Resetting the epigenetic balance of Polycomb and COMPASS function at enhancers for cancer therapy

Abstract

The lysine methyltransferase KMT2C (also known as MLL3), a subunit of the COMPASS complex, implements monomethylation of Lys4 on histone H3 (H3K4) at gene enhancers. KMT2C (hereafter referred to as MLL3) frequently incurs point mutations across a range of human tumor types, but precisely how these lesions alter MLL3 function and contribute to oncogenesis is unclear. Here we report a cancer mutational hotspot in MLL3 within the region encoding its plant homeodomain (PHD) repeats and demonstrate that this domain mediates association of MLL3 with the histone H2A deubiquitinase and tumor suppressor BAP1. Cancer-associated mutations in the sequence encoding the MLL3 PHD repeats disrupt the interaction between MLL3 and BAP1 and correlate with poor patient survival. Cancer cells that had PHD-associated MLL3 mutations or lacked BAP1 showed reduced recruitment of MLL3 and the H3K27 demethylase KDM6A (also known as UTX) to gene enhancers. As a result, inhibition of the H3K27 methyltransferase activity of the Polycomb repressive complex 2 (PRC2) in tumor cells harboring BAP1 or MLL3 mutations restored normal gene expression patterns and impaired cell proliferation in vivo. This study provides mechanistic insight into the oncogenic effects of PHD-associated mutations in MLL3 and suggests that restoration of a balanced state of Polycomb–COMPASS activity may have therapeutic efficacy in tumors that bear mutations in the genes encoding these epigenetic factors.

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Fig. 1: MLL3 COMPASS associates with the BAP1 complex.
Fig. 2: Cancer-associated MLL3PHD mutations disrupt MLL3–BAP1 binding and correlate with decreased patient survival.
Fig. 3: BAP1-dependent recruitment of MLL3 COMPASS to enhancers.
Fig. 4: MLL3 regulates tumor suppressor expression from BAP1-depdendent enhancers.
Fig. 5: BAP1 depletion leads to increased H3K27me3 due to loss of UTX-containing MLL3 COMPASS from chromatin.
Fig. 6: Mutations within MLL3PHD sensitize cells to PRC2 inhibition.

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Acknowledgements

We would like to thank F. Zhang (MIT) for the kind gifts of the Px330 and lentiCRISPR v2 vectors. L.W. is supported by the Training Program in Signal Transduction and Cancer (T32 CA070085). Z.Z. is supported by the Robert H. Lurie Comprehensive Cancer Center—Translational Bridge Program Fellowship in Lymphoma Research. E.R.S. is supported by NCI grant R50CA211428. Studies in J.N.S.'s laboratory are supported by NIDCD grant DC013805, and studies related to COMPASS in A.S.'s laboratory are supported by NCI's Outstanding Investigator Award R35CA197569.

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Authors

Contributions

L.W. and A.S. designed the study; L.W. and Z.Z. performed the majority of the experiments and part of the analyses, and wrote the first draft of the manuscript; R.H., N.L., X.H., L.W. and A.S. designed the in vivo studies; R.H., N.L. and X.H. performed and analyzed the in vivo experiments; S.A.M. and E.J.R. generated and sequenced the NGS libraries; K.A.C. and J.N.S. performed the mass spectrometry experiments and analyzed the results; C.K.C. performed the initial bioinformatics analyses on the studies related to the role of BAP1 and MLL3 at enhancers; P.A.O. performed all of the other bioinformatics analyses; J.J.M. provided clinical supervision in the interpretation of data; L.Z. and D.F. performed clinical data analysis; P.N. helped with the UTX ChIP-seq; Y.T. performed size-exclusion chromatography; L.W., Z.Z., M.A.M., E.R.S. and A.S. revised the manuscript.

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Correspondence to Ali Shilatifard.

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Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–20

Reporting Summary

Supplementary Table 1

Raw data for mass spectrometry of MLL3-NTD purification

Supplementary Table 2

Raw data for MLL3 mutations in human cancers

Supplementary Table 3

Genes that are significantly regulated by MLL3, BAP1 and UTX

Supplementary Table 4

Genes that are upregulated by MLL3 for more than twofold

Supplementary Table 5

Genes that are involved in epithelial cell differentiation pathway

Supplementary Table 6

GSEA pathway analysis for genes upregulated by MLL3

Supplementary Table 7

Genes that are upregulated by MLL3 and BAP1 and GSEA pathway analysis

Supplementary Table 8

Genes that are up-regulated by MLL3, BAP1 and UTX and GSEA pathway analysis

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Wang, L., Zhao, Z., Ozark, P.A. et al. Resetting the epigenetic balance of Polycomb and COMPASS function at enhancers for cancer therapy. Nat Med 24, 758–769 (2018). https://doi.org/10.1038/s41591-018-0034-6

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