The Type II Hsp40 Sis1 cooperates with Hsp70 and the E3 ligase Ubr1 to promote degradation of terminally misfolded cytosolic protein.
Mechanisms for cooperation between the cytosolic Hsp70 system and the ubiquitin proteasome system during protein triage are not clear. Herein, we identify new mechanisms for selection of misfolded cytosolic proteins for degradation via defining functional interactions between specific cytosolic Hsp70/Hsp40 pairs and quality control ubiquitin ligases. These studies revolved around the use of S. cerevisiae to elucidate the degradation pathway of a terminally misfolded reporter protein, short-lived GFP (slGFP). The Type I Hsp40 Ydj1 acts with Hsp70 to suppress slGFP aggregation. In contrast, the Type II Hsp40 Sis1 is required for proteasomal degradation of slGFP. Sis1 and Hsp70 operate sequentially with the quality control E3 ubiquitin ligase Ubr1 to target slGFP for degradation. Compromise of Sis1 or Ubr1 function leads slGFP to accumulate in a Triton X-100-soluble state with slGFP degradation intermediates being concentrated into perinuclear and peripheral puncta. Interestingly, when Sis1 activity is low the slGFP that is concentrated into puncta can be liberated from puncta and subsequently degraded. Conversely, in the absence of Ubr1, slGFP and the puncta that contain slGFP are relatively stable. Ubr1 mediates proteasomal degradation of slGFP that is released from cytosolic protein handling centers. Pathways for proteasomal degradation of misfolded cytosolic proteins involve functional interplay between Type II Hsp40/Hsp70 chaperone pairs, PQC E3 ligases, and storage depots for misfolded proteins.
Department of Cell and Developmental Biology, School of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA.
Daniel W
DW
Summers
Katie J
KJ
Wolfe
Hong Yu
HY
Ren
Douglas M
DM
Cyr
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000595:Amino Acid Sequence; D003513:Cycloheximide / Q000494:pharmacology; D003600:Cytosol / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000378:metabolism; D050956:HSP40 Heat-Shock Proteins / Q000378:metabolism*; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D017510:Protein Folding / Q000187:drug effects; D055550:Protein Stability; D059748:Proteolysis / Q000187:drug effects; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D013379:Substrate Specificity / Q000187:drug effects; D025801:Ubiquitin / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism*
D050956:HSP40 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins; C070320:SIS1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D025801:Ubiquitin; D049452:Green Fluorescent Proteins; D003513:Cycloheximide; C066066:UBR1 protein, S cerevisiae; D044767:Ubiquitin-Protein Ligases; D046988:Proteasome Endopeptidase Complex; D000251:Adenosine Triphosphatases; C095973:SSA1 protein, S cerevisiae