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. Author manuscript; available in PMC: 2022 Dec 6.
Published in final edited form as: Neuromuscul Disord. 2019 Nov 19;30(1):38–46. doi: 10.1016/j.nmd.2019.11.005

Fig. 6.

Fig. 6.

Impaired function of mutant DNAJB6b. (A) Filter-trap assay showed significant loss of 120Q-HTT anti-aggregation activity for p.A50V and p.E54A, but not p.S57L, compared to wild-type DNAJB6b. The previously described p.F93L variant is included as reference and wild-type DNAJB6a (inactive towards cytoplasmic aggregation) as a negative control. Induction delay 5 h. (B) The p.H31Q mutation showed an additive impairing effect on DNAJB6b anti-aggregation activity towards 120Q-HTT when combined with p.A50V in filter-trap assay. Induction delay 4 h. Bars show mean ± SD of aggregation score (ratio of aggregated/soluble 120Q-HTT in induced versus uninduced cells) of 9 (A) or 12 (B) replicates. Representative FTA membranes show aggregated 120Q-HTT (in technical duplicates), whereas western blots show soluble 120Q-HTT and V5-DNAJB6 in uninduced (−) and induced (+) cells. For all FTA and WB images, see online supplementary file 2. (C) All investigated DNAJB6b variants (p.A50V, p.E54A, p.S57L, and p.F93L) increased nuclear TDP-43 aggregation after heat shock, whereas wild-type and p.H31Q DNAJB6b and GFP had no such effect. Bars show the percentage of cells with TDP-43 accumulations (mean ± SD) in 8 replicates. Representative cell images of DNAJB6b wild-type and p.A50V are shown under the graph; see online supplementary file 3 for the full series of representative images. *p<0.05, **p<0.01, ***p<0.001 compared to wild-type DNAJB6b (A) and (C) or for the indicated pairs (B).