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. 2022 Jun 17;298(8):102158. doi: 10.1016/j.jbc.2022.102158

Figure 2.

Figure 2

Cytosolic Hsp70 and Hsp40 activity buffer against protein misfolding and aggregation.A, lower slope values derived from flow cytometry analysis of HEK293T cells cotransfected with the biosensors, DNAJB1 and HSPA1A or control (a nonfluorescent derivative of GFP (Y66L Emerald (12)) for 48 h. Data points indicate biological replicates, and bars indicate means ± SD. Student’s t test results are shown (2-tailed; control versus overexpression); ∗∗∗p < 0.001, ∗∗p < 0.01, ∗p < 0.05, ns ≥ 0.05. B, aggregation analysis (A50%) using the same treatments and conditions as for A. C, shown are immunofluorescence micrographs of HEK293T cells transiently transfected with either V5-tagged Hsp40 or Hsp70 proteins (DNAJB1 and HSPA1A, respectively). The nucleus is stained with Hoechst 33342 and chaperone with Cy5-labeled anti-V5 antibody (or isotype control for specificity). Graphs indicate quantitation with paired Student’s t test results shown (2-tailed, paired); ∗∗∗p < 0.001, ∗∗p < 0.01. Data points represent immunofluorescence intensity in single cells (paired by mean). HEK293T, human embryonic kidney 293T cell line; ns, not significant.