# Dynamics and regulation of sister chromosome cohesion in E. coli

> **NIH NIH R01** · BAYLOR COLLEGE OF MEDICINE · 2020 · $320,000

## Abstract

PROJECT SUMMARY/ABSTRACT
Premature arrest of DNA replication forks is a major cause of the most severe kinds of DNA
damage that occur in cells, including DNA double strand breaks and chromosome
rearrangements. Genetic change arising from spontaneous fork arrest is now predicted to
exceed that occurring from exogenous sources, and thus is a major source of the genome
instability that causes disease in humans and antibiotic resistance in bacteria. Despite the
potential effects of arrested forks on human health, the root mechanisms that cause and prevent
fork arrest are poorly understood. The long-term goal of this research is to advance our ability to
identify and mitigate the primary causes of replication fork arrest in humans and pathogenic
bacteria by establishing a comprehensive understanding of how, when, and why fork arrest
occurs in E. coli. The objective of this proposal is to determine the sources and mechanisms of
spontaneous replication fork arrest in E. coli, and to identify the cellular mechanisms that
prevent it. The central hypothesis is that replication fork arrest occurs primarily by a topological
mechanism in which DNA helical strain between the replication fork and bound protein blocks
strand unwinding, rather than by direct steric interference between the replisome and protein.
The rationale for this proposal is that understanding the mechanism of fork arrest is critical, as
sterical and topological mechanisms would differ in subsequent effects and regulation. The
objective of this proposal will be achieved through the following specific aims: (1) Determine the
location and source of spontaneous RFBs in the E. coli genome. Utilizing synchronized cells
and a novel chromosome supercoiling assay, this aim will produce an aerial view of how and
where fork pausing occurs over the chromosome, and will investigate the role of DNA topology
in fork arrest. Spontaneous fork pausing will be correlated with binding of major nucleoid
proteins and transcription by ChIP-seq. (2) Define and differentiate replication arrest and restart
in models of two canonical RFBs. Using physiological engineered barriers consisting of either a
stable DNA-bound protein complex or an actively transcribing gene oriented head-on with
replication, this aim will provide a highly quantitative investigation of how fork arrest and
recovery differs between the two major identified classes of RFBs. (3) Identify cellular
mechanisms that increase the stability and progression of replication forks at RFBs. This aim
will test two models of replication fork stability through reduction in helical stress at the fork;
factory replication and chromosome cohesion (catenation of sister duplexes behind the fork). It
will also reveal new and unexpected mechanisms of fork stability through an unbiased screen.

## Key facts

- **NIH application ID:** 9865315
- **Project number:** 1R01GM135368-01
- **Recipient organization:** BAYLOR COLLEGE OF MEDICINE
- **Principal Investigator:** David Bates
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $320,000
- **Award type:** 1
- **Project period:** 2020-03-01 → 2022-02-28

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9865315

## Citation

> US National Institutes of Health, RePORTER application 9865315, Dynamics and regulation of sister chromosome cohesion in E. coli (1R01GM135368-01). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/9865315. Licensed CC0.

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