# Genetic Requirements for Protein Degradation at the Endoplasmic Reticulum Translocon

> **NIH NIH R15** · BALL STATE UNIVERSITY · 2022 · $447,000

## Abstract

PROJECT SUMMARY
Most endomembrane system and secreted eukaryotic proteins traverse the endoplasmic reticulum (ER)
translocon during or shortly after synthesis. Underscoring the critical nature of maintaining functional translocons,
eukaryotes possess multiple conserved translocon quality control (TQC) mechanisms that promote degradation
of channel-clogging proteins. TQC remains poorly understood relative to other ER protein quality control
pathways. The highly conserved yeast zinc metalloprotease Ste24 (ZMPSte24 in mammals) catalyzes
degradation of translocon-clogging proteins. Preliminary data indicate that ER rhomboid-family pseudoprotease
Dfm1 (homologous to mammalian derlin proteins) contributes to TQC, likely via the Ste24 mechanism.
Consistent with the mission of the National Institute of General Medical Sciences, the objective of the proposed
work is an improved understanding of TQC, a medically important facet of cell biology, using yeast as a model
system. Ste24 promotes healthy aging and, by virtue of its function in TQC, is likely to play a protective role
against the progression of diabetes. Further, at least two proteins with profound significance for metabolic
physiology persistently engage translocons. The proposed experiments will test the hypothesis that Dfm1 and
its cofactor, the Cdc48 ATPase, partially extract ubiquitylated clogging proteins from the translocon to enable
cleavage by Ste24, whereupon the cytosolic fragment is degraded by the proteasome and the luminal fragment
is trafficked through the endomembrane system to the vacuole for degradation. The specific aims of this project
are to (1) characterize STE24 function in TQC and (2) determine the role of Dfm1 and Cdc48 in TQC. To address
these objectives, several mechanistic aspects of Ste24 and Dfm1/Cdc48 function in TQC will be explored. The
breadth of Ste24 and Dfm1/Cdc48 TQC substrates will be determined. Experiments will address whether Ste24
TQC substrates are ubiquitylated prior to recognition, if TQC substrates are cleaved in a Ste24-dependent
manner, and how Ste24 TQC substrates are ultimately degraded (i.e. by the proteasome or vacuole). Dfm1
association with the translocon and TQC substrates will be assessed, as will the contributions of Dfm1 and
Cdc48 to Ste24-mediated and Ste24-independent TQC. The role of conserved Dfm1 elements in TQC will be
determined. Finally, yeast lacking DFM1 expressing TQC substrates exhibit a profound fitness defect that is
rapidly and stably suppressed. The genetic determinants of suppression will be identified. Undergraduate and
master’s students will participate in all aspects of this project, including experiment design, data collection, and
communication of results. These experiments will yield novel mechanistic insights into conserved, biomedically
relevant mechanisms of TQC. Upon completion, the proposed work has the strong potential to inform the
development of improved therapeutic strategies for multiple human conditions, i...

## Key facts

- **NIH application ID:** 10512586
- **Project number:** 2R15GM111713-03
- **Recipient organization:** BALL STATE UNIVERSITY
- **Principal Investigator:** Eric Meyer Rubenstein
- **Activity code:** R15 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $447,000
- **Award type:** 2
- **Project period:** 2014-09-01 → 2026-04-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10512586

## Citation

> US National Institutes of Health, RePORTER application 10512586, Genetic Requirements for Protein Degradation at the Endoplasmic Reticulum Translocon (2R15GM111713-03). Retrieved via AI Analytics 2026-05-26 from https://api.ai-analytics.org/grant/nih/10512586. Licensed CC0.

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