Fig. 8.

MFM-causing BAG-domain mutations also cause HSP70-dependent aggregation. a Immunofluorescence pictures of HeLa cells expressing the indicated BAG3-disease causing mutants, with or without the additional R480A mutation that abrogates the ability of the (mutant) BAG3s to interact with Hsp70. Scale bar = 5 μm. b Analyses of the number of cells expressing the indicated BAG3-disease causing mutants (with or without the additional R480A mutation that abrogates their ability BAG3s to interact with Hsp70) that contain over 20 punctae. The increase in cells with punctae in the single mutant backgrounds and their abrogation in the double mutant backgrounds are significant with a P value of <0.00005 (t test). c Pedigree structures and known genotype of the proband (*) and parents in Family 1 and Family 2 with a P470S mutation in the BAG-domain. d Histochemical and immunohistochemical analysis of muscle biopsies from two novel MFM patients carrying a P470S mutation in the BAG domain of BAG3: Patient 1 (a–c) and Patient 2 (d, e) (see Fig. S6B, C for further patient info); a hematoxylin and eosin (H&E) staining demonstrates region of grouped atrophy with small fibers harboring vacuoles; b modified gomori trichrome (GT) staining demonstrating both cytoplasmic inclusion and rimmed vacuoles; c immunohistochemistry with an antibody to TDP-43 demonstrates protein accumulation; d H&E staining shows large centrally located eosinophilic inclusion; e GT staining shows dark centrally located inclusions and a fiber with a centrally located vacuole. Arrows denote protein inclusions or vacuoles. Scale bar = 50 μm. e Immunoprecipitation of GFP-tagged BAG3 variants from cells expressing both GFP-BAG3 variants and FLAG-tagged HSP70. Western blots using the indicated antibodies is shown. Source data are provided as a Source data file