# The Cellular and Molecular Effects of Synonymous Mutations

> **NIH NIH R01** · UNIVERSITY OF COLORADO · 2020 · $311,985

## Abstract

Synonymous mutations have traditionally been considered to be silent because they do not change the
encoded amino acid. However, evidence is mounting that synonymous mutations can alter the structure,
stability and/or function of mRNAs. Synonymous mutations have been implicated in cancer and Crohn’s
disease, the development of antibiotic resistance, and bacterial adaptation to novel conditions. Few studies
have investigated the mechanistic basis of the effects of synonymous mutations on mRNA structure, and
investigations that have relied on computational predictions have often failed to identify the reasons for
fitness effects. Further, the effects of non-synonymous mutations often ripple through the metabolic and
regulatory networks in cells; there is no reason to think that synonymous mutations will affect only the
encoded mRNA and nothing else. No previous study has addressed the system-wide effects of
synonymous mutations. The goals of this project are 1) to identify high-impact synonymous mutations that
increase the fitness of E. coli under strong selective pressures, and 2) to elucidate the mechanistic basis of
these fitness effects at the system-wide and molecular levels.
Aim 1 describes the introduction of all possible synonymous mutations into 300 E. coli genes encoding
metabolic enzymes and transcriptional regulators – a total of 312,000 mutations. The mutant cells will be
screened under several different selective pressures to identify high-impact mutations. The vast scale of this
screen sets this project apart from many previous investigations that have focused on mildly deleterious
mutations in individual genes.
Aim 2 addresses the effects of 30 high-impact synonymous mutations on the levels of the encoded mRNAs
and proteins with the goal of identifying 10 that likely operate via different mechanisms. The system-wide
effects of these 10 mutations will be investigated to help us understand why each mutation increases fitness
under specific conditions.
Aim 3 describes investigations of the molecular mechanisms by which 10 high-impact mutations affect the
structure, stability and function of the encoded mRNAs. Possible mechanisms include creation of new
transcriptional start sites, alteration of the binding of small regulatory RNAs, changes in structure that affect
mRNA stability and/or translation, and changes in the structure of an encoded protein due to alterations of
the tempo of translation, which can affect protein folding.
This project will provide an unprecedented look at the frequencies and fitness effects of beneficial
synonymous mutations and a detailed mechanistic understanding of the effects of 10 or more high-impact
synonymous mutations that affect mRNA structure, stability and function in different ways.

## Key facts

- **NIH application ID:** 9926908
- **Project number:** 5R01GM124365-04
- **Recipient organization:** UNIVERSITY OF COLORADO
- **Principal Investigator:** SHELLEY D. COPLEY
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $311,985
- **Award type:** 5
- **Project period:** 2017-09-01 → 2023-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9926908

## Citation

> US National Institutes of Health, RePORTER application 9926908, The Cellular and Molecular Effects of Synonymous Mutations (5R01GM124365-04). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/9926908. Licensed CC0.

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