Single-cell epigenetic analysis reveals principles of chromatin states in H3.3-K27M gliomas

Nofar Harpaz; Tamir Mittelman; Olga Beresh; Ofir Griess; Noa Furth; Tomer-Meir Salame; Roni Oren; Liat Fellus-Alyagor; Alon Harmelin; Sanda Alexandrescu; Joana Graca Marques; Mariella G Filbin; Guy Ron; Efrat Shema

doi:10.1016/j.molcel.2022.05.023

Single-cell epigenetic analysis reveals principles of chromatin states in H3.3-K27M gliomas

Nofar Harpaz, Tamir Mittelman, Olga Beresh, Ofir Griess, Noa Furth, Tomer-Meir Salame, Roni Oren, Liat Fellus-Alyagor, Alon Harmelin, Sanda Alexandrescu, Joana Graca Marques, Mariella G Filbin, Guy Ron, Efrat Shema^*

^*Corresponding author for this work

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Abstract

Cancer cells are highly heterogeneous at the transcriptional level and epigenetic state. Methods to study epigenetic heterogeneity are limited in throughput and information obtained per cell. Here, we adapted cytometry by time-of-flight (CyTOF) to analyze a wide panel of histone modifications in primary tumor-derived lines of diffused intrinsic pontine glioma (DIPG). DIPG is a lethal glioma, driven by a histone H3 lysine 27 mutation (H3-K27M). We identified two epigenetically distinct subpopulations in DIPG, reflecting inherent heterogeneity in expression of the mutant histone. These two subpopulations are robust across tumor lines derived from different patients and show differential proliferation capacity and expression of stem cell and differentiation markers. Moreover, we demonstrate the use of these high-dimensional data to elucidate potential interactions between histone modifications and epigenetic alterations during the cell cycle. Our work establishes new concepts for the analysis of epigenetic heterogeneity in cancer that could be applied to diverse biological systems.

Original language	English
Pages (from-to)	2696-2713
Number of pages	18
Journal	Molecular Cell
Volume	82
Issue number	14
Early online date	17 Jun 2022
DOIs	https://doi.org/10.1016/j.molcel.2022.05.023
Publication status	Published - 21 Jul 2022

Bibliographical note

Funding Information:
We thank Y. Aylon and I. Tirosh for their important input. We are grateful to M. Monje for generously sharing with us the SU-DIPG48, SU-DIPG6, SU-DIPG6-GFP, SU-DIPG38, SU-DIPG25, and SU-DIPG13 cultures, and N. Jabado for her kind gift of the isogenic BT245 and SU-DIPG36 cultures. We thank S. Baker, L. Kasper, and the St. Jude Children's Research Hospital Pediatric Brain Tumor Portal, https://pbtp.stjude.cloud, for the SJ-HGGX39 cell line and the shK27M constructs. We thank C. Raanan, M. Zerbib, and I. Savchenko for their contribution in establishing the DIPG mouse model and G. Jona for his assistance with the lentivirus system. The graphical abstract and Figure 1A were created using biorender.com. Funding: E.S. is an incumbent of the Lisa and Jeffrey Aronin Family Career Development chair. This research was supported by grants from the European Research Council (ERC801655), the Israeli Science Foundation (1881/19), The German-Israeli Foundation for Scientific Research and Development, the Israeli Council for Higher Education (CHE) via the Weizmann Data Science Research Center, and Minerva. N.H. T.M. G.R. and E.S. designed the study, performed data analysis, and wrote the manuscript. N.H. T.M. and O.B. conducted the experiments. T.-M.S. assisted in CyTOF runs and panel design. N.F. and O.B. generated lentiviral expression systems. O.G. assisted with RNA sequencing. A.H. R.O. and L.F.-A. established the xenograft mice model and performed immunohistochemistry. S.A. J.G.M. and M.G.F. performed patients’ autopsy analysis. The authors declare no competing interests.

All Science Journal Classification (ASJC) codes

Molecular Biology
Cell Biology

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10.1016/j.molcel.2022.05.023

es_MolCell_SingleCellEpigeneticAnalysis_AM2022Accepted author manuscript, 26.6 MBLicence: CC BY-NC-ND

https://doi.org/10.1101/2021.11.02.466907Licence: CC BY-NC-ND

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Harpaz, N., Mittelman, T., Beresh, O., Griess, O., Furth, N., Salame, T.-M., Oren, R., Fellus-Alyagor, L., Harmelin, A., Alexandrescu, S., Marques, J. G., Filbin, M. G., Ron, G., & Shema, E. (2022). Single-cell epigenetic analysis reveals principles of chromatin states in H3.3-K27M gliomas. Molecular Cell, 82(14), 2696-2713. https://doi.org/10.1016/j.molcel.2022.05.023

@article{8752362b33e04d82abd9587044e4200b,

title = "Single-cell epigenetic analysis reveals principles of chromatin states in H3.3-K27M gliomas",

abstract = "Cancer cells are highly heterogeneous at the transcriptional level and epigenetic state. Methods to study epigenetic heterogeneity are limited in throughput and information obtained per cell. Here, we adapted cytometry by time-of-flight (CyTOF) to analyze a wide panel of histone modifications in primary tumor-derived lines of diffused intrinsic pontine glioma (DIPG). DIPG is a lethal glioma, driven by a histone H3 lysine 27 mutation (H3-K27M). We identified two epigenetically distinct subpopulations in DIPG, reflecting inherent heterogeneity in expression of the mutant histone. These two subpopulations are robust across tumor lines derived from different patients and show differential proliferation capacity and expression of stem cell and differentiation markers. Moreover, we demonstrate the use of these high-dimensional data to elucidate potential interactions between histone modifications and epigenetic alterations during the cell cycle. Our work establishes new concepts for the analysis of epigenetic heterogeneity in cancer that could be applied to diverse biological systems.",

author = "Nofar Harpaz and Tamir Mittelman and Olga Beresh and Ofir Griess and Noa Furth and Tomer-Meir Salame and Roni Oren and Liat Fellus-Alyagor and Alon Harmelin and Sanda Alexandrescu and Marques, {Joana Graca} and Filbin, {Mariella G} and Guy Ron and Efrat Shema",

note = "Funding Information: We thank Y. Aylon and I. Tirosh for their important input. We are grateful to M. Monje for generously sharing with us the SU-DIPG48, SU-DIPG6, SU-DIPG6-GFP, SU-DIPG38, SU-DIPG25, and SU-DIPG13 cultures, and N. Jabado for her kind gift of the isogenic BT245 and SU-DIPG36 cultures. We thank S. Baker, L. Kasper, and the St. Jude Children's Research Hospital Pediatric Brain Tumor Portal, https://pbtp.stjude.cloud, for the SJ-HGGX39 cell line and the shK27M constructs. We thank C. Raanan, M. Zerbib, and I. Savchenko for their contribution in establishing the DIPG mouse model and G. Jona for his assistance with the lentivirus system. The graphical abstract and Figure 1A were created using biorender.com. Funding: E.S. is an incumbent of the Lisa and Jeffrey Aronin Family Career Development chair. This research was supported by grants from the European Research Council (ERC801655), the Israeli Science Foundation (1881/19), The German-Israeli Foundation for Scientific Research and Development, the Israeli Council for Higher Education (CHE) via the Weizmann Data Science Research Center, and Minerva. N.H. T.M. G.R. and E.S. designed the study, performed data analysis, and wrote the manuscript. N.H. T.M. and O.B. conducted the experiments. T.-M.S. assisted in CyTOF runs and panel design. N.F. and O.B. generated lentiviral expression systems. O.G. assisted with RNA sequencing. A.H. R.O. and L.F.-A. established the xenograft mice model and performed immunohistochemistry. S.A. J.G.M. and M.G.F. performed patients{\textquoteright} autopsy analysis. The authors declare no competing interests. ",

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T1 - Single-cell epigenetic analysis reveals principles of chromatin states in H3.3-K27M gliomas

AU - Harpaz, Nofar

AU - Mittelman, Tamir

AU - Beresh, Olga

AU - Griess, Ofir

AU - Furth, Noa

AU - Salame, Tomer-Meir

AU - Oren, Roni

AU - Fellus-Alyagor, Liat

AU - Harmelin, Alon

AU - Alexandrescu, Sanda

AU - Marques, Joana Graca

AU - Filbin, Mariella G

AU - Ron, Guy

AU - Shema, Efrat

N1 - Funding Information: We thank Y. Aylon and I. Tirosh for their important input. We are grateful to M. Monje for generously sharing with us the SU-DIPG48, SU-DIPG6, SU-DIPG6-GFP, SU-DIPG38, SU-DIPG25, and SU-DIPG13 cultures, and N. Jabado for her kind gift of the isogenic BT245 and SU-DIPG36 cultures. We thank S. Baker, L. Kasper, and the St. Jude Children's Research Hospital Pediatric Brain Tumor Portal, https://pbtp.stjude.cloud, for the SJ-HGGX39 cell line and the shK27M constructs. We thank C. Raanan, M. Zerbib, and I. Savchenko for their contribution in establishing the DIPG mouse model and G. Jona for his assistance with the lentivirus system. The graphical abstract and Figure 1A were created using biorender.com. Funding: E.S. is an incumbent of the Lisa and Jeffrey Aronin Family Career Development chair. This research was supported by grants from the European Research Council (ERC801655), the Israeli Science Foundation (1881/19), The German-Israeli Foundation for Scientific Research and Development, the Israeli Council for Higher Education (CHE) via the Weizmann Data Science Research Center, and Minerva. N.H. T.M. G.R. and E.S. designed the study, performed data analysis, and wrote the manuscript. N.H. T.M. and O.B. conducted the experiments. T.-M.S. assisted in CyTOF runs and panel design. N.F. and O.B. generated lentiviral expression systems. O.G. assisted with RNA sequencing. A.H. R.O. and L.F.-A. established the xenograft mice model and performed immunohistochemistry. S.A. J.G.M. and M.G.F. performed patients’ autopsy analysis. The authors declare no competing interests.

PY - 2022/7/21

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N2 - Cancer cells are highly heterogeneous at the transcriptional level and epigenetic state. Methods to study epigenetic heterogeneity are limited in throughput and information obtained per cell. Here, we adapted cytometry by time-of-flight (CyTOF) to analyze a wide panel of histone modifications in primary tumor-derived lines of diffused intrinsic pontine glioma (DIPG). DIPG is a lethal glioma, driven by a histone H3 lysine 27 mutation (H3-K27M). We identified two epigenetically distinct subpopulations in DIPG, reflecting inherent heterogeneity in expression of the mutant histone. These two subpopulations are robust across tumor lines derived from different patients and show differential proliferation capacity and expression of stem cell and differentiation markers. Moreover, we demonstrate the use of these high-dimensional data to elucidate potential interactions between histone modifications and epigenetic alterations during the cell cycle. Our work establishes new concepts for the analysis of epigenetic heterogeneity in cancer that could be applied to diverse biological systems.

AB - Cancer cells are highly heterogeneous at the transcriptional level and epigenetic state. Methods to study epigenetic heterogeneity are limited in throughput and information obtained per cell. Here, we adapted cytometry by time-of-flight (CyTOF) to analyze a wide panel of histone modifications in primary tumor-derived lines of diffused intrinsic pontine glioma (DIPG). DIPG is a lethal glioma, driven by a histone H3 lysine 27 mutation (H3-K27M). We identified two epigenetically distinct subpopulations in DIPG, reflecting inherent heterogeneity in expression of the mutant histone. These two subpopulations are robust across tumor lines derived from different patients and show differential proliferation capacity and expression of stem cell and differentiation markers. Moreover, we demonstrate the use of these high-dimensional data to elucidate potential interactions between histone modifications and epigenetic alterations during the cell cycle. Our work establishes new concepts for the analysis of epigenetic heterogeneity in cancer that could be applied to diverse biological systems.

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Single-cell epigenetic analysis reveals principles of chromatin states in H3.3-K27M gliomas

Abstract

Bibliographical note

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