# AUTOPHAGIC CLEARANCE OF INACTIVE PROTEASOMES AND RIBOSOMES AS MODELS FOR  PROTEIN QUALITY CONTROL

> **NIH NIH R01** · WASHINGTON UNIVERSITY · 2020 · $289,750

## Abstract

PROJECT SUMMARY
Maintenance of proteostasis is central to cellular fitness and is achieved through sophisticated protein quality control
(PQC) pathways that remove dysfunctional or unwanted proteins and protein complexes that become cytotoxic if allowed
to accumulate. In fact, protein aggregation encouraged by defects in PQC is a hallmark of aging, cancer, and numerous
human ‘aggregation-prone’ pathologies, including amyotrophic lateral sclerosis, Alzheimer’s, Parkinson’s, Huntington’s,
and prion-mediated diseases. Consequently, fully understanding PQC could offer new strategies to mitigate protein
aggregation and subsequent proteotoxic stress, especially as they relate to these disease states.
Recently, ancient and mechanistically conserved PQC pathways were discovered that direct the autophagic elimination of
unwanted or inactive proteasomes and ribosomes, termed proteaphagy and ribophagy, respectively. Notably, a selective
proteaphagic route triggered by inhibition shares features with amyloidogenic protein removal, including sequestration by
the Hsp42 chaperone, ubiquitylation, and subsequent recognition by the autophagic receptors Cue5 or RPN10, suggesting
that inhibitor-induced proteaphagy, and likely ribophagy, offer tractable models to dissect features underpinning PQC and
protein aggregate clearance.
This project aims to characterize proteaphagy and ribophagy induced by inhibition in both Arabidopsis and yeast with the
goal of discovering components central to autophagic PQC. Specifically, we will define how, where, and why the yeast
ubiquitin (Ub) ligases Hul5, Rsp5, and San1 (and their Arabidopsis orthologs) recognize and ubiquitylate dysfunctional
proteasomes, and how this modification triggers autophagy via the Ub-binding Cue5/RPN10 autophagic receptors, using
facile confocal fluorescence microscopic and GFP-fusion cleavage assays together with co-localization studies. A special
focus will be on the mechanism(s) involving Hsp42 that coalesce inactive proteasomes into large cytoplasmic aggregates
similar to those observed for various aggregation-prone proteins. We will determine which proteasome subunits become
modified, where the Ubs are attached, which types of poly-Ub chains are assembled, and identify other factors/post-
translational modifications that might be important, by mass spectrometric analysis of proteasomes purified before and
after inhibition. Using similar methodologies, we will examine how ribosomes are degraded by autophagy after exposure
to translation inhibitors, and examine the role(s) of ubiquitylation and the corresponding ligases, Hsp42, and receptors like
Cue5/RPN10 in this clearance. We will also confirm proteaphagy in human cells and the involvement of the Cue5
ortholog Tollip in this process, and determine if the pathway(s) used for proteophagy and ribophagy also help clear
amyloidogenic proteins. Finally, we will study a new class of autophagic receptors related to RPN10 that might
substantially expand th...

## Key facts

- **NIH application ID:** 9824581
- **Project number:** 5R01GM124452-03
- **Recipient organization:** WASHINGTON UNIVERSITY
- **Principal Investigator:** RICHARD DAVID VIERSTRA
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $289,750
- **Award type:** 5
- **Project period:** 2017-12-11 → 2021-11-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9824581

## Citation

> US National Institutes of Health, RePORTER application 9824581, AUTOPHAGIC CLEARANCE OF INACTIVE PROTEASOMES AND RIBOSOMES AS MODELS FOR  PROTEIN QUALITY CONTROL (5R01GM124452-03). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/9824581. Licensed CC0.

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