Nucleotide exchange factors Fes1 and HspBP1 mimic substrate to release misfolded proteins from Hsp70

Abstract

Protein quality control depends on the tight regulation of interactions between molecular chaperones and polypeptide substrates. Substrate release from the chaperone Hsp70 is triggered by nucleotide-exchange factors (NEFs) that control folding and degradation fates via poorly understood mechanisms. We found that the armadillo-type NEFs budding yeast Fes1 and its human homolog HspBP1 employ flexible N-terminal release domains (RDs) with substrate-mimicking properties to ensure the efficient release of persistent substrates from Hsp70. The RD contacts the substrate-binding domain of the chaperone, competes with peptide substrate for binding and is essential for proper function in yeast and mammalian cells. Thus, the armadillo domain engages Hsp70 to trigger nucleotide exchange, whereas the RD safeguards the release of substrates. Our findings provide fundamental mechanistic insight into the functional specialization of Hsp70 NEFs and have implications for the understanding of proteostasis-related disorders, including Marinesco-Sjögren syndrome.

Recent advances in the structural and mechanistic aspects of Hsp70 molecular chaperones

“…The NBDs of all structures are aligned to subdomains IA, IB, and IIA of the NBD of bovine Hsc70. Dashed lines at the N termini of GrpE and Sil1 indicate the unstructured regions not present in the crystal structure and proposed to bind into the substrate-binding pocket of Hsp70 (115,116). The N-terminal methionine (M1) and the first residue in the structure are indicated.…”

“…NEFs can also have a function beyond accelerating nucleotide exchange. Yeast Fes1 and human HspBP1 have a long unstructured N-terminal sequence that was shown to bind into the substrate-binding pocket and to prevent rebinding of substrates after nucleotide exchange and ATP-binding-mediated release (115). This function is essential for Fes1 activity in yeast.…”

The ribosome-associated complex RAC serves in a relay that directs nascent chains to Ssb

“…In line with this notion the Zuo1 LP-motif negatively modulates nascent chain binding to Ssz1. This previously unknown function of Zuo1 in some respect resembles a recently discovered supplementary function of the yeast Hsp70 nucleotide exchange factor (NEF) Fes1 36 . Fes1 does not only act as a NEF, but via its flexible N-terminal domain mimics Hsp70 substrates.…”

“…Fes1 does not only act as a NEF, but via its flexible N-terminal domain mimics Hsp70 substrates. Accordingly, binding of the Fes1 N-terminal domain to the Hsp70-SBD promotes substrate release 36 . Ssz1 does not hydrolyze ATP and thus does not require a classical NEF as a co-chaperone.…”

Hsp70–Hsp110 chaperones deliver ubiquitin-dependent and -independent substrates to the 26S proteasome for proteolysis in yeast

“…The transfer of misfolded proteins from chaperone systems to the UPS may rely on simple kinetic competition for substrate binding as well as on more intricate mechanisms that physically link chaperones to the UPS machinery. For example in the yeast cytoplasm, the degradation of misfolded proteins that are associated with the chaperone Hsp70 depends on the nucleotide exchange factor Fes1 being released from the chaperone and then recognized by PQC ubiquitin ligases (Gowda et al, 2016(Gowda et al, , 2013(Gowda et al, , 2018. In metazoan cells, mechanisms involve the direct docking of the ubiquitin ligase CHIP to Hsp70, and the nucleotide exchange factor BAG-1 coordinates the release of the misfolded protein from Hsp70 with proteasome interactions (Demand et al, 2001;Hohfeld and Jentsch, 1997;Lüders et al, 2000).…”