ABSTRACT
Mutations can disrupt the native function of protein by causing misfolding, which is generally handled by an intricate protein quality control network. To better understand the triaging mechanisms for misfolded cytosolic proteins, we screened a human mutation library to identify a panel of unstable mutations. The degradation of these mutated cytosolic proteins is largely dependent on the ubiquitin proteasome system. Using BioID proximity labelling, we found that the co-chaperones DNAJA1 and DNAJA2 are key interactors with one of the mutated proteins. Notably, the absence of DNAJA2 increases the turnover of the mutant but not the wild-type protein. Our work indicates that specific missense mutations in cytosolic proteins can promote enhanced interactions with molecular chaperones. Assessment of the broader panel of cytosolic mutant proteins shows that the co-chaperone DNAJA2 exhibits two distinct behaviours – acting to stabilize a wide array of cytosolic proteins, including wild-type variants, and to specifically ‘buffer’ some mutant proteins to reduce their turnover. Our work illustrates how distinct elements of the protein homeostasis network are utilized in the presence of a cytosolic misfolded protein.
Footnotes
Author contributions
Conceptualization: H.A.B., J.P.B., V.C., M.T., B.R., T.M.; Methodology: H.A.B., J.P.B., V.C., M.T., B.R.; Validation: H.A.B., J.P.B., V.C., S.K.; Formal analysis: H.A.B., V.C., F.A.Y.; Investigation: H.A.B., J.P.B., V.C., A.M., S.K., H.E., J.L., F.A.Y., E.H.; Resources: H.A.B., J.P.B., V.C., A.M., S.C., G.C.; Writing - original draft: H.A.B., T.M.; Writing - review & editing: H.A.B., T.M.; Visualization: H.A.B., J.P.B., T.M.; Supervision: M.T., B.R., T.M.; Project administration: T.M.; Funding acquisition: M.T., T.M.
Funding
This work was supported by the Canadian Institute of Health Research (CIHR, PJT-159804 to T.M. and M.T.). H.B. is recipient of the CIHR Doctoral Research Award and the Li Tze Fong Memorial Scholarship. J.B. was recipient of a CIHR Postdoctoral Fellowship (MFE-171278). G.C. was supported by a DFG Walter Benjamin fellowship (CA 2559/1-1) and a Michael Smith Foundation of Health Research Research Trainee fellowship (RT-2020-0517).
Data availability
The raw mass spectrometry data has been uploaded to MassIVE (https://massive.ucsd.edu/) with the accession number MSV000094061 (doi:10.25345/C5HQ3S903; BioID data) and MSV000095862 (doi:10.25345/C5Q52FQ9N; pooled CRISPR KO). The data that support the findings of this study are also available from the corresponding author upon request.
