, from day 3. Scale bar is 20 μm.Proximity ligation assay as in (B), but in cells without AS expression. Scale bar is 20 μm.The signal from proximity ligation assay was quantification as total red fluorescence signal, divided by the DAPI signal to normalise cellular content. In each experiment, 10 microscopic images containing > 100 cells were quantified. Bars represent mean ± SD signal normalised to AS‐expressing cells from three experiments (one‐way ANOVA multiple comparisons with Sidak post hoc test, *P = 0.0001).
Cristine Betzer et al. EMBO Rep. 2018;embr
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