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A widespread family of heat-resistant obscure (Hero) proteins protect against protein instability and aggregation

Fig 4

Hero proteins prevent pathogenic protein aggregation in cells.

(A) Representative microscopic images of GFP signals in the cells transfected with GFP-tagged TDP-43ΔNLS, HTTQ103, or GA50, together with the GST control or each Hero protein. Note that signals from aggregations are saturated. (B) Filter trap assay of aggregation-prone proteins expressed in cells. TDP-43ΔNLS, HTTQ103, or GA50 was expressed in a GFP-fusion form in HEK293T cells, together with each Hero protein or the GST control. The original or 5-fold diluted samples were loaded on a cellulose acetate membrane in the presence of 1% SDS and the trapped aggregates were probed with anti-GFP antibody. (C) Filter trap assay of aggregation-prone proteins expressed in cells. GFP-TDP-43ΔNLS was expressed in HEK293T cells, together with each Hero protein, its KR-less mutant, in which positively charged amino acids (Lys and Arg) were substituted with neutral Gly, or the GST negative control. Five-fold serial dilutions of the cell lysates were subjected to the filter trap assay and probed with the anti-GFP antibody. Experiments were simultaneously performed, and the presented data were spliced from the same membrane. (D) Filter trap assay of aggregation-prone proteins expressed in cells. GFP-TDP-43ΔNLS was expressed in HEK293T cells, together with the GST control, each Hero protein, or their shuffled amino acid sequences with 42– or 100–amino acid length. Five-fold serial dilutions of the cell lysates were subjected to the filter trap assay and probed with anti-GFP antibody. Experiments were simultaneously performed, and the presented data were spliced from the same membrane. aa, amino acid; GFP, green fluorescent protein; GST, glutathione S-transferase; Hero, heat-resistant obscure; KR, lysine and arginine; TDP-43ΔNLS, TDP-43 lacking the nuclear localization signal; WT, wild-type.

Fig 4

doi: https://doi.org/10.1371/journal.pbio.3000632.g004