# Defining the mechanisms regulating MLLT3 expression in human hematopoietic stem cells

> **NIH NIH R01** · UNIVERSITY OF CALIFORNIA LOS ANGELES · 2020 · $280,800

## Abstract

SUMMARY
Hematopoietic stem cells (HSCs) sustain life-long blood formation due to their ability to self-renew and
differentiate into all mature blood cell types (referred to as “stemness”). Transplantation of HSC-containing grafts
is a life-saving therapy for multiple blood disorders; however, shortage of immunologically matched donors limits
the number of patients that can be treated. Expansion/generation of human HSCs in culture would greatly
improve transplantation therapy but has been unsuccessful due to poor understanding of the underlying biology
of HSC stemness. We identified MLLT3 as HSC regulator that is highly enriched in human HSC at all stages
once they have emerged from AGM and expand in the fetal liver (FL); however, MLLT3 expression declines in
cultured HSC. Loss and gain of function studies on human FL and cord blood (CB) HSCs showed that MLLT3 is
critical for their self-renewal, and when its expression is restored, it enables HSC expansion in culture by
protecting their “stemness” program. Importantly, FL and CB HSC expanded in culture showed 10-30 fold
increase in human engraftment in NSG mice, and ability to sustain the HSPC compartment and multilineage
hematopoiesis without malignant transformation or differentiation block. This finding offers an unprecedented
opportunity to understand how human HSC stemness is controlled, and harness MLLT3 for clinical use. So far,
the regulatory mechanisms that govern MLLT3 expression in HSC are completely unknown. Using a combination
of epigenetic studies and bioinformatic approaches, we identified several candidate enhancers in MLLT3 gene
that may regulate its two isoforms. Our data suggests that these enhancers are epigenetically remodeled during
differentiation and silenced during HSC culture, providing new avenues to understand why cultured HSC lose
MLLT3 expression. To understand how MLLT3 expression is regulated in human HSCs, we propose two
complementary approaches: 1) CRISPR/Cas9-mediated epigenome editing to dissect the role of MLLT3
enhancers, 2) combination of bioinformatic and experimental approaches using lentiviral overexpression to
identify MLLT3 upstream regulators. Success in these approaches could help generate/expand human HSCs in
culture and thereby increase the availability of HSCs for transplantation.

## Key facts

- **NIH application ID:** 9894797
- **Project number:** 5R01DK121557-02
- **Recipient organization:** UNIVERSITY OF CALIFORNIA LOS ANGELES
- **Principal Investigator:** Hanna Katri Annikki Mikkola
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $280,800
- **Award type:** 5
- **Project period:** 2019-03-16 → 2021-12-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9894797

## Citation

> US National Institutes of Health, RePORTER application 9894797, Defining the mechanisms regulating MLLT3 expression in human hematopoietic stem cells (5R01DK121557-02). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/9894797. Licensed CC0.

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