Fig. 1: DnaJB8 architecture defined by domain-domain interactions.

a Domain maps for DnaJB8 used in the in vitro experiments and the DnaJB–Clover and Clover constructs used in the mammalian cell experiments. DnaJB8 is colored according to domain annotation: JD (red), G/F rich (blue), S/T rich (cyan), and CTD (green). Clover is colored pale green. b Representative images of triplicate populations of 300,000 cells expressing DnaJB8–mClover3 (left) and mClover3 (right). Clover and DAPI fluorescence signals are shown in green and blue, respectively. Scale bar, 5 μm, is shown in white. c Quantification of DnaJB8–Clover and Clover puncta in high (3×) and low (1×) protein level expressing cell lines. In each analysis at least 2000 cells were counted by the CellProfiler software. Puncta were manually counted by two independent observers, with data reported as averages with standard deviation. d XL–MS contact map of DnaJB8–Clover cross-links identified using DMTMM and ADH. The axes are colored in red and green for JD and CTD, respectively. Cross-link pairs between JD–CTD, JD–JD, and CTD–CTD are shown in dashed boxes colored gray, red, and green, respectively. Contacts to helix 5 are denoted with H5. e Histogram of overall R_h of DnaJB8 in 1× PBS 150 mM NaCl from DLS at times 0 h (black), 10 h (blue), and 20 h (gold), with arrows indicating R_h peaks for each time point. Over time, there was a depletion in particle sizes <10 nm and an increase in particles ~100–1000 nm. f XL–MS contact map of DnaJB8 cross-links identified using DMTMM and ADH (black) and DSS (gray). The axes are colored in red and green for JD and CTD, respectively. Cross-link pairs between JD–CTD, JD–JD, and CTD–CTD are shown in a dashed box colored gray, red and green, respectively. Contacts to helix 5 are denoted with H5. g Histogram of overall R_h of DnaJB8 in 1× PBS 285 mM NaCl at times 0 h (blue), 10 h (black), and 20 h (red), with arrows indicating R_h peaks for each time point. Over time, there is no change in the species of particle sizes <10 nm and no appearance of particles ~100–1000 nm. h Contact map of DnaJB8 cross-links identified using ADH/DMTMM in the presence of 285 mM NaCl. The axes are colored in red and green for JD and CTD, respectively. JD–JD and CTD–CTD cross-links are shown in dashed boxes colored in red and green, respectively.