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Published July 2, 2018 | Supplemental Material
Journal Article Open

Substrate relay in an Hsp70‐cochaperone cascade safeguards tail‐anchored membrane protein targeting


Membrane proteins are aggregation‐prone in aqueous environments, and their biogenesis poses acute challenges to cellular protein homeostasis. How the chaperone network effectively protects integral membrane proteins during their post‐translational targeting is not well understood. Here, biochemical reconstitutions showed that the yeast cytosolic Hsp70 is responsible for capturing newly synthesized tail‐anchored membrane proteins (TAs) in the soluble form. Moreover, direct interaction of Hsp70 with the cochaperone Sgt2 initiates a sequential series of TA relays to the dedicated TA targeting factor Get3. In contrast to direct loading of TAs to downstream chaperones, stepwise substrate loading via Hsp70 maintains the solubility and targeting competence of TAs, ensuring their efficient delivery to the endoplasmic reticulum (ER). Inactivation of cytosolic Hsp70 severely impairs TA translocation in vivo. Our results demonstrate a new role of cytosolic Hsp70 in directly assisting the targeting of an essential class of integral membrane proteins and provide a paradigm for how "substrate funneling" through a chaperone cascade preserves the conformational quality of nascent membrane proteins during their biogenesis.

Additional Information

© 2018 The Authors. Received 16 February 2018; Revised 24 May 2018; Accepted 29 May 2018. We thank E. A. Craig for the Ssa antibody and the SSA1 and ssa1ts strains, V. Denic for the SGT2FLAG∆get3 and sgt2TPRmtFLAG strains, U. S. Chio, M. Rao, A. Montequin, F.C. Liang, A. Siegel for reagents, and the Dougherty lab for use of HPLC. We thank Bil Clemons, Ray Deshaies, and members of the Shan, Clemons, and Deshaies laboratories for discussions and critical comments on the manuscript. This work was supported by NIH grant GM107368 and grant from the Weston Havens Foundation to S.S. Author contributions: HC and SS designed experiment; HC performed experiments and analyzed the data; HC and SS wrote the paper. The authors declare that they have no conflict of interest.

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