G9a/GLP-dependent histone H3K9me2 patterning during human hematopoietic stem cell lineage commitment

Molecular and Cell Biology (MCB) Program.
Genes & development (Impact Factor: 10.8). 10/2012; 26(22). DOI: 10.1101/gad.200329.112
Source: PubMed


G9a and GLP are conserved protein methyltransferases that play key roles during mammalian development through mono- and dimethylation of histone H3 Lys 9 (H3K9me1/2), modifications associated with transcriptional repression. During embryogenesis, large H3K9me2 chromatin territories arise that have been proposed to reinforce lineage choice by affecting high-order chromatin structure. Here we report that in adult human hematopoietic stem and progenitor cells (HSPCs), H3K9me2 chromatin territories are absent in primitive cells and are formed de novo during lineage commitment. In committed HSPCs, G9a/GLP activity nucleates H3K9me2 marks at CpG islands and other genomic sites within genic regions, which then spread across most genic regions during differentiation. Immunofluorescence assays revealed the emergence of H3K9me2 nuclear speckles in committed HSPCs, consistent with progressive marking. Moreover, gene expression analysis indicated that G9a/GLP activity suppresses promiscuous transcription of lineage-affiliated genes and certain gene clusters, suggestive of regulation of HSPC chromatin structure. Remarkably, HSPCs continuously treated with UNC0638, a G9a/GLP small molecular inhibitor, better retain stem cell-like phenotypes and function during in vitro expansion. These results suggest that G9a/GLP activity promotes progressive H3K9me2 patterning during HSPC lineage specification and that its inhibition delays HSPC lineage commitment. They also inform clinical manipulation of donor-derived HSPCs.

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Available from: Beverly Torok-Storb, Oct 01, 2015
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    • "Our previous results revealed that H3K9me2 patterning progresses through distinct stages during HSC differentiation. First, H3K9me2 marks appear low or absent in HSC-enriched CD34+CD90+CD38loCD45RA– cells [6]. Next, in CD34+ HSPCs (composed mainly of committed progenitors), a nucleation stage ensues in which H3K9me2 marks appear at discrete loci across the genome [6]. "
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    ABSTRACT: Background The formation of chromatin domains is an important step in lineage commitment. In human hematopoietic stem and progenitor cells (HSPCs), G9a/GLP-dependent H3K9me2 chromatin territories form de novo during lineage specification and are nucleated at punctate sites during lineage commitment. Here, we examined the patterning of G9a/GLP-dependent H3K9me2 in HSPCs and the consequences for chromatin structure. Results We profiled chromatin accessibility across the genome of HSPCs treated with either a small molecule inhibitor of G9a/GLP or DMSO. We observed that chromatin accessibility is dramatically altered at the regions of H3K9me2 nucleation. We have characterized the regions of H3K9me2 nucleation, with our analysis revealing that H3K9me2 is nucleated in HSPCs at CpG islands (CGIs) and CGI-like sequences across the genome. Our analysis furthermore revealed a bias of H3K9me2 nucleation towards regions with low rates of C- > T deamination, which typically lack DNA methylation. Lastly, we examined the interaction of H3K9me2 and DNA methylation and determined that chromatin accessibility changes upon loss of H3K9me2 are dependent on the presence of DNA methylation. Conclusions These results indicate that H3K9me2 nucleation is established at specific sequences that have base composition similar to CGIs. Our results furthermore indicate that H3K9me2 nucleation leads to local changes in chromatin accessibility and that H3K9me2 and DNA methylation work synergistically to regulate chromatin accessibility.
    Epigenetics & Chromatin 09/2014; 7(1):23. DOI:10.1186/1756-8935-7-23 · 5.33 Impact Factor
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    • "We found that in PF mice, the reduction of IGF-1/pAkt and PKA/pCREB signaling was associated with an induction of Foxo1 expression and a reduction of G9a (Figures 6B and S5E), but it did not affect the expressions of Foxo3a and Foxo4 (Figures S5F–S5H). Also, the results indicated that the numbers of ST-HSC and MPP were significantly increased after treatment with PKA small interfering RNA (siRNA) as well as after treatment with IGF-1 siRNA (Figures 6C and S5D; Table S5), in agreement with the finding that inhibition of G9a increases primitive HSCs (Chen et al., 2012). Given that inhibition of mTOR, another key effector of nutrient signaling, is known to enhance HSC self-renewal and maintenance autonomously and nonautonomously, we examined the crosstalk between mTOR and PKA in HSCs and MPPs (Chen et al., 2009; Huang et al., 2012). "
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    ABSTRACT: Immune system defects are at the center of aging and a range of diseases. Here, we show that prolonged fasting reduces circulating IGF-1 levels and PKA activity in various cell populations, leading to signal transduction changes in long-term hematopoietic stem cells (LT-HSCs) and niche cells that promote stress resistance, self-renewal, and lineage-balanced regeneration. Multiple cycles of fasting abated the immunosuppression and mortality caused by chemotherapy and reversed age-dependent myeloid-bias in mice, in agreement with preliminary data on the protection of lymphocytes from chemotoxicity in fasting patients. The proregenerative effects of fasting on stem cells were recapitulated by deficiencies in either IGF-1 or PKA and blunted by exogenous IGF-1. These findings link the reduced levels of IGF-1 caused by fasting to PKA signaling and establish their crucial role in regulating hematopoietic stem cell protection, self-renewal, and regeneration.
    Cell Stem Cell 06/2014; 14(6):810-823. DOI:10.1016/j.stem.2014.04.014 · 22.27 Impact Factor
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    • "This has been demonstrated in certain contexts, as G9a mediates T-helper cell diversification (Lehnertz et al. 2010) and embryonic stem cell (ESC) differentiation (Feldman et al. 2006). Furthermore, G9A/GLP inhibition delays the differentiation of human hematopoietic stem cells (HSCs) ex vivo (Chen et al. 2012), suggesting additional roles in early hematopoiesis. Despite recent ad- Ó 2014 Lehnertz et al. "
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    ABSTRACT: Chromatin modulators are emerging as attractive drug targets, given their widespread implication in human cancers and susceptibility to pharmacological inhibition. Here we establish the histone methyltransferase G9a/EHMT2 as a selective regulator of fast proliferating myeloid progenitors with no discernible function in hematopoietic stem cells (HSCs). In mouse models of acute myeloid leukemia (AML), loss of G9a significantly delays disease progression and reduces leukemia stem cell (LSC) frequency. We connect this function of G9a to its methyltransferase activity and its interaction with the leukemogenic transcription factor HoxA9 and provide evidence that primary human AML cells are sensitive to G9A inhibition. Our results highlight a clinical potential of G9A inhibition as a means to counteract the proliferation and self-renewal of AML cells by attenuating HoxA9-dependent transcription.
    Genes & development 02/2014; 28(4):317-27. DOI:10.1101/gad.236794.113 · 10.80 Impact Factor
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