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

Hosokawa, Hiroyuki; Koizumi, Maria; Masuhara, Kaori; Romero-Wolf, Maile; Tanaka, Tomoaki; Nakayama, Toshinori; Rothenberg, Ellen V.

doi:10.1084/jem.20202648

Cited by 16 publications

(7 citation statements)

References 73 publications

(167 reference statements)

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“…After T lineage commitment and passage through bselection, E proteins can regulate gene products that are not expressed at the DN stage, including CD4, TCRa, and Rorgt (7,(15)(16)(17). Thus, regulation of different suites of T cell genes at different stages of development is a key feature of E protein activity during T cell development, as has also been observed for GATA3 and Runx factors (18,19).…”

Section: Introductionmentioning

confidence: 72%

Regulation of the Signal-Dependent E Protein HEBAlt Through a YYY Motif Is Required for Progression Through T Cell Development

Yoganathan

Rocha

et al. 2022

Front. Immunol.

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The E protein transcription factors E2A and HEB are critical for many developmental processes, including T cell development. We have shown that the Tcf12 locus gives rise to two distinct HEB proteins, with alternative (HEBAlt) and canonical (HEBCan) N-terminal domains, which are co-expressed during early T cell development. While the functional domains of HEBCan have been well studied, the nature of the HEBAlt-specific (Alt) domain has been obscure. Here we provide compelling evidence that the Alt domain provides a site for the molecular integration of cytokine signaling and E protein activity. Our results indicate that phosphorylation of a unique YYY motif in the Alt domain increases HEBAlt activity by 10-fold, and that this increase is dependent on Janus kinase activity. To enable in vivo studies of HEBAlt in the T cell context, we generated ALT-Tg mice, which can be induced to express a HA-tagged HEBAlt coding cassette in the presence of Cre recombinases. Analysis of ALT-Tg mice on the Vav-iCre background revealed a minor change in the ratio of ISP cells to CD8+ SP cells, and a mild shift in the ratio of T cells to B cells in the spleen, but otherwise the thymus, spleen, and bone marrow lymphocyte subsets were comparable at steady state. However, kinetic analysis of T cell development in OP9-DL4 co-cultures revealed a delay in early T cell development and a partial block at the DN to DP transition when HEBAlt levels or activity were increased. We also observed that HEBCan and HEBAlt displayed significant differences in protein stability that were resolved in the thymocyte context. Finally, a proteomic screen identified STAT1 and Xpo1 as potential members of HEBAlt-containing complexes in thymocytes, consistent with JAK-induced activation of HEBAlt accompanied by translocation to the nucleus. Thus, our results show that the Alt domain confers access to multiple layers of post-translational control to HEBAlt that are not available to HEBCan, and thus may serve as a rheostat to tune E protein activity levels as cells move through different thymic signaling environments during T cell development.

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Section: Introductionmentioning

confidence: 72%

Regulation of the Signal-Dependent E Protein HEBAlt Through a YYY Motif Is Required for Progression Through T Cell Development

Yoganathan

Rocha

et al. 2022

Front. Immunol.

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“…The majority of precommitment-specific binding sites for Runx factors (Group 1 sites) have been shown to be actual co-binding sites with PU.1 1, 13 . Besides PU.1, in later pro-T cells other TFs such as GATA3 and Bcl11b can also collaborate with Runx factors at different sites 32, 33 . Notably, the presence of PU.1 can redirect Runx1 occupancy to the PU.1 sites, while depleting Runx1 binding (“theft”) from alternative Runx sites 13 .…”

Section: Resultsmentioning

confidence: 99%

“…2e). PU.1 ( Spi1 ) is normally repressed by Runx1 in Phase 2 only 1, 33 . Its expression was not affected by Runx1 overexpression at day 2 post-infection, but was significantly downregulated even in the DN1 population at day 4 (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Multiple TFs also change activity 21, 30, 31 . Runx TFs themselves can interact physically with multiple TFs at distinct binding sites, suggesting that TF cooperativity could be a major influence on Runx activity 13, 32, 33 . Yet how important are the Runx factors themselves in dictating which factor complexes and which target sites will be active?…”

Section: Introductionmentioning

confidence: 99%

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Runx factors launch T-cell and innate lymphoid programs via direct and gene network-based mechanisms

Shin

Zhou

Wang

et al. 2022

Preprint

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Runx factors are essential for lineage specification of various hematopoietic cells, including T lymphocytes. However, they regulate context-specific genes and occupy distinct genomic regions in different cell types. Here, we show that dynamic Runx binding shifts in early T-cell development are mostly not restricted by local chromatin state but regulated by Runx dosage and functional partners. Runx co-factors compete to recruit a limited pool of Runx factors in early T-progenitors, and a modest increase in Runx protein availability at pre-commitment stages causes premature Runx occupancy at post-commitment binding sites. This results in striking T-lineage developmental acceleration by selectively activating T-identity and innate lymphoid cell programs. These are collectively regulated by Runx together with other, Runx-induced transcription factors that co-occupy Runx target genes and propagate gene network changes.

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“…Runx transcription factors are also potent at counteracting the stem/progenitor module by repressing the majority of key transcription factor coding genes supporting this program ( Mef2c, Lmo2, Lyl1, Bcl11a, Tal1 , and Meis1 ) in a highly concentration-sensitive way ( 84 ). As the cells pass from Phase 1 to Phase 2, Runx1 begins to collaborate with GATA3 to repress Spi1 (PU.1) ( 51 , 210 ) and turns off the downstream target genes of PU.1 as well, Cd34 and Cd44 ( 51 ).…”

Section: Gene Regulatory Network Models For Early T Cell Development ...mentioning

confidence: 99%

Multi-modular structure of the gene regulatory network for specification and commitment of murine T cells

Shin

Rothenberg

2023

Front. Immunol.

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T cells develop from multipotent progenitors by a gradual process dependent on intrathymic Notch signaling and coupled with extensive proliferation. The stages leading them to T-cell lineage commitment are well characterized by single-cell and bulk RNA analyses of sorted populations and by direct measurements of precursor-product relationships. This process depends not only on Notch signaling but also on multiple transcription factors, some associated with stemness and multipotency, some with alternative lineages, and others associated with T-cell fate. These factors interact in opposing or semi-independent T cell gene regulatory network (GRN) subcircuits that are increasingly well defined. A newly comprehensive picture of this network has emerged. Importantly, because key factors in the GRN can bind to markedly different genomic sites at one stage than they do at other stages, the genes they significantly regulate are also stage-specific. Global transcriptome analyses of perturbations have revealed an underlying modular structure to the T-cell commitment GRN, separating decisions to lose “stem-ness” from decisions to block alternative fates. Finally, the updated network sheds light on the intimate relationship between the T-cell program, which depends on the thymus, and the innate lymphoid cell (ILC) program, which does not.

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

Cited by 16 publications

References 73 publications

Regulation of the Signal-Dependent E Protein HEBAlt Through a YYY Motif Is Required for Progression Through T Cell Development

Regulation of the Signal-Dependent E Protein HEBAlt Through a YYY Motif Is Required for Progression Through T Cell Development

Runx factors launch T-cell and innate lymphoid programs via direct and gene network-based mechanisms

Multi-modular structure of the gene regulatory network for specification and commitment of murine T cells

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