# Functional assessment of enhancer-gene interactions in vivo

> **NIH NIH R00** · UNIVERSITY OF CALIFORNIA-IRVINE · 2021 · $245,091

## Abstract

PROJECT SUMMARY
Transcriptional enhancers are a predominant category of functional elements in the non-coding portion of the
human genome, far outnumbering the ~20,000 protein-coding genes. Mutations affecting enhancers have
been implicated in human disease, and comprehensively understanding the genome-wide architecture and
function of enhancers remains a major unsolved challenge arising from The Human Genome Project. Despite
substantial progress in mapping of these elements (e.g., by the ENCODE consortium), the in vivo target genes
of enhancers are generally unknown, and the mechanisms of their long range regulation during development
are not well explored. Recently, I developed a novel method that allows manipulation of enhancers at their
endogenous genomic location in mice using CRISPR/Cas9 genome editing (Kvon et al., Cell, 2016). In this
application, I propose to better understand the mechanisms of gene regulation by distant-acting enhancers
through in vivo mouse studies, exploiting this highly efficient CRISPR/Cas9 genome editing technology to
create enhancer knock-out and knock-in mice and employing novel methods to map enhancer-promoter
interactions. I will address the following questions regarding distal enhancer function in the genome: 1) Which
genes do different classes of enhancers regulate? 2) Is there enhancer-promoter specificity for distant-acting
enhancers? and 3) What are the consequences of enhancer loss or replacement on an organism's function?
Mentored phase: First, I propose to adapt CRISPR/Cas9 genome editing for studying long-range enhancer-
gene interactions in vivo using mouse embryonic limb as a model system. I will create a series of enhancer
knock-outs to identify their target gene(s) and a series of enhancer knock-ins to study enhancer-promoter
specificity. Second, I will adopt and optimize the Capture-C technology to identify interaction partners of
enhancers directly in mouse tissues. Independent phase: I will use methods developed in the mentored
phase to systematically map target genes for important developmental enhancers in vivo and to gain a detailed
understanding of the mechanisms governing long-range enhancer-promoter interactions on a genomic scale. I
will also use elucidated enhancer-promoter interactions to study basic principles of long-range enhancer
regulation in the mammalian genome using CRISPR/Cas9 technology. This will enable me to develop several
long-term research directions, focused on the role of enhancer-gene interactions in human evolution, disease,
and development. The main areas of research training will include: 1) Further advancing the use of
CRISPR/Cas9 in mice, 2) Capture-C technology development in mouse tissues, and 3) computational genome
analysis. My mentor (Dr. Len Pennacchio) and co-mentor (Dr. Axel Visel) are leaders in these fields. My career
development activities will focus on skills in key areas of my research, attending courses and workshops,
developing leadership and mentor...

## Key facts

- **NIH application ID:** 10140405
- **Project number:** 5R00HG009682-04
- **Recipient organization:** UNIVERSITY OF CALIFORNIA-IRVINE
- **Principal Investigator:** Evgeny Kvon
- **Activity code:** R00 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $245,091
- **Award type:** 5
- **Project period:** 2020-04-07 → 2023-01-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10140405

## Citation

> US National Institutes of Health, RePORTER application 10140405, Functional assessment of enhancer-gene interactions in vivo (5R00HG009682-04). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10140405. Licensed CC0.

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