# Regulation of Opioid Sensitivity and Tolerance by Ubiquitin Ligase Signaling

> **NIH NIH R01** · SEATTLE CHILDREN'S HOSPITAL · 2022 · $629,577

## Abstract

SUMMARY
Opioid drugs are the most widely used analgesics, but also abused substances. The adverse actions of these
drugs, including peripheral side effects, dependence, tolerance and withdrawal, severely limit their utility for
long term pain management. The -opioid receptor (MOR) is the primary target of opioid analgesia, addiction
and withdrawal. Thus, efforts aimed at developing safer opioid treatments and managing problematic side
effects, such as addiction and withdrawal, require a much deeper understanding of the genetic and molecular
mechanisms that regulate MOR and the behavioral effects of opioid drugs.
 Our long-term goal is to use unbiased, large-scale forward genetics and proteomics, as well as
targeted biochemistry and pharmacology to understand how ubiquitin ligase signaling affects opioid sensitivity
and tolerance at a behavioral level. Towards this goal, we deploy a transgenic MOR (tgMOR) model, in which
mammalian MOR is expressed in the nervous system of C. elegans. This imbues C. elegans with opioid-
sensitive behaviors that we evaluate using computationally automated assays. Importantly, tgMOR animals
exhibit the behavioral hallmarks of opioid responses in higher organisms including acute depressant effects,
desensitization and tolerance. We previously used forward genetics and our tgMOR platform to unveil a novel,
conserved anti-opioid system that functions from C. elegans through rodent behavioral models. This approach
has now identified a ubiquitin ligase that regulates opioid sensitivity and behavioral tolerance to repeated
opioid exposure. Our proposal represents an unprecedented opportunity to decipher how ubiquitin ligase
signaling affects opioid drug responses on a behavioral genetic level.
 Our first aim will use two primary approaches to determine how this ubiquitin ligase shapes opioid
behavioral responses. 1) We will use unbiased, large-scale forward suppressor genetics combined with whole-
genome sequencing and CRISPR/Cas9 editing to identify mutations that rescue abnormal opioid responses in
tgMOR ubiquitin ligase mutants. This will identify genes and genetic networks that are inhibited by ubiquitin
ligase activity to influence opioid responses. 2) We will deploy unbiased, large-scale targeted proteomics with
this ubiquitin ligase that will determine its interactome, binding proteins and putative substrates. Importantly,
this aim features two approaches that are unbiased and large-scale but also complementary. In the second
aim, we use in depth biochemical and structural studies to evaluate candidate substrates for this ubiquitin
ligase. Using orthologous human ubiquitin ligases provides an important translational component to our
proposal. We further aim to develop a high throughput screen (HTS) compatible assay for these human
ubiquitin ligases. Finally, we use this HTS compatible assays in a pilot screen for small molecule inhibitors of
these ubiquitin ligases. Our aims represent first of their kind studies on...

## Key facts

- **NIH application ID:** 10490609
- **Project number:** 1R01DA056370-01
- **Recipient organization:** SEATTLE CHILDREN'S HOSPITAL
- **Principal Investigator:** Brock Grill
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $629,577
- **Award type:** 1
- **Project period:** 2022-07-15 → 2027-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10490609

## Citation

> US National Institutes of Health, RePORTER application 10490609, Regulation of Opioid Sensitivity and Tolerance by Ubiquitin Ligase Signaling (1R01DA056370-01). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10490609. Licensed CC0.

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