# Translation, targeting, and decay of yeast nonsense-containing mRNAs

> **NIH NIH R35** · UNIV OF MASSACHUSETTS MED SCH WORCESTER · 2024 · $577,959

## Abstract

Project Summary/Abstract
 This proposal addresses the translation and degradation of mRNAs subject to premature
translation termination. Such mRNAs are usually thought to be derived from genes harboring nonsense
mutations, but also include cytoplasmic products of failed or alternative splicing, transcripts of
pseudogenes or unproductive gene rearrangements, and mRNAs with upstream open reading frames, as
well as mRNAs arising from non-standard transcription initiation or undergoing out-of-frame translation
initiation or unexpected frameshifting. Translational elongation on these transcripts generally leads to
ribosomal recognition of a premature termination codon (PTC) and to the triggering of accelerated mRNA
decay in a process known as nonsense-mediated mRNA decay (NMD). The central regulators of this
mRNA quality control pathway are the three Upf proteins (Upf1, Upf2, and Upf3). Although NMD has been
studied extensively in multiple eukaryotes, the precise mechanisms by which the Upf proteins recognize a
ribosome undergoing an atypical termination event and respond to it by triggering accelerated degradation
of the associated mRNA remain unknown. In several recent studies we have begun to gain significant
mechanistic insight into NMD in the yeast model system and to discern what appear to be at least four
definable steps during which: 1) Upf1 binds stochastically to elongating ribosomes while monitoring
translational termination, 2) Upf1 association with a ribosome engaged in premature termination is
stabilized and a commitment to NMD is promoted by Upf2 and Upf3 binding, 3) activation of the Upf1
helicase activity by ATP hydrolysis promotes dissociation of the termination complex and recruitment of
the mRNA decapping enzyme, and 4) post-decapping, the body of the mRNA is digested exonucleolytically
by Xrn1. In the experiments of this proposal we will use the tools of yeast genetics, molecular biology, and
cryo-electron microscopy to pursue several critical aspects of this model by addressing three goals that
seek to: elucidate the mechanism of decapping activation of NMD substrates by Upf1, determine the roles
and functional order for factors that link premature termination to mRNA decapping, and define the basis
for Pab1 enhancement of translation termination efficiency, likely a key step for distinguishing premature
from normal termination. At the conclusion of this study we anticipate being able to formulate a more
integrated model detailing the atypical aspects of premature translation termination, the mechanism by
which premature termination is targeted by the three Upf proteins, and the molecular events linking the
functions of Upf proteins to targeted mRNA decay. We expect that our refined model for yeast NMD will
be generally applicable to the molecular events of NMD in higher organisms and will provide insights to
improving therapeutic approaches to human diseases caused by nonsense mutations.

## Key facts

- **NIH application ID:** 10906732
- **Project number:** 5R35GM148277-02
- **Recipient organization:** UNIV OF MASSACHUSETTS MED SCH WORCESTER
- **Principal Investigator:** Allan S Jacobson
- **Activity code:** R35 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $577,959
- **Award type:** 5
- **Project period:** 2023-08-15 → 2028-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10906732

## Citation

> US National Institutes of Health, RePORTER application 10906732, Translation, targeting, and decay of yeast nonsense-containing mRNAs (5R35GM148277-02). Retrieved via AI Analytics 2026-05-26 from https://api.ai-analytics.org/grant/nih/10906732. Licensed CC0.

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