# Regulatory elements of replication timing and 3D genome organization

> **NIH NIH R35** · UNIVERSITY OF MINNESOTA · 2021 · $197,426

## Abstract

PROJECT SUMMARY
Increasing evidence indicates that three-dimensional (3D) genome organization is required to regulate
gene function and its alterations are associated with many diseases. The genome is organized into
compartments that align with the temporal order of DNA replication (replication timing – RT). However,
little is known about the mechanisms underlying RT control and 3D genome organization. Recently, we
identified a novel class of cis regulatory elements that control RT and 3D genome organization: early
replicating control elements – ERCEs. Our long-term goals are to study what are the regulatory elements
of genome organization in human differentiated cell types, investigate how trans-acting factors control
these elements, and define how 3D genome organization is remodeled during development and
evolution. To achieve these goals, we will delete and insert candidate ERCEs into the genome of human
differentiated cell types and test their effect on RT, 3D genome organization and gene expression. We
will track ERCE activation using highly-synchronous human embryonic stem cells differentiation
systems. Finally, we will analyze 3D genome organization evolution using primary cells derived from
different species. We will use the recently developed technologies for cell purification based on chip-
enclosed microfluidic flow cytometry to isolate live, single cells to establish clonal cell lines carrying
ERCE deletions and insertions. Moreover, we will use this technology for precise identification and
isolation of cell populations in specific cell cycle stages to systematically investigate the connections
between RT, genome architecture and gene expression. Overall, we anticipate that the application of
microfluidic flow cytometry will allow us to optimize our cell line development, as well as our sample
purification for downstream genome-wide analyses. We expect that our work will contribute significantly
to understand the fundamental principles of genome organization and its relationship to gene function.

## Key facts

- **NIH application ID:** 10387585
- **Project number:** 3R35GM137950-02S1
- **Recipient organization:** UNIVERSITY OF MINNESOTA
- **Principal Investigator:** Juan Carlos Rivera-Mulia
- **Activity code:** R35 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $197,426
- **Award type:** 3
- **Project period:** 2020-07-01 → 2025-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10387585

## Citation

> US National Institutes of Health, RePORTER application 10387585, Regulatory elements of replication timing and 3D genome organization (3R35GM137950-02S1). Retrieved via AI Analytics 2026-05-26 from https://api.ai-analytics.org/grant/nih/10387585. Licensed CC0.

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