# Arginyl-tRNA beyond translation: mechanism and regulation of protein arginylation

> **NIH NIH R35** · CASE WESTERN RESERVE UNIVERSITY · 2024 · $402,500

## Abstract

Project Summary
Transfer RNA (tRNA) is best known to function in ribosome-mediated protein synthesis. However, in a
less known role, arginyl-tRNA is essential for catalyzing a unique and poorly understood protein post-
translational modification, namely arginylation, that regulates protein turnover. In this arginylation reaction,
ATE1 (Arginyltransferase 1) facilitates arginine transfer to protein targets using a mechanism that
depends on, and is selective for, arginyl-tRNA(Arg) as the donor cofactor. ATE1-mediated protein
arginylation was identified on hundreds of proteins and is recognized as a global regulator of eukaryotic
cellular processes, including embryogenesis, stress responses, and aging. Deregulation of ATE1 is found
in patients with Parkinson’s disease and metastatic prostate, liver, and skin cancers. Nonetheless, how
ATE1 (and other aminoacyl-tRNA transferases) hijacks tRNA from the highly efficient ribosomal protein
synthesis pathways and catalyzes the arginylation reaction remains a mystery. This proposal aims to
elucidate the catalytic mechanism and regulation of ATE1-mediated protein arginylation in vitro and in
cells. We will focus on interrogating the activity of ATE1 and the consequences of arginylation at three
scales. Firstly, we will determine the molecular mechanisms ATE1 selects for arginyl-tRNA(Arg) and
recognizes specific sites in protein targets through an integrative approach combining structural,
biochemical, and biophysical methods. Once determined, this research will allow a better understanding
of the growing classes of aminoacyl-tRNA transferases in general. Secondly, we will quantitatively
determine the consequences of arginylation on target protein turnover in living cells. Protein degradation
usually depends on poly-ubiquitination, a downstream or concurrent event following arginylation, and
through either proteasomal or autophagy-lysosomal pathways. By examining specific model substrates
for proteasome or autophagosome under normal or stressed conditions, we will determine the crosstalk
between arginylation and ubiquitination; delineate the contribution of each degradation pathway; reveal
the kinetics in cells. Lastly, we will investigate whether and how core components of the ribosomal
translation machinery and nutrients affect protein arginylation. Mechanistically, these studies will expand
our knowledge of the regulatory roles of amino acids and tRNAs, enrich our toolbox to study
macromolecule regulation by tRNA-dependent aminoacylation, and reshape how we consider the
functions of the charged tRNAs beyond protein synthesis. Together, this research provides fundamental
knowledge about arginylation, lays the groundwork for discovering novel therapeutic strategies by
modulating ATE1 activity and protein arginylation in Parkinson’s disease and metastatic cancers, and
enables us to build integrative platforms for future research.

## Key facts

- **NIH application ID:** 10895440
- **Project number:** 5R35GM150678-02
- **Recipient organization:** CASE WESTERN RESERVE UNIVERSITY
- **Principal Investigator:** Yi Zhang
- **Activity code:** R35 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $402,500
- **Award type:** 5
- **Project period:** 2023-08-01 → 2028-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10895440

## Citation

> US National Institutes of Health, RePORTER application 10895440, Arginyl-tRNA beyond translation: mechanism and regulation of protein arginylation (5R35GM150678-02). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10895440. Licensed CC0.

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