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Stage-specific action of Runx1 and GATA3 controls silencing of PU.1 expression in mouse pro-T cells

Hosokawa, Hiroyuki and Koizumi, Maria and Masuhara, Kaori and Romero-Wolf, Maile and Tanaka, Tomoaki and Nakayama, Toshinori and Rothenberg, Ellen V. (2021) Stage-specific action of Runx1 and GATA3 controls silencing of PU.1 expression in mouse pro-T cells. Journal of Experimental Medicine, 218 (8). Art. No. e20202648. ISSN 0022-1007. PMCID PMC8241539. doi:10.1084/jem.20202648. https://resolver.caltech.edu/CaltechAUTHORS:20210701-152448019

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Abstract

PU.1 (encoded by Spi1), an ETS-family transcription factor with many hematopoietic roles, is highly expressed in the earliest intrathymic T cell progenitors but must be down-regulated during T lineage commitment. The transcription factors Runx1 and GATA3 have been implicated in this Spi1 repression, but the basis of the timing was unknown. We show that increasing Runx1 and/or GATA3 down-regulates Spi1 expression in pro–T cells, while deletion of these factors after Spi1 down-regulation reactivates its expression. Leveraging the stage specificities of repression and transcription factor binding revealed an unconventional but functional site in Spi1 intron 2. Acute Cas9-mediated deletion or disruption of the Runx and GATA motifs in this element reactivates silenced Spi1 expression in a pro–T cell line, substantially more than disruption of other candidate elements, and counteracts the repression of Spi1 in primary pro–T cells during commitment. Thus, Runx1 and GATA3 work stage specifically through an intronic silencing element in mouse Spi1 to control strength and maintenance of Spi1 repression during T lineage commitment.


Item Type:Article
Related URLs:
URLURL TypeDescription
https://doi.org/10.1084/jem.20202648DOIArticle
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE159960Related ItemData
ORCID:
AuthorORCID
Hosokawa, Hiroyuki0000-0002-9592-2889
Koizumi, Maria0000-0001-8275-5594
Masuhara, Kaori0000-0002-6648-2431
Romero-Wolf, Maile0000-0002-8024-7198
Tanaka, Tomoaki0000-0002-9761-1750
Nakayama, Toshinori0000-0002-1434-2007
Rothenberg, Ellen V.0000-0002-3901-347X
Additional Information:© 2021 Hosokawa et al. This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms/). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 4.0 International license, as described at https://creativecommons.org/licenses/by-nc-sa/4.0/). Submitted: 11 December 2020; Revised: 1 May 2021; Accepted: 10 June 2021. We thank D. Perez, J. Tijerina, P. Cannon, and R. Diamond for cell sorting and advice, I. Soto for mouse colony care, H. Amrhein and D. Trout for computational assistance, I. Antoshechkin for sequencing management, members of the Support Center for Medical Research and Education at Tokai University for their technical help, and the members of the Rothenberg group for valuable discussion and reagents. E.V. Rothenberg was supported by the U.S. Public Health Service (grants R01 AI135200, R01AI083514, and R01HD076915). H. Hosokawa was supported by the Japan Society for the Promotion of Science KAKENHI grant JP19H03692, the Uehara Memorial Foundation, the Naito Foundation, the Takeda Science Foundation, the Yasuda Memorial Medical Foundation, the SENSHIN Medical Research Foundation, the Daiichi Sankyo Foundation of Life Science, the Tokyo Biochemical Research Foundation, a Princess Takamatsu Cancer Research Fund research grant, and the 2020 Tokai University School of Medicine Research Aid. T. Tanaka was supported by Japan Society for the Promotion of Science KAKENHI grant JP19H03708, the Uehara Memorial Foundation, the Naito Foundation, the Takeda Science Foundation, the Yasuda Memorial Medical Foundation, the SENSHIN Medical Research Foundation, the NOVARTIS Foundation (Japan) for the Promotion of Science, a Princess Takamatsu Cancer Research Fund research grant, the KOSE Cosmetology Research Foundation, and the Medical Institute of Bioregulation Kyushu University Cooperative Research Project Program. This work was also partly supported by the Tokai University General Research Organization Research and Study Project (H. Hosokawa), the L.A. Garfinkle Memorial Laboratory Fund and the Al Sherman Foundation, Provost and Division of Biology & Biological Engineering of Caltech special project funds, and the Albert Billings Ruddock Professorship (E.V. Rothenberg). Author contributions: H. Hosokawa designed and supervised the study, performed experiments, analyzed data, and wrote the manuscript; M. Koizumi, K. Masuhara, and M. Romero-Wolf performed experiments and analyzed data; T. Tanaka performed experiments, analyzed data, and edited the manuscript; T. Nakayama provided unique biological reagents and helpful discussions and edited the manuscript; E.V. Rothenberg designed and supervised the study, analyzed data, and wrote the manuscript. Disclosures: E.V. Rothenberg reported personal fees from Century Therapeutics and personal fees from Kite Pharma outside the submitted work. No other disclosures were reported. Data availability: The GEO accession number for all new deep-sequencing data reported in this paper is GSE159960.
Funders:
Funding AgencyGrant Number
NIHR01AI135200
NIHR01AI083514
NIHR01HD076915
Japan Society for the Promotion of Science (JSPS)JP19H03692
Uehara Memorial FoundationUNSPECIFIED
Naito FoundationUNSPECIFIED
Takeda Science FoundationUNSPECIFIED
Yasuda Memorial Medical FoundationUNSPECIFIED
SENSHIN Medical Research FoundationUNSPECIFIED
Daiichi Sankyo Foundation of Life ScienceUNSPECIFIED
Tokyo Biochemical Research FoundationUNSPECIFIED
Princess Takamatsu Cancer Research FundUNSPECIFIED
Tokai UniversityUNSPECIFIED
Japan Society for the Promotion of Science (JSPS)JP19H03708
NOVARTIS FoundationUNSPECIFIED
KOSE Cosmetology Research FoundationUNSPECIFIED
Kyushu UniversityUNSPECIFIED
Louis A. Garfinkle Memorial Laboratory FundUNSPECIFIED
Al Sherman FoundationUNSPECIFIED
Caltech Division of Biology and Biological EngineeringUNSPECIFIED
Albert Billings Ruddock ProfessorshipUNSPECIFIED
Issue or Number:8
PubMed Central ID:PMC8241539
DOI:10.1084/jem.20202648
Record Number:CaltechAUTHORS:20210701-152448019
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20210701-152448019
Official Citation:Hiroyuki Hosokawa, Maria Koizumi, Kaori Masuhara, Maile Romero-Wolf, Tomoaki Tanaka, Toshinori Nakayama, Ellen V. Rothenberg; Stage-specific action of Runx1 and GATA3 controls silencing of PU.1 expression in mouse pro–T cells. J Exp Med 2 August 2021; 218 (8): e20202648. doi: https://doi.org/10.1084/jem.20202648
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:109693
Collection:CaltechAUTHORS
Deposited By: Tony Diaz
Deposited On:06 Jul 2021 17:23
Last Modified:09 Jul 2021 21:05

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