Auto-regulatory J-domain interactions control Hsp70 recruitment in the oligomeric DNAJB8 co-chaperone

Molecular chaperones closely regulate cellular protein homeostasis, with the Hsp40/Hsp70 families managing to be both versatile and specific. Traditionally seen as “co-chaperones”, Hsp40s are now recognized as crucially important, but their structure and function are still poorly understood. The oligomeric members have especially proven resistive to in-depth characterization. Here we present a hybrid structure-function study of a homo-oligomeric Hsp40, DnaJB8, which is known for its potent suppression of aggregation by polyglutamine and other amyloidogenic proteins. Using a combination of biochemical, NMR, crosslinking-mass spectrometry, and in silico approaches, we examine the architecture and dynamics of DnaJB8 oligomers. We identify an electrostatically driven intra-oligomer interaction between the DnaJ-characteristic J-Domain (JD) and DnaJB89s C-terminal domain (CTD). This inter-domain contact is autoinhibitory of Hsp70 interaction by occluding the JD surface essential for Hsp70 binding. Disruption of the JD-CTD interaction liberates and mobilizes the JD to facilitate Hsp70 binding. These results identify the JD-CTD interaction as an evolutionarily conserved built-in regulatory element that controls recruitment of Hsp70 to activated Hsp40 co-chaperones.