# Genetic regulation of progenitor cells in appendicular skeletal development

> **NIH NIH R01** · UNIVERSITY OF MINNESOTA · 2022 · $335,412

## Abstract

PROJECT SUMMARY
Congenital limb malformations, caused by abnormal limb development, occur in one in 1,000 live human births.
Therefore, understanding the mechanisms of limb development is relevant to biology and medicine. Limb
development starts with the specification of a discrete region of the lateral plate mesoderm into limb
progenitors, which gives rise to the limb bud. In the last several decades, the research field intensely focused
on understanding the mechanisms by which signaling centers in limb buds regulate patterning of limb buds,
leading to formation of limb skeletons. However, we have limited knowledge about how limb progenitors are
specified and what mechanisms regulate their initial differentiation before the establishment of limb bud
signaling centers; yet, these processes are essential for correct limb development. Studies in the last decades
showed that distinct mechanisms operate on lateral plate mesoderm and limb progenitors prior to establishing
limb bud signaling centers. For example, we found that deletion of Sall4, encoding a zinc finger transcription
factor, approximately two days before the onset of limb development, resulted in severe defects specifically in
hindlimbs, while deletion at later stages had no or subtle effect. In our preliminary studies, we found that
simultaneous inactivation of Sall4, Irx3 and Irx5 (Irx3/5) caused the absence of hindlimbs with the loss of
expression of hindlimb progenitor-specific genes, such as Isl1. This result indicates that combined function of
Sall4 and Irx3/5 specifies lateral plate mesoderm into hindlimb progenitors. In Aim 1, our goal is to elucidate
the molecular mechanisms of hindlimb progenitor specification. We will determine whether SALL4 and IRX3/5
redundantly and directly regulate Isl1 through its enhancer. We will determine genes that act downstream of
Sall4 and Irx3/5 to specify hindlimb progenitors by genomic experiments. We will determine their functions in
specifying hindlimb progenitors by genetic knockout approaches. We also obtained data, strongly suggesting
that Sall4 knockout causes increased glycolysis in limb progenitors, when endogenous glycolysis is
transitioning from high to low activity. Recent studies provided evidence that, beyond supplying energy,
glycolysis mediates fibroblast growth factor signaling in the tail bud and regulates body elongation. Fibroblast
growth factor signaling is one of earliest signaling that regulates limb progenitor differentiation. In Aim 2, we will
test an intriguing hypothesis that Sall4-dependent repression of glycolysis regulates differentiation of limb
progenitors. We will construct metabolomes of wild type and Sall4 mutant limb progenitors to understand
metabolic status and the changes by loss of Sall4. We will test the role of glycolysis by reducing glycolysis in
Sall4 mutant embryos and determine their differentiation, as well as increasing glycolysis in embryos without
mutations. This proposal will generate important b...

## Key facts

- **NIH application ID:** 10478035
- **Project number:** 5R01AR064195-10
- **Recipient organization:** UNIVERSITY OF MINNESOTA
- **Principal Investigator:** Yasuhiko Kawakami
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $335,412
- **Award type:** 5
- **Project period:** 2013-04-01 → 2024-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10478035

## Citation

> US National Institutes of Health, RePORTER application 10478035, Genetic regulation of progenitor cells in appendicular skeletal development (5R01AR064195-10). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/10478035. Licensed CC0.

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