Functional interrogation of HOXA9 regulome in MLLr leukemia via reporter-based CRISPR/Cas9 screen
- University of Alabama at Birmingham, United States
- St Jude Children's Research Hospital, United States
- St Jude Children's Research Hospital, United States [US]
Abstract
Aberrant HOXA9 expression is a hallmark of most aggressive acute leukemias, notably those with KMT2A (MLL) gene rearrangements. HOXA9 overexpression not only predicts poor diagnosis and outcome but also plays a critical role in leukemia transformation and maintenance. However, our current understanding of HOXA9 regulation in leukemia is limited, hindering development of therapeutic strategies. Here, we generated the HOXA9-mCherry knock-in reporter cell lines to dissect HOXA9 regulation. By utilizing the reporter and CRISPR/Cas9 screens, we identified transcription factors controlling HOXA9 expression, including a novel regulator, USF2, whose depletion significantly down-regulated HOXA9 expression and impaired MLLr leukemia cell proliferation. Ectopic expression of Hoxa9 rescued impaired leukemia cell proliferation upon USF2 loss. Cut&Run analysis revealed the direct occupancy of USF2 at HOXA9 promoter in MLLr leukemia cells. Collectively, the HOXA9 reporter facilitated the functional interrogation of the HOXA9 regulome and has advanced our understanding of the molecular regulation network in HOXA9-driven leukemia.
Data availability
All plasmids created in this study will be deposited to Addgene. Raw data collected from Cut&Run were deposited at NCBI GEO (GSE140664). Raw data collected from CRISPR screening were included in Supplementary File 2. Publicly available dataset used in this study were cited accordingly including Figures 1E and S5D: GSE120781; Figure 1-supplement 1A-C: GSE13159; Figure 3-supplement 2C: GSE126619, GSE74812, GSE89485; Figure 3-supplement 3A: ENCODE (HCT116); Figure 5-supplement 3A-C: European Genome-phenome Archive (EGA) under accession number EGAS00001003266, EGAS00001000654, EGAS00001001952, EGAS00001001923, EGAS00001002217 and EGAS00001000447.
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Acute deletion of CTCF disrupted enhancer-promoter regulation of MYC in human cancer cellsNCBI Gene Expression Omnibus, GSE120781.
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Microarray Innovations in LEukemia (MILE) study: Stage 1 dataNCBI Gene Expression Omnibus, GSE13159.
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Acute deletion of CTCF disrupted enhancer-promoter regulation of MYC in human cancer cellsNCBI Gene Expression Omnibus, GSE126619.
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Therapeutic targeting MLL degradation pathways in MLL-rearranged leukemiaNCBI Gene Expression Omnibus, GSE89485.
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PAX5-driven Subtypes of B-cell Acute Lymphoblastic LeukemiaEuropean Genome-phenome Archive, EGAS00001003266.
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PAX5-driven Subtypes of B-cell Acute Lymphoblastic LeukemiaEuropean Genome-phenome Archive, EGAS00001000654.
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PAX5-driven Subtypes of B-cell Acute Lymphoblastic LeukemiaEuropean Genome-phenome Archive, EGAS00001001952.
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PAX5-driven Subtypes of B-cell Acute Lymphoblastic LeukemiaEuropean Genome-phenome Archive, EGAS00001001923.
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PAX5-driven Subtypes of B-cell Acute Lymphoblastic LeukemiaEuropean Genome-phenome Archive, EGAS00001002217.
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PAX5-driven Subtypes of B-cell Acute Lymphoblastic LeukemiaEuropean Genome-phenome Archive, EGAS00001000447.
Article and author information
Author details
Funding
Leukemia Research Foundation
- Rui Lu
American Cancer Society (IRG15-59-IRG)
- Rui Lu
National Cancer Insititute (P30CA021765-37)
- Chunliang Li
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Xiaobing Shi, Van Andel Institute, United States
Version history
- Received: April 15, 2020
- Accepted: September 30, 2020
- Accepted Manuscript published: October 1, 2020 (version 1)
- Version of Record published: October 30, 2020 (version 2)
Copyright
© 2020, Zhang et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Functional interrogation of HOXA9 regulome in MLLr leukemia via reporter-based CRISPR/Cas9 screen
eLife 9:e57858.
https://doi.org/10.7554/eLife.57858
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