LIF-independent JAK signalling to chromatin in embryonic stem cells uncovered from an adult stem cell disease

Abstract

Activating mutations in the tyrosine kinase Janus kinase 2 (JAK2) cause myeloproliferative neoplasms, clonal blood stem cell disorders with a propensity for leukaemic transformation. Leukaemia inhibitory factor (LIF) signalling through the JAK-signal transducer and activator of transcription (STAT) pathway enables self-renewal of embryonic stem (ES) cells. Here we show that mouse ES cells carrying the human JAK2V617F mutation were able to self-renew in chemically defined conditions without cytokines or small-molecule inhibitors, independently of JAK signalling through the STAT3 or phosphatidylinositol-3-OH kinase pathways. Phosphorylation of histone H3 tyrosine 41 (H3Y41) by JAK2 was recently shown to interfere with binding of heterochromatin protein 1α (HP1α). Levels of chromatin-bound HP1α were lower in JAK2V617F ES cells but increased following inhibition of JAK2, coincident with a global reduction in histone H3Y41 phosphorylation. JAK2 inhibition reduced levels of the pluripotency regulator Nanog, with a reduction in H3Y41 phosphorylation and concomitant increase in HP1α levels at the Nanog promoter. Furthermore, Nanog was required for factor independence of JAK2V617F ES cells. Taken together, these results uncover a previously unrecognized role for direct signalling to chromatin by JAK2 as an important mediator of ES cell self-renewal.

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Figure 1: JAK2V617F sustains ES cells in a self-renewing state without any additional factors.
Figure 2: Factor-independent JAK2V617 ES cells are capable of multilineage differentiation in vitro and in vivo.
Figure 3: JAK2V617F does not activate canonical signalling pathways independently of cytokines, but JAK is required for ES cell self-renewal.
Figure 4: JAK2 is present in the nucleus of ES cells and dynamically regulates HP1α access to chromatin by phosphorylating histone H3Y41.
Figure 5: JAK2 regulates H3Y41ph at the Nanog promoter and Nanog is critical for factor-independent self-renewal.
Figure 6: JAK2 is not the only JAK that can phosphorylate H3Y41.

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Acknowledgements

We gratefully acknowledge the assistance of S. Kinston for technical support, M. Anderson and T. Hamilton for 8-cell-stage injections, K. Griffiths and A. Johnston for assistance with statistics, N. Ivanova for the Nanog shRNA vector, S. Pollard for use of Incucyte, A. Smith for STAT3-null and Nanog-overexpressing ES cells and for helpful discussions and A. Bradley for helpful discussions. Research in the authors' laboratories is supported by Cancer Research UK, Leukaemia and Lymphoma Research, The Leukaemia and Lymphoma Society and Medical Research Council.

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D.S.G. designed the experiments and performed most of the experiments. B.G. conceived the study and wrote the paper. J.L., P.L. and A.R.G. designed and made the JAK2V617F ES cells. M.A.D., A.J.B. and T.K. performed ChIPs, western for H3Y41ph and in vitro kinase assay. M.W.B.T. analysed microarray data. W.M. and J.N. generated teratocarcinomas and derived JAK2-null ES cells. Y.-H.C. and A.M.S. generated Fig. 2b. D.S.G., A.R.G and B.G. wrote the paper.

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Correspondence to Dean S. Griffiths or Berthold Göttgens.

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Griffiths, D., Li, J., Dawson, M. et al. LIF-independent JAK signalling to chromatin in embryonic stem cells uncovered from an adult stem cell disease. Nat Cell Biol 13, 13–21 (2011). https://doi.org/10.1038/ncb2135

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