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Haematopoietic malignancies caused by dysregulation of a chromatin-binding PHD finger.

Wang GG, Song J, Wang Z, Dormann HL, Casadio F, Li H, Luo JL, Patel DJ, Allis CD - Nature (2009)

Bottom Line: In these processes, a PHD finger that specifically recognizes H3K4me3/2 marks was essential for leukaemogenesis.Mutations in PHD fingers that abrogated H3K4me3 binding also abolished leukaemic transformation.Collectively, our studies represent, to our knowledge, the first report that deregulation of the PHD finger, an 'effector' of specific histone modification, perturbs the epigenetic dynamics on developmentally critical loci, catastrophizes cellular fate decision-making, and even causes oncogenesis during mammalian development.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Chromatin Biology & Epigenetics, The Rockefeller University, New York, New York 10065, USA.

ABSTRACT
Histone H3 lysine 4 methylation (H3K4me) has been proposed as a critical component in regulating gene expression, epigenetic states, and cellular identities1. The biological meaning of H3K4me is interpreted by conserved modules including plant homeodomain (PHD) fingers that recognize varied H3K4me states. The dysregulation of PHD fingers has been implicated in several human diseases, including cancers and immune or neurological disorders. Here we report that fusing an H3K4-trimethylation (H3K4me3)-binding PHD finger, such as the carboxy-terminal PHD finger of PHF23 or JARID1A (also known as KDM5A or RBBP2), to a common fusion partner nucleoporin-98 (NUP98) as identified in human leukaemias, generated potent oncoproteins that arrested haematopoietic differentiation and induced acute myeloid leukaemia in murine models. In these processes, a PHD finger that specifically recognizes H3K4me3/2 marks was essential for leukaemogenesis. Mutations in PHD fingers that abrogated H3K4me3 binding also abolished leukaemic transformation. NUP98-PHD fusion prevented the differentiation-associated removal of H3K4me3 at many loci encoding lineage-specific transcription factors (Hox(s), Gata3, Meis1, Eya1 and Pbx1), and enforced their active gene transcription in murine haematopoietic stem/progenitor cells. Mechanistically, NUP98-PHD fusions act as 'chromatin boundary factors', dominating over polycomb-mediated gene silencing to 'lock' developmentally critical loci into an active chromatin state (H3K4me3 with induced histone acetylation), a state that defined leukaemia stem cells. Collectively, our studies represent, to our knowledge, the first report that deregulation of the PHD finger, an 'effector' of specific histone modification, perturbs the epigenetic dynamics on developmentally critical loci, catastrophizes cellular fate decision-making, and even causes oncogenesis during mammalian development.

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The PHD finger-containing NUP98-JARID1A fusion isoform (NJL), but not that lacking the PHD finger (NJS), confers leukomogenic potentials to hematopoietic stem/progenitor cellsa, NUP98-JARID1A and NUP98-PHF23 structure (see Supplementary Fig.1 for details). b, Immunoblot of hematopoietic cells transduced with empty vector (lanes 1-2) or that encoding FLAG-tagged NJS (lanes 3-4) or NJL (lanes 5-6). c, Proliferation kinetics of lineage-negative hematopoietic cells after transduction of empty vector, NJL or NJS. Data are presented as mean ±s.d. of 6 experiments. d, Wright-Giemsa staining (insert, microscopy image) and e, FACS of NJL-transformed cells. f, Leukemia kinetics in mice (12 each group) after transplantation of bone marrow transduced with vector, NJL or NLS. g, Hematoxylin-Eosin staining of spleen section and h, Wright-Giemsa staining of bone marrow from NJL-induced AML mice. Scale bar, 20μM.
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Figure 1: The PHD finger-containing NUP98-JARID1A fusion isoform (NJL), but not that lacking the PHD finger (NJS), confers leukomogenic potentials to hematopoietic stem/progenitor cellsa, NUP98-JARID1A and NUP98-PHF23 structure (see Supplementary Fig.1 for details). b, Immunoblot of hematopoietic cells transduced with empty vector (lanes 1-2) or that encoding FLAG-tagged NJS (lanes 3-4) or NJL (lanes 5-6). c, Proliferation kinetics of lineage-negative hematopoietic cells after transduction of empty vector, NJL or NJS. Data are presented as mean ±s.d. of 6 experiments. d, Wright-Giemsa staining (insert, microscopy image) and e, FACS of NJL-transformed cells. f, Leukemia kinetics in mice (12 each group) after transplantation of bone marrow transduced with vector, NJL or NLS. g, Hematoxylin-Eosin staining of spleen section and h, Wright-Giemsa staining of bone marrow from NJL-induced AML mice. Scale bar, 20μM.

Mentions: In clinically reported AML patients4,5, chromosomal translocation fuses the C-terminal PHD finger of JARID1A (also known as RBP2/KDM5A) or PHF23, together with nuclear localization signals, to NUP98, a common leukemia fusion partner that harbors transactivation activities15-17 (Supplementary Fig.1). Notably, the JARID1APHD3 motif is excluded from an alternatively spliced isoform of JARID1A and the corresponding NUP98-JARID1A fusion (hereafter referred to as NJS), while it is retained in the longer fusion isoform (hereafter referred to as NJL; Fig.1a). We asked whether JARID1APHD3 as a putative chromatin-‘reading’ module is involved in hematopoietic malignancies. To test this, we examined leukemogenic potential of both fusion isoforms using hematopoeitic progenitor transformation assay18 (Supplementary Fig.2a). While bone marrow-derived hematopoietic stem/progenitor cells transduced with empty retrovirus or retrovirus encoding NJS proliferated transiently and differentiated into mature cells, those transduced with NJL proliferated indefinitely as undifferentiated progenitors (Fig.1b-c). NJL-transduced marrow cells proliferated in a cell-autonomous manner, exhibited typical myeloblast morphology (Fig.1d) and expressed early myeloid progenitor antigens (c-Kit+/Cd11b+/Cd34+/Gr-1-/Cd19-/B220-/low; Fig.1e and Supplementary Fig. 2b). The arres of myeloid differentiation by NJL indicated that it would induce leukemia in vivo. Indeed, all of 12 mice transplanted with bone marrow progenitors transduced with NJL died of AML in an average of 69 days, whereas those reconstituted with vector- or NJS-transduced progenitors remained healthy after one year (Fig.1f). NJL-induced leukemia exhibited a myeloid phenotype (Supplementary Fig.2c-d), and typically presented with an enlarged spleen, packed progenitors in bone marrow, and massive increase in peripheral white blood cells (Supplementary Table 1; Fig.1g-h). Taken together, NJL represents a potent leukemia oncogene in both cellular and animal models.


Haematopoietic malignancies caused by dysregulation of a chromatin-binding PHD finger.

Wang GG, Song J, Wang Z, Dormann HL, Casadio F, Li H, Luo JL, Patel DJ, Allis CD - Nature (2009)

The PHD finger-containing NUP98-JARID1A fusion isoform (NJL), but not that lacking the PHD finger (NJS), confers leukomogenic potentials to hematopoietic stem/progenitor cellsa, NUP98-JARID1A and NUP98-PHF23 structure (see Supplementary Fig.1 for details). b, Immunoblot of hematopoietic cells transduced with empty vector (lanes 1-2) or that encoding FLAG-tagged NJS (lanes 3-4) or NJL (lanes 5-6). c, Proliferation kinetics of lineage-negative hematopoietic cells after transduction of empty vector, NJL or NJS. Data are presented as mean ±s.d. of 6 experiments. d, Wright-Giemsa staining (insert, microscopy image) and e, FACS of NJL-transformed cells. f, Leukemia kinetics in mice (12 each group) after transplantation of bone marrow transduced with vector, NJL or NLS. g, Hematoxylin-Eosin staining of spleen section and h, Wright-Giemsa staining of bone marrow from NJL-induced AML mice. Scale bar, 20μM.
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Figure 1: The PHD finger-containing NUP98-JARID1A fusion isoform (NJL), but not that lacking the PHD finger (NJS), confers leukomogenic potentials to hematopoietic stem/progenitor cellsa, NUP98-JARID1A and NUP98-PHF23 structure (see Supplementary Fig.1 for details). b, Immunoblot of hematopoietic cells transduced with empty vector (lanes 1-2) or that encoding FLAG-tagged NJS (lanes 3-4) or NJL (lanes 5-6). c, Proliferation kinetics of lineage-negative hematopoietic cells after transduction of empty vector, NJL or NJS. Data are presented as mean ±s.d. of 6 experiments. d, Wright-Giemsa staining (insert, microscopy image) and e, FACS of NJL-transformed cells. f, Leukemia kinetics in mice (12 each group) after transplantation of bone marrow transduced with vector, NJL or NLS. g, Hematoxylin-Eosin staining of spleen section and h, Wright-Giemsa staining of bone marrow from NJL-induced AML mice. Scale bar, 20μM.
Mentions: In clinically reported AML patients4,5, chromosomal translocation fuses the C-terminal PHD finger of JARID1A (also known as RBP2/KDM5A) or PHF23, together with nuclear localization signals, to NUP98, a common leukemia fusion partner that harbors transactivation activities15-17 (Supplementary Fig.1). Notably, the JARID1APHD3 motif is excluded from an alternatively spliced isoform of JARID1A and the corresponding NUP98-JARID1A fusion (hereafter referred to as NJS), while it is retained in the longer fusion isoform (hereafter referred to as NJL; Fig.1a). We asked whether JARID1APHD3 as a putative chromatin-‘reading’ module is involved in hematopoietic malignancies. To test this, we examined leukemogenic potential of both fusion isoforms using hematopoeitic progenitor transformation assay18 (Supplementary Fig.2a). While bone marrow-derived hematopoietic stem/progenitor cells transduced with empty retrovirus or retrovirus encoding NJS proliferated transiently and differentiated into mature cells, those transduced with NJL proliferated indefinitely as undifferentiated progenitors (Fig.1b-c). NJL-transduced marrow cells proliferated in a cell-autonomous manner, exhibited typical myeloblast morphology (Fig.1d) and expressed early myeloid progenitor antigens (c-Kit+/Cd11b+/Cd34+/Gr-1-/Cd19-/B220-/low; Fig.1e and Supplementary Fig. 2b). The arres of myeloid differentiation by NJL indicated that it would induce leukemia in vivo. Indeed, all of 12 mice transplanted with bone marrow progenitors transduced with NJL died of AML in an average of 69 days, whereas those reconstituted with vector- or NJS-transduced progenitors remained healthy after one year (Fig.1f). NJL-induced leukemia exhibited a myeloid phenotype (Supplementary Fig.2c-d), and typically presented with an enlarged spleen, packed progenitors in bone marrow, and massive increase in peripheral white blood cells (Supplementary Table 1; Fig.1g-h). Taken together, NJL represents a potent leukemia oncogene in both cellular and animal models.

Bottom Line: In these processes, a PHD finger that specifically recognizes H3K4me3/2 marks was essential for leukaemogenesis.Mutations in PHD fingers that abrogated H3K4me3 binding also abolished leukaemic transformation.Collectively, our studies represent, to our knowledge, the first report that deregulation of the PHD finger, an 'effector' of specific histone modification, perturbs the epigenetic dynamics on developmentally critical loci, catastrophizes cellular fate decision-making, and even causes oncogenesis during mammalian development.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Chromatin Biology & Epigenetics, The Rockefeller University, New York, New York 10065, USA.

ABSTRACT
Histone H3 lysine 4 methylation (H3K4me) has been proposed as a critical component in regulating gene expression, epigenetic states, and cellular identities1. The biological meaning of H3K4me is interpreted by conserved modules including plant homeodomain (PHD) fingers that recognize varied H3K4me states. The dysregulation of PHD fingers has been implicated in several human diseases, including cancers and immune or neurological disorders. Here we report that fusing an H3K4-trimethylation (H3K4me3)-binding PHD finger, such as the carboxy-terminal PHD finger of PHF23 or JARID1A (also known as KDM5A or RBBP2), to a common fusion partner nucleoporin-98 (NUP98) as identified in human leukaemias, generated potent oncoproteins that arrested haematopoietic differentiation and induced acute myeloid leukaemia in murine models. In these processes, a PHD finger that specifically recognizes H3K4me3/2 marks was essential for leukaemogenesis. Mutations in PHD fingers that abrogated H3K4me3 binding also abolished leukaemic transformation. NUP98-PHD fusion prevented the differentiation-associated removal of H3K4me3 at many loci encoding lineage-specific transcription factors (Hox(s), Gata3, Meis1, Eya1 and Pbx1), and enforced their active gene transcription in murine haematopoietic stem/progenitor cells. Mechanistically, NUP98-PHD fusions act as 'chromatin boundary factors', dominating over polycomb-mediated gene silencing to 'lock' developmentally critical loci into an active chromatin state (H3K4me3 with induced histone acetylation), a state that defined leukaemia stem cells. Collectively, our studies represent, to our knowledge, the first report that deregulation of the PHD finger, an 'effector' of specific histone modification, perturbs the epigenetic dynamics on developmentally critical loci, catastrophizes cellular fate decision-making, and even causes oncogenesis during mammalian development.

Show MeSH
Related in: MedlinePlus