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• W4205687150 abstract "Misfolded proteins in the endoplasmic reticulum (ER) are retrotranslocated to the cytosol for ubiquitination and degradation by the proteasome. During this process, known as ER-associated degradation (ERAD), the ER-embedded Hrd1 ubiquitin ligase plays a central role in recognizing, ubiquitinating, and retrotranslocating scores of lumenal and integral membrane proteins. To better define the mechanisms underlying Hrd1 function in Saccharomyces cerevisiae, several model substrates have been developed. One substrate is Sec61-2, a temperature sensitive allele of the Sec61 translocation channel. Cells expressing Sec61-2 grow at 25 °C because the protein is stable, but sec61-2 yeast are inviable at 38 °C because the mutated protein is degraded in a Hrd1-dependent manner. Therefore, deleting HRD1 stabilizes Sec61-2 and hence sec61-2hrd1∆ double mutants are viable at 38 °C. This unique phenotype allowed us to perform a non-biased screen for loss-of-function alleles in HRD1. Based on its importance in mediating substrate retrotranslocation, the screen was also developed to focus on mutations in sequences encoding Hrd1's transmembrane-rich domain. Ultimately, a group of recessive mutations was identified in HRD1, including an ensemble of destabilizing mutations that resulted in the delivery of Hrd1 to the ERAD pathway. A more stable mutant resided in a buried transmembrane domain, yet the Hrd1 complex was disrupted in yeast expressing this mutant. Together, these data confirm the importance of Hrd1 complex integrity during ERAD, suggest that allosteric interactions between transmembrane domains regulate Hrd1 complex formation, and provide the field with new tools to define the dynamic interactions between ERAD components during substrate retrotranslocation." @default.
• W4205687150 created "2022-01-25" @default.
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• W4205687150 date "2022-01-18" @default.
• W4205687150 modified "2023-09-28" @default.
• W4205687150 title "A positive genetic selection for transmembrane domain mutations in HRD1 underscores the importance of Hrd1 complex integrity during ERAD" @default.
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• W4205687150 doi "https://doi.org/10.1007/s00294-022-01227-1" @default.
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