# The molecular grammar of human RNA biology

> **NIH NIH R35** · UNIVERSITY OF CALIFORNIA, SAN FRANCISCO · 2024 · $134,932

## Abstract

Project Summary/Abstract
The overarching goal of my research is to define how the molecular grammar of RNA molecules regulates
gene expression. To accomplish this goal, I have pursued extensive and interdisciplinary training in the
molecular, cellular, and systems biology understanding of human RNA biology. I seek to build upon my
documented track record in RNA biology and method development to address pressing questions in RNA
biology. This proposal identifies two emphasis areas of importance for inquiry in the next five years. The first
emphasis of this proposal builds on my decade of research into the RNA–binding protein DDX3 by identifying
and exploring critical gaps in understanding. DDX3 is an essential ATP-dependent RNA–binding protein that
couples ATP binding to local RNA duplex unwinding and ribonucleoprotein remodeling. Prior work from my
group and others has implicated DDX3 in translational control for mRNA molecules containing a variety of
mRNA elements, but the precise mechanism, important mRNA features, and genetic interactions remain
incompletely understood. Here, we seek to define how DDX3 interacts with the ribosome to mediate
translational control, to use a new assay we developed to define DDX3-dependent translation in an unbiased
manner, to define how depletion of DDX3 versus missense variants differ in genetic interactions, and to
establish the mechanism leading to changes in RNA levels following DDX3 depletion. The second emphasis of
this proposal advances new developments in my group that enable single-molecule measurement of RNA-
protein interactions in cells. We evolved a new deamination-based molecular recorder to capture RNA-protein
interactions by modifying the sequence of RNA adjacent to an RNA-protein interaction. Through long-read
sequencing we can then identify regions in RNA that were bound by a protein tagged with the deaminase.
Using this approach, we find unexpected heterogeneity in RNA–binding protein sites on individual mRNA
molecules. We propose to build upon these findings, both to understand the mechanistic and functional
implications of this heterogeneity and to extend our approach to new RNA–binding proteins. Overall, the
proposed research is aligned with my research goal by defining the mechanism of important RNA–binding
proteins and by developing new approaches to measure single-molecule RNA biology. I expect the results of
the proposed research to advance the understanding of RNA biology with implications for the fundamental
understanding of RNA, human disease, and mRNA therapeutics.

## Key facts

- **NIH application ID:** 11041342
- **Project number:** 3R35GM149255-01S1
- **Recipient organization:** UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
- **Principal Investigator:** Stephen Nicholas Floor
- **Activity code:** R35 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $134,932
- **Award type:** 3
- **Project period:** 2023-05-01 → 2024-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/11041342

## Citation

> US National Institutes of Health, RePORTER application 11041342, The molecular grammar of human RNA biology (3R35GM149255-01S1). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/11041342. Licensed CC0.

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