Fig. 5

DNAJB6 is associated with tau and HSP70. A Co-immunoprecipitation assay indicated that DNAJB6 is associated with tau and HSPA8. SH-SY5Y cells expressing tau P301L were immunoprecipitated using an anti-DNAJB6 antibody to pull down endogenous DNAJB6. An anti-GST antibody served as a negative control. Precipitated and co-precipitated proteins were analyzed by immunoblotting using indicated antibodies. The images are representatives of three biological replicates. B SH-SY5Y cells co-expressing tau P301L and an empty vector, V5-DNAJB6a, or V5-DNAJB6b for 48 h were immunoprecipitated using an anti-V5 antibody to pull down V5-DNAJB6a or V5-DNAJB6b. Co-immunoprecipitated proteins were detected as described in A. C Quantification of the ratio of co-immunoprecipitated tau to immunoprecipitated DNAJB6a or DNAJB6b in B. The values were given as mean ± S.D. (n = 3, ***P < 0.001, unpaired two-tailed Student’s t-test). D V5-DNAJB6b and EGFP-tau plasmids were co-transfected into SH-SY5Y cells. 24 h post-transfection, cells were seeded on slides for an additional 24 h. Cells seeded on slides were then hybridized with tau and V5 primary antibodies to detect tau and DNAJB6b, respectively. When tau and DNAJB6 interact, the PLA probes on secondary antibodies were ligated and amplified. The red PLA signals showing protein interaction were detected by fluorescent microscopy. Scale bar: 10 μm. E Quantification of red PLA signal of D. The percentage of PLA-positive SH-SY5Y cells to the tau P301L transfected cells (EGFP-positive) was quantified. The values were given as mean ± S.D. (n = 3, **P < 0.01, unpaired two-tailed Welch’s t-test). The individual data values of the replicates in C and E are listed in Additional file 2