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. 2023 Jun 30;12:e86936. doi: 10.7554/eLife.86936

Figure 1. A proteomic approach to identify tau aggregate interactors.

(A) Tau aggregates were partially purified from DS1 and DS10 HEK293 cells expressing tauRD-YFP. Detergent fractionation enabled generation of a sarkosyl-insoluble fraction containing tau aggregates. Proteins were resolved by SDS-PAGE and then extracted from individual lanes for analysis by LC-MS/MS. (B) Volcano plot showing proteins enriched in the sarkosyl-insoluble fraction as a fold enrichment from cells expressing tauRD-YFP aggregates (DS10, dark blue dots) over cells expressing tauRD-YFP that does not form aggregates (DS1, gray dots). The red line indicates a false discovery rate of 1.5%. Gene ontology (GO) term enrichment analyses of biological processes is also shown for select GO terms: orange dots, chaperone-mediated protein folding (chaperone folding); green dots, regulation of ubiquitination (Ub regulation); teal dots, ubiquitin-dependent protein catabolic process (Ub degradation). (C) Spectral indices for a selection of the proteins identified only in the DS10-insoluble fraction. Viable knockouts are shown as green bars. Non-viable knockouts are shown as black bars. Error bars represent the SEM of three extracted protein SDS-PAGE gel bands.

Figure 1.

Figure 1—figure supplement 1. Partial purification of tau aggregates.

Figure 1—figure supplement 1.

Sarkosyl-soluble and sarkosyl-insoluble fractions were run on gels and proteins were stained with SimplyBlue protein stain. DS9 and DS10 featured significant enrichment of insoluble tauRD-YFP (arrow on gel). Whole lanes for pellet fractions were analyzed by mass spectrometry (biological triplicates). The DS9 cell line, which contains tau aggregates of a distinct strain, is included as a tau-aggregate containing control. Source data for this figure are provided in Figure 1—figure supplement 1—source data 1.
Figure 1—figure supplement 1—source data 1. This source data file contains the original uncropped images for the SDS-PAGE gels shown in Figure 1—figure supplement 1.