HRD Complex Self-Remodeling Enables a Novel Route of Membrane Protein Retrotranslocation

Neal, Sonya E.; Syau, Della; Nejatfard, Anahita; Nadeau, Samantha; Hampton, Randolph Y.

doi:10.1016/j.isci.2020.101493

Cited by 19 publications

(35 citation statements)

References 55 publications

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“…Upon deletion of the DFM1 gene, Hrd1 levels increase and the Hrd1 complex may be remodeled, thereby enabling a novel route of membrane protein retro-translocation. This supports the functional flexibility of the Hrd1 complex in response to ER stress [ 42 , 43 ].…”

Section: Er-associated Degradation In Yeastsupporting

confidence: 66%

“…During ubiquitination, Ubc6 attaches the first ubiquitin to a substrate, and Ubc7 extends ubiquitin chains using mostly lysine 48 linkages [ 56 ]. Ubiquitinated substrates are then retro-translocated to the cytosol by Cdc48/p97, which is recruited to the ER membrane by Ubx2 and/or Dfm1 [ 43 , 53 , 57 ]. The mechanism underlying the retro-translocation of membrane substrates, including Doa10 substrates, is ill-defined.…”

Section: Er-associated Degradation In Yeastmentioning

confidence: 99%

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Potential Physiological Relevance of ERAD to the Biosynthesis of GPI-Anchored Proteins in Yeast

Nakatsukasa

2021

IJMS

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Misfolded and/or unassembled secretory and membrane proteins in the endoplasmic reticulum (ER) may be retro-translocated into the cytoplasm, where they undergo ER-associated degradation, or ERAD. The mechanisms by which misfolded proteins are recognized and degraded through this pathway have been studied extensively; however, our understanding of the physiological role of ERAD remains limited. This review describes the biosynthesis and quality control of glycosylphosphatidylinositol (GPI)-anchored proteins and briefly summarizes the relevance of ERAD to these processes. While recent studies suggest that ERAD functions as a fail-safe mechanism for the degradation of misfolded GPI-anchored proteins, several pieces of evidence suggest an intimate interaction between ERAD and the biosynthesis of GPI-anchored proteins.

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Section: Er-associated Degradation In Yeastsupporting

confidence: 66%

Section: Er-associated Degradation In Yeastmentioning

confidence: 99%

Potential Physiological Relevance of ERAD to the Biosynthesis of GPI-Anchored Proteins in Yeast

Nakatsukasa

2021

IJMS

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“…Remarkably, suppression of dfm1Δ alleviates proteotoxic stress by fully restoring retrotranslocation, and chromosome XV is duplicated for the sole purpose of increasing the gene dosage of HRD1 . In a recent analysis, we showed that overexpression of HRD1 allows for self-remodeling of the HRD complex, allowing Hrd1 to retrotranslocate ERAD-M substrates without Dfm1 ( Neal et al , 2020 ); in normal circumstances, ERAD-M retrotranslocation is completely dependent on Dfm1, with no involvement of Hrd1. Thus, elucidating the mechanisms of dfm1Δ suppression led to the discovery of new functions for the HRD complex and an expanded view of Hrd1’s molecular abilities.…”

Section: Resultsmentioning

confidence: 99%

“…Growth assays were carried out as described previously ( Neal et al , 2020 ). Briefly, cells were grown into log phase (OD 600 0.2–0.3) in medium with 2% dextrose.…”

Section: Methodsmentioning

confidence: 99%

Inner-nuclear-membrane–associated degradation employs Dfm1-independent retrotranslocation and alleviates misfolded transmembrane-protein toxicity

Flagg

Wangeline

Holland

et al. 2021

MBoC

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Prior to their delivery to and degradation by the 26S proteasome, misfolded transmembrane proteins of the ER and inner-nuclear membrane must be extracted from lipid bilayers. This extraction process, known as retrotranslocation, requires both quality-control E3 ubiquitin ligases and dislocation factors that diminish the energetic cost of dislodging the transmembrane segments of a protein. Recently, we showed that retrotranslocation of all ER transmembrane proteins requires the Dfm1 rhomboid pseudoprotease. However, we did not investigate whether Dfm1 also mediated retrotranslocation of transmembrane substrates in the inner-nuclear membrane (INM), which is contiguous with the ER but functionally separated from it by nucleoporins. Here, we show that canonical retrotranslocation occurs during INM-associated degradation (INMAD) but proceeds independently of Dfm1. Despite this independence, ERAD-M and INMAD cooperate to mitigate proteotoxicity. We show a novel misfolded-transmembrane-protein toxicity that elicits genetic suppression, demonstrating the cell's ability to tolerate a toxic burden of misfolded transmembrane proteins without functional INMAD or ERAD-M. This strikingly contrasted the suppression of the dfm1Δ null, which leads to the resumption of ERAD-M through HRD-complex remodeling. Thus, we conclude that INM retrotranslocation proceeds through a novel, private channel, which can be studied by virtue of its role in alleviating membrane-associated proteotoxicity.

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“…2A,B) (Neal et al, 2018). Notably, because Hrd1 has been shown to be required for restoring retrotranslocation function when Dfm1 is absent (Neal et al, 2018(Neal et al, , 2020, our screening strain also has Hrd1 missing to prevent suppression of strains during the random mutagenesis screen. The resulting transformants were screened for high colony fluorescence due to buildup of SUS-GFP, indicating Dfm1 loss of function and inability to retrotranslocate SUS-GFP.…”

Section: Dfm1 L1 and Tm2 Mutants Are Unable To Retrotranslocate Er Membrane Substratesmentioning

confidence: 99%

Derlin rhomboid pseudoproteases employ substrate engagement and lipid distortion function for retrotranslocation of ER multi-spanning membrane substrates

Nejatfard

Wauer

Bhaduri

et al. 2021

Preprint

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Nearly one-third of proteins are initially targeted to the endoplasmic reticulum (ER) membrane where they are correctly folded, assembled, and then delivered to their final cellular destinations. In order to prevent the accumulation of misfolded membrane proteins, ER associated degradation (ERAD) moves these clients from the ER membrane to the cytosol; a process known as retrotranslocation. Our recent work in S. cerevisiae has revealed a derlin rhomboid pseudoprotease Dfm1 is involved in the retrotranslocation of ubiquitinated ERAD membrane substrates. In this study we sought to understand the mechanism associated with Dfm1's actions and found that Dfm1's conserved rhomboid residues are critical for membrane protein retrotranslocation. Specifically, we identified several retrotranslocation-deficient Loop 1 mutants that display impaired binding to membrane substrates. Furthermore, Dfm1 has retained the lipid thinning functions of its rhomboid protease predecessors to facilitate in the removal of ER membrane substrates. We find this substrate engagement and lipid thinning feature is conserved in its human homolog, Derlin-1. Utilizing interaction studies and molecular dynamics simulations, this work reveals that rhomboid pseudoprotease derlins employ novel mechanisms of substrate engagement and lipid thinning for catalyzing extraction of multi-spanning membrane substrates.

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HRD Complex Self-Remodeling Enables a Novel Route of Membrane Protein Retrotranslocation

Cited by 19 publications

References 55 publications

Potential Physiological Relevance of ERAD to the Biosynthesis of GPI-Anchored Proteins in Yeast

Potential Physiological Relevance of ERAD to the Biosynthesis of GPI-Anchored Proteins in Yeast

Inner-nuclear-membrane–associated degradation employs Dfm1-independent retrotranslocation and alleviates misfolded transmembrane-protein toxicity

Derlin rhomboid pseudoproteases employ substrate engagement and lipid distortion function for retrotranslocation of ER multi-spanning membrane substrates

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