# Rewiring of the pluripotency enhancer network during early mammalian development

> **NIH NIH R01** · UNIVERSITY OF CALIFORNIA, SAN FRANCISCO · 2020 · $414,878

## Abstract

PROJECT ABSTRACT:
Pluripotency is the remarkable ability of a single cell to give rise to every cell type of the mammalian body plan.
Pluripotent cells exist in epiblast of early implantation embryos. There are two well-described pluripotent cell
types, those of the early versus late epiblast that can be modeled in vitro as naïve embryonic stem cells versus
primed epiblast cells respectively. These cells differ minimally in terms of their expression profiles, yet vastly in
terms of their epigenomes. In particularly, largely distinct enhancers drive expression of the same genes in the
two states. The reason for the extensive enhancer rewiring in the absence of gene expression changes is
unknown, but appears to be a critical aspect of early mammalian development. Preliminary results begin to
address this problem by following the function of a single transcription factor Grhl2. Grhl2 is upregulated during
embryonic stem to epiblast cell transition and is able to induce previously latent enhancers to a fully active
state driving expression of proximal genes. Yet, these genes do not change expression during the transition.
Evaluation of potential enhancers regulating the same genes in the embryonic stem cells uncovered the
Klf2/4/5- related transcription factors as likely regulators of the genes in the naïve state. Indeed Grhl2 is
upregulated just as Klf2/4/5 is downregulated. However, Klf2/4/5 regulates a much larger network of genes in
the naive state than Grhl2 does in the primed state. Therefore, it appears that Grhl2 assumes control of a
subset of Klf2/4/5 targets during the transition and that other transcription factors must assume control of other
parts of the very large Klf2/4/5 network. These findings led to the hypothesis that during the early to late
epiblast transition, large naïve regulatory networks are broken down into much smaller primed regulatory
networks, providing the late epiblast cells the flexibility to differentiate down the divergent somatic lineages that
form at gastrulation, immediately following the late epiblast stage. Then each of the smaller networks can be
selectively maintained among the different lineages. Indeed, the Grhl2 network is excluded from the primitive
streak while remaining expressed in the surrounding epiblast cells. To test the hypothesis, there are three
specific aims. In aim 1, cutting edge technologies are used to identify all Klf2/4/5 and Grhl2 driven enhancer
promoter interactions in the embryonic stem and epiblast cell states respectively in order to directly determine
whether Grhl2 results in the rewiring of enhancer-promoter interactions among a subset of Klf2/4/5 targets. In
aim 2, bioinformatics and novel biochemical methods are used to uncover additional epiblast cell transcription
factors that rewire other subsets of the Klf2/4/5 driven network. In aim 3, single cell sequencing of wild-type
and knockout embryos is used to explore the biological role for enhancer rewiring in vivo. Successful
c...

## Key facts

- **NIH application ID:** 9988247
- **Project number:** 5R01GM122439-04
- **Recipient organization:** UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
- **Principal Investigator:** Robert Blelloch
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $414,878
- **Award type:** 5
- **Project period:** 2017-09-07 → 2023-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9988247

## Citation

> US National Institutes of Health, RePORTER application 9988247, Rewiring of the pluripotency enhancer network during early mammalian development (5R01GM122439-04). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/9988247. Licensed CC0.

---

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