# Mechanisms of DNA Replication, Chromosome Compaction, and Chromosome Unlinking

> **NIH NIH R35** · SLOAN-KETTERING INST CAN RESEARCH · 2021 · $1,028,611

## Abstract

Summary
Accurate transmission of the genetic information requires complete duplication of the chromosomal DNA each
cell division cycle. The orderly progression of replication forks is challenged by encounters with template
damage, slow moving and arrested RNA polymerases, and frozen DNA-protein complexes that stall the fork.
Stalled forks are foci for genomic instability that causes genetic alterations and can give rise to cancer. Stalled
forks must be remodeled/repaired and replication restarted/continued in order to maintain genomic stability.
 We have developed an Escherichia coli DNA replication system that allows us to analyze the
consequences of collision of the replisome with leading-strand template damage and with which we can model
all aspects of replisome stalling in vitro. In this proposal we investigate the integrated network of responses to
DNA damage that the bacterium uses to preserve genomic integrity. We ask: (i) how do stalled forks contribute
to induction of the DNA damage (SOS) response? (ii) What is the mechanism of the UmuDC DNA replication
checkpoint elaborated by the SOS response? (iii) What are the dynamics of exchange between DNA
polymerase IV and DNA polymerase III during replisome-mediated trans-lesion bypass? And (iv), how do
replisomes overcome collisions with RNA polymerases that are themselves stalled by DNA template damage.
We will begin to apply our expertise to address these questions using human replication proteins and are also
expanding our analyses by using single molecule approaches.
 Coordinating the structural organization of chromosomes is essential for DNA replication, transcription,
and chromosome segregation during cell division. Failure to achieve proper chromosomal organization during
separation can result in DNA breakage, leading to an uneven distribution of the genetic material to the next
generation. Chromosomal organization involves two principal mechanisms: topological maintenance and
protein-mediated packaging of the DNA. The former prevents entanglement by regulating the topology of the
DNA, resolving unwanted catenanes and knots. The latter shapes the conformation of chromosomes,
increasing the efficiency of any particular macromolecular transaction. Our analyses focus on the interaction
between the cellular condesin, MukB, and the cellular decatenase topoisomerase IV that we discovered and
that we have shown to be required for proper chromosome compaction and segregation. We ask: (i) what is
the role of the MukB accessory proteins MukE and MukF and MukB ATP hydrolysis in chromosome
compaction? (ii) What are the DNA-MukB-Topo IV structures that are formed that lead to chromosome
compaction? (iii) How do defects in chromosome compaction affect DNA metabolic processes such as DNA
repair? And (iv) how does the presumptive bacterial cohesin, RecN, function in double-strand break repair and
daughter-strand gap repair?

## Key facts

- **NIH application ID:** 10129200
- **Project number:** 5R35GM126907-04
- **Recipient organization:** SLOAN-KETTERING INST CAN RESEARCH
- **Principal Investigator:** KENNETH J MARIANS
- **Activity code:** R35 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $1,028,611
- **Award type:** 5
- **Project period:** 2018-04-01 → 2023-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10129200

## Citation

> US National Institutes of Health, RePORTER application 10129200, Mechanisms of DNA Replication, Chromosome Compaction, and Chromosome Unlinking (5R35GM126907-04). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10129200. Licensed CC0.

---

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