# Mechanistic studies of stalled DNA replication fork rescue

> **NIH NIH R01** · STATE UNIVERSITY OF NEW YORK AT BUFFALO · 2020 · $33,639

## Abstract

Abstract
There is a fundamental gap in understanding how stalled DNA replication forks are rescued. Continued existence
of this gap represents an important problem because, until it is filled, a complete and clear understanding of the
mechanism of stalled fork reactivation will be lacking. This understanding is crucial as defects in these repair
mechanisms in higher organisms lead to the accumulation of mutations leading to cancer, and the proposed
studies are therefore directly relevant to human disease. Consequently, the long term goal is to understand the
mechanism of stalled DNA replication fork reactivation. The main objective of this proposal is to understand the
interplay between the single stranded DNA binding protein (SSB) and key fork rescue enzymes on nucleoid
templates and of the subsequent processing events leading to restoration of a fork structure. To achieve this
objective, this proposal is divided into three specific aims: 1), Determine the mechanism(s) of fork regression;
2,) To determine how fork impediments affect fork regression; and 3), Ascertain the effects of nucleoid associated
proteins on fork rescue enzymes. Under the first aim, magnetic tweezers and atomic force microscopy (both in
air and high-speed in buffer) will be used to determine how SSB loading and regression by RecG are affected
by PriA and to ascertain whether RecA and RuvAB are able to catalyze an efficient and unidirectional fork
regression reaction. When the proposed studies for Aim 1 are complete, a clear picture of the events at a
nascent, stalled replication fork will be provided. Under the second aim, the same two single DNA molecule
approaches will be used to provide insight into the effects of replisome impediments on stalled fork rescue, with
high spatial and temporal resolution. At the conclusion of the proposed studies for Aim 2, the effects of DNA
lesions and protein-DNA complexes on fork rescue will be made clear and it is anticipated that the mechanism(s)
for displacing stalled RNA polymerase in the vicinity of forks will be obtained. Under the final aim, magnetic
tweezers to manipulate single molecules of DNA will be used to ascertain the effects of nucleoid associated
proteins (NAPs) on fork rescue. When the proposed studies for Aim 3 are complete, it will be ascertained whether
NAPs catalyze regression on their own and if they assist or inhibit fork rescue enzymes. The proposed research
is innovative because of the combinatorial strategy taken. It is also innovative because of the exciting and novel
single molecule approaches used, the focus on nucleoid templates and an understanding to be gained of how
the primary protein barrier(s) causing replisome stalling are removed. Finally, the work is also innovative because
of the care taken in elucidating how recombination helicases function in the presence of SSB. The proposed
research is significant because it will allow, for the first time, the development of clear models of the mechanistic
eve...

## Key facts

- **NIH application ID:** 9961613
- **Project number:** 5R01GM100156-08
- **Recipient organization:** STATE UNIVERSITY OF NEW YORK AT BUFFALO
- **Principal Investigator:** Piero R Bianco
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $33,639
- **Award type:** 5
- **Project period:** 2013-06-07 → 2020-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9961613

## Citation

> US National Institutes of Health, RePORTER application 9961613, Mechanistic studies of stalled DNA replication fork rescue (5R01GM100156-08). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/9961613. Licensed CC0.

---

*[NIH grants dataset](/datasets/nih-grants) · CC0 1.0*