KW: Investigation, Funding acquisition, Validation, Writing - original draft, Writing - review and editing, Formal analysis
KR: Investigation, Validation, Writing - review and editing, Formal analysis
ME: Investigation, Writing - review and editing
AS: Investigation, Writing - review and editing
BV: Investigation, Writing - review and editing, Funding acquisition
CI: Investigation, Writing - review and editing
JO: Supervision, Writing - review and editing
PS: Supervision, Writing - review and editing, Funding acquisition
ER: Conceptualization, Methodology, Supervision, Writing - review and editing, Formal analysis, Writing - original draft, Funding acquisition, Project administration
The heterotrimeric Asi ubiquitin ligase (encoded by
Organelle proteome maintenance is essential for eukaryotic life. Several dedicated proteolytic systems promote organelle-specific turnover of misfolded or excess proteins. Inner nuclear membrane (INM) proteins are targeted for proteasomal destruction via INM-associated degradation (INMAD). At least three ubiquitin ligases mediate INMAD in the budding yeast
Asi contributes to protein quantity control (e.g. degradation of orphan subunits of oligosaccharyl transferase and glycosylphosphatidylinositol transamidase complexes (Natarajan
The aminoglycoside hygromycin B produced by the bacterium
We hypothesized that Asi is an important mediator of PQC. We predicted that the Asi complex would be required for resistance to conditions expected to increase the abundance of aberrant proteins. To test this, we cultured wild type yeast, yeast lacking genes encoding each subunit of the Asi complex, and a panel of PQC mutant yeast strains in the absence and presence of increasing concentrations of hygromycin B (Figure 1A). Consistent with previous results, loss of
Loss of
The finding that loss of Hul5p does not enhance sensitivity to hygromycin B was surprising, given multiple characterized functions of Hul5p in PQC. Among other roles, Hul5p promotes degradation of substrates that have escaped detection by the ribosome quality control ubiquitin ligase Ltn1p (Sitron and Brandman, 2019) and promotes turnover of misfolded proteins following heat shock (Fang
Multiple lines of evidence suggest that a subcomplex of the Asi ubiquitin ligase including Asi1p and Asi3p (but not Asi2p) mediates PQC degradation of misfolded proteins, potentially in complex with unidentified substrate adaptors. First, as demonstrated here, deletion of
Asi2p function is also dispensable for degradation of some Asi1/3p substrates that do not possess features rendering them predicted PQC substrates (Khmelinskii
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VJY60 (alias W303) |
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1B | (Thomas and Rothstein, 1989) |
VJY85 |
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1B |
(Wang
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VJY360 |
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1A |
(Tong
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VJY469 |
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1A |
(Tong
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VJY476 (alias BY4741) |
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1A, 1C |
(Tong
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VJY511 |
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1A, 1C |
(Tong
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VJY662 |
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1A |
(Tong
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VJY667 |
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1A |
(Tong
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VJY696 |
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1A |
(Tong
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VJY852 |
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1A, 1C |
(Tong
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VJY969 |
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1C | This study |
VJY970 |
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1C | This study |
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This work was funded by NIH grant R15 GM111713 (EMR). Work in the lab of PJS is funding by NIH grant R15 G067291 and NIH grant R15 CA252996. KAW and BMV were supported by Ball State University Honors Undergraduate Fellowships. This project was conceived while EMR was supported in part by a Ball State University Excellence in Teaching award (sponsored by the Ball State University Division of Online and Strategic Learning and the Office of the Provost).