Differentiation of human pluripotent stem cells into neurons or cortical organoids requires transcriptional co-regulation by UTX and 53BP1

Abstract

UTX is a chromatin modifier required for development and neural lineage specification, but how it controls these biological processes is unclear. To determine the molecular mechanisms of UTX, we identified novel UTX protein interaction partners. Here we show that UTX and 53BP1 directly interact and co-occupy promoters in human embryonic stem cells and differentiating neural progenitor cells. Human 53BP1 contains a UTX-binding site that diverges from its mouse homolog by 41%, and disruption of the 53BP1–UTX interaction abrogated human, but not mouse, neurogenesis in vitro. The 53BP1–UTX interaction is required to upregulate key neurodevelopmental genes during the differentiation of human embryonic stem cells into neurons or into cortical organoids. 53BP1 promotes UTX chromatin binding, and in turn H3K27 modifications and gene activation, at a subset of genomic regions, including neurogenic genes. Overall, our data suggest that the 53BP1–UTX interaction supports the activation of key genes required for human neurodevelopment.

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Fig. 1: 53BP1 binds UTX in hESCs.
Fig. 2: UTX and 53BP1 co-occupy promoters genome wide.
Fig. 3: UTX and 53BP1 binding correlates to gene activation in hNPCs.
Fig. 4: 53BP1 is required for the differentiation of neurons on monolayer culture and three-dimensional cortical organoids.
Fig. 5: UTX and 53BP1 promote a common set of genes related to nervous system development.
Fig. 6: The 53BP1–UTX interaction promotes the development of cortical organoids.
Fig. 7: 53BP1 affects H3K27me3 and H3K27ac during neural differentiation.

Code availability

Bash script for ChIP-seq is available from figshare https://doi.org/10.6084/m9.figshare.7411835.

Data availability

All sequencing data have been deposited in the Gene Expression Omnibus with the accession code GSE108116. Data that support the findings of this study are available from the corresponding author upon reasonable request.

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Acknowledgements

The authors thank M. A. Dyer for helpful discussions; X. Cao, and Y.-G. Han for helpful comments; A. Andersen and C. Guess for editing the manuscript; J. Houston for cell sorting; S. Olsen and D. Roeber for sequencing samples; A. Nityanandam for optimizing the cortical organoid protocol and technical advice; V. Stewart for mouse embryo injection; S. Porter and S. Miller for mi-seq analysis of in-frame clones and mouse embryos; L. Ding and M. Rusch for sequencing mapping; V. Frohlich, J. Klein, L. Griffiths, J. Peters, L. Milburn, and Y. Li for experimental assistance. M.E. is funded by the NIH (1F32HD093276). J.P. is funded by American Lebanese Syrian Associated Charities and NIH (R01GM114260, R01AG047928, and R01AG053987). J.C.P is funded by American Lebanese Syrian Associated Charities and American Cancer Society (132096-RSG-18-032-01-DDC).

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X.Y.: IP-WB, ChIP, 53BP1-KO, 53BP1 in-frame deletions, cortical organoids. B.X.: all seq analysis except whole-genome sequencing. B.M.: initial IP for mass spectrometry, IP-WB, and UTX mutant. M.E.: 53bp1 mutant analyses. S.J.: recombinant protein co-IP-WB. Y.-D.W.: whole-genome sequence analysis. V.P. and J.P.: proteomic analyses. A.P.: transcript profilings and image quantification. Y.F.: B.X. supervision. J.C.P: project design, ChIP, IP, data analysis, and manuscript writing.

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Correspondence to Jamy C. Peng.

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Yang, X., Xu, B., Mulvey, B. et al. Differentiation of human pluripotent stem cells into neurons or cortical organoids requires transcriptional co-regulation by UTX and 53BP1. Nat Neurosci 22, 362–373 (2019). https://doi.org/10.1038/s41593-018-0328-5

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