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Cytoplasmic Hsp70 and Hsp90 regulate functionally distinct ER quality control checkpoints during thiazide‐sensitive NaCl cotransporter biogenesis
Author(s) -
Donnelly Bridget F.,
Needham Patrick G.,
Snyder Avin C.,
Roy Ankita,
Khadem Shaheen,
Brodsky Jeffrey L.,
Subramanya Arohan R.
Publication year - 2013
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.27.1_supplement.1210.1
Subject(s) - hsp90 , endoplasmic reticulum associated protein degradation , hsp70 , chaperone (clinical) , microbiology and biotechnology , biology , heat shock protein , endoplasmic reticulum , unfolded protein response , chemistry , biochemistry , medicine , gene , pathology
Mutations of the thiazide‐sensitive NaCl cotransporter (NCC) cause Gitelman syndrome, a recessive renal salt‐wasting disorder whose carrier state protects against essential hypertension. Given its complex topology, NCC is inefficiently processed and prone to ER‐associated degradation (ERAD). In Gitelman syndrome, misfolded mutants are targeted for ERAD more strongly, highlighting the importance of NCC quality control in human disease and hypertension resistance. Previously, we reported that the cytoplasmic chaperone Hsp70 stimulates NCC ERAD. Here, we identify Hsp90 as a chaperone that promotes NCC folding. In MDCK cells, disruption of the Hsp90 chaperone cycle accelerated NCC ERAD, and augmenting NCC transfer from Hsp70 to Hsp90 by coexpressing the organizer cochaperone HOP stabilized NCC turnover. In a yeast expression system, Hsp70/Ssa1 inactivation effectively stabilized NCC, while ablation of Hsp90 or HOP homologs only mildly increased NCC turnover, suggesting that Hsp70 selects NCC for ERAD prior to Hsp90 engagement. Consistent with these findings, adjusting the folding environment in MDCK and HEK cells via reduced temperature (25°C) enhanced NCC ER export, increased Hsp90‐NCC interaction, and diminished Hsp70 association. These observations indicate that Hsp70 and Hsp90 comprise distinct checkpoints that sequentially monitor the early stages of NCC biogenesis. Support: VA, AHA, NIH