# Regulation of Splicing During Hematopoietic Stem and Progenitor Cell Formation

> **NIH NIH F31** · ALBERT EINSTEIN COLLEGE OF MEDICINE · 2024 · $48,974

## Abstract

Project Summary/Abstract
Hematopoietic stem and progenitor cells (HSPCs) sustain lifelong hematopoiesis through self-renewal and
differentiation into all blood cell types. They form during early embryogenesis through a regulated and conserved
process termed the endothelial-to-hematopoietic transition (EHT). During the EHT, a subset of endothelial cells
(ECs) adapt a hematopoietic transcriptional program to form the hemogenic endothelium (HE) followed by
morphological changes to make HSPCs. As de novo production of HSPCs occurs solely during early embryonic
development, HSPC formation has profound consequences on all embryonic and adult hematopoiesis. Defects
in EHT regulators are prevalent in hematologic disorders, therefore lessons learned by studying EHT could
inform the pathways driving these diseases. Studying EHT regulation is critical for understanding processes key
to hematopoietic health from embryo to adult life. Though some factors controlling EHT are known, our
knowledge of HE/HSPC regulators remains poorly understood. In prior studies, the Bowman lab determined that
proper pre-mRNA splicing is required for EHT as HEs were largely absent in zebrafish mutants for the
spliceosomal component splicing factor 3b, subunit 1 (sf3b1). These data indicate that splicing is important for
HE formation, but the mechanisms regulating the splicing changes critical for EHT are largely unknown. To
explore this process, I first defined the alternative splicing signature between embryonic zebrafish EC and
HE/HSPC. Cis-acting regulatory elements within pre-mRNA guide splice isoform selection thus, to identify
potential mechanisms controlling alternative splicing during EHT, I surveyed alternative splicing events between
EC and HE/HSPC for differences in splicing regulatory sequence features. Through this preliminary analysis, I
uncovered that the EHT alternative events were enriched for weaker splice sites and shorter intron length
suggesting these features could have a regulatory function in dictating EHT specific-splice isoform choice. In
addition to sequence-driven mechanisms, alternative splicing can be modified by transcriptional checkpoints
such as promoter-proximal pausing. In pilot studies, I showed that pharmacological or genetic inhibition of
promoter-proximal pausing factors can diminish HE/HSPC production. Based on my data and the literature, I will
test the hypothesis that EHT-specific-splice isoform selection is guided by distinct cis-acting-regulatory elements
(aim 1) and regulated by promoter proximal pausing factors (aim 2). These studies of cell-type specific splicing
regulation in a complex, multicellular context will enable understanding of how splicing fine-tunes the EHT fate
decision. Completion of this study will reveal critical regulation for the genesis of HSPC.

## Key facts

- **NIH application ID:** 10876900
- **Project number:** 5F31HL168882-02
- **Recipient organization:** ALBERT EINSTEIN COLLEGE OF MEDICINE
- **Principal Investigator:** Ilana N Karp
- **Activity code:** F31 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $48,974
- **Award type:** 5
- **Project period:** 2023-04-26 → 2025-04-25

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10876900

## Citation

> US National Institutes of Health, RePORTER application 10876900, Regulation of Splicing During Hematopoietic Stem and Progenitor Cell Formation (5F31HL168882-02). Retrieved via AI Analytics 2026-06-15 from https://api.ai-analytics.org/grant/nih/10876900. Licensed CC0.

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