Dynamic Reprogramming of DNA Methylation at an Epigenetically Sensitive Allele in Mice

There is increasing evidence in both plants and animals that epigenetic marks are not always cleared between generations. Incomplete erasure at genes associated with a measurable phenotype results in unusual patterns of inheritance from one generation to the next, termed transgenerational epigenetic inheritance. The Agouti viable yellow (A(vy)) allele is the best-studied example of this phenomenon in mice. The A(vy) allele is the result of a retrotransposon insertion upstream of the Agouti gene. Expression at this locus is controlled by the long terminal repeat (LTR) of the retrotransposon, and expression results in a yellow coat and correlates with hypomethylation of the LTR. Isogenic mice display variable expressivity, resulting in mice with a range of coat colours, from yellow through to agouti. Agouti mice have a methylated LTR. The locus displays epigenetic inheritance following maternal but not paternal transmission; yellow mothers produce more yellow offspring than agouti mothers. We have analysed the DNA methylation in mature gametes, zygotes, and blastocysts and found that the paternally and maternally inherited alleles are treated differently. The paternally inherited allele is demethylated rapidly, and the maternal allele is demethylated more slowly, in a manner similar to that of nonimprinted single-copy genes. Interestingly, following maternal transmission of the allele, there is no DNA methylation in the blastocyst, suggesting that DNA methylation is not the inherited mark. We have independent support for this conclusion from studies that do not involve direct analysis of DNA methylation. Haplo-insufficiency for Mel18, a polycomb group protein, introduces epigenetic inheritance at a paternally derived A(vy) allele, and the pedigrees reveal that this occurs after zygotic genome activation and, therefore, despite the rapid demethylation of the locus

Bone Marrow Allograft Rejection Mediated by a Novel Murine NK Receptor, NKG2I

Natural killer (NK) cells mediate bone marrow allograft rejection. However, the molecular mechanisms underlying such a rejection remain elusive. In previous analyses, it has been shown that NK cells recognize allogeneic target cells through Ly-49s and CD94/NKG2 heterodimers. Here, we describe identification and characterization of a novel murine NK receptor, NKG2I, belonging to the NKG2 family. NKG2I, which was composed of 226 amino acids, showed ∼40% homology to the murine NKG2D and CD94 in the C-type lectin domain. Flow cytometric analysis with anti-NKG2I monoclonal antibody (mAb) revealed that expression of NKG2I was largely confined to NK and NKT cells, but was not seen in T cells. Furthermore, anti-NKG2I mAb inhibited NK cell–mediated cytotoxicity, whereas cross-linking of NKG2I enhanced interleukin 2– and interleukin 12–dependent interferon-γ production. Similarly, the injection of anti-NKG2I mAb before the allogeneic bone marrow transfer in vivo impinged on the function of NKG2I, resulting in the enhanced colony formation in the spleen. NKG2I is a novel activating receptor mediating recognition and rejection of allogeneic target cells

Maintenance of Undifferentiated State and Self-Renewal of Embryonic Neural Stem Cells by Polycomb Protein Ring1B

12 pages, 7 figures.-- PMID: 19544461 [PubMed].-- Printed version published Jul 2009.Supporting information (Suppl. figures S1-S9, tables S1-S2) available at: http://www3.interscience.wiley.com/journal/122302104/suppinfoCell lineages generated during development and tissue maintenance are derived from self-renewing stem cells by differentiation of their committed progeny. Recent studies suggest that epigenetic mechanisms, and in particular the Polycomb group (PcG) of genes, play important roles in controlling stem cell self-renewal. Here, we address PcG regulation of stem cell self-renewal and differentiation through inactivation of Ring1B, a histone H2A E3 monoubiquitin ligase, in embryonic neural stem cells (NSCs) from the olfactory bulb of a conditional mouse mutant line. We show that neural stem/progenitor cell proliferation in vivo and in neurosphere assays is impaired, lacking Ring1B, and their self-renewal and multipotential abilities, assessed as sphere formation and differentiation from single cells, are severely affected. We also observed unscheduled neuronal, but not glial, differentiation of mutant stem/progenitor cells under proliferating conditions, an alteration enhanced in cells also lacking Ring1A, the Ring1B paralog, some of which turned into morphologically identifiable neurons. mRNA analysis of mutant cells showed upregulation of some neuronal differentiation-related transcription factors and the cell proliferation inhibitor Cdkn1a/p21, as well as downregulation of effectors of the Notch signaling pathway, a known inhibitor of neuronal differentiation of stem/progenitor cells. In addition, differentiation studies of Ring1B-deficient progenitors showed decreased oligodendrocyte formation in vitro and enhanced neurogenesis and reduced gliogenesis in vivo. These data suggest a role for Ring1B in maintenance of the undifferentiated state of embryonic neural stem/progenitor cells. They also suggest that Ring1B may modulate the differentiation potential of NSCs to neurons and glia.M.R-T. and H. M-G. were recipients of FPU and FPI fellowships, from the Ministerio de Educacion y Ciencia and Comunidad de Madrid, respectively. This work was supported by grants SAF2007-65957-C02-01 (M.V.), the Onco-Cycle program from the Comunidad de Madrid (M.V.), SAF2004-05798, and CIBERNED CB06/05/0065 from Instituto de Salud Carlos III (C.V-A.).Peer reviewe

Variant PRC1 competes with retinoic acid-related signals to repress Meis2 in the mouse distal forelimb bud

10 p.-3 fig.-1 tab.Suppression of Meis genes in the distal limb bud is required for proximal-distal (PD) specification of the forelimb. Polycomb group (PcG) factors play a role in downregulation of retinoic acid (RA)-related signals in the distal forelimb bud, causing Meis repression. It is, however, not known whether downregulation of RA-related signals and PcG-mediated proximal gene repression are functionally linked. Here, we reveal that PcG factors and RA-related signals antagonize each other to polarize Meis2 expression along the PD axis in mouse. Supported by mathematical modeling and simulation, we propose that PcG factors are required to adjust the threshold for RA-related signaling to regulate Meis2 expression. Finally, we show that a variant Polycomb repressive complex 1 (PRC1), incorporating PCGF3 and PCGF5, represses Meis2 expression in the distal limb bud. Taken together, we reveal a previously unknown link between PcG proteins and downregulation of RA-related signals to mediate the phase transition of Meis2 transcriptional status during forelimb patterning.This work was supported by Grants-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology of Japan (MEXT) (23249015 to H.K.), the Japan Agency for Medical Research and Development (AMED) (JP18gm0510016 to H.K. and T.K.), the Special Postdoctoral Researcher Program of RIKEN (to N.Y.-K.), the Regional Innovation Program from MEXT (to T.K.) and the Cross-ministerial Strategic Innovation Promotion Program (SIP) from Cabinet Office, Government of Japan (to T.K. and H.K.).Peer reviewe

Human NK cell development in hIL-7 and hIL-15 knockin NOD/SCID/IL2rgKO mice.

The immune system encompasses acquired and innate immunity that matures through interaction with microenvironmental components. Cytokines serve as environmental factors that foster functional maturation of immune cells. Although NOD/SCID/IL2rgKO (NSG) humanized mice support investigation of human immunity in vivo, a species barrier between human immune cells and the mouse microenvironment limits human acquired as well as innate immune function. To study the roles of human cytokines in human acquired and innate immune cell development, we created NSG mice expressing hIL-7 and hIL-15. Although hIL-7 alone was not sufficient for supporting human NK cell development in vivo, increased frequencies of human NK cells were confirmed in multiple organs of hIL-7 and hIL-15 double knockin (hIL-7xhIL-15 KI) NSG mice engrafted with human hematopoietic stem cells. hIL-7xhIL-15 KI NSG humanized mice provide a valuable in vivo model to investigate development and function of human NK cells

PRC1-mediated epigenetic programming is required to generate the ovarian reserve

10 p.-4 fig.The ovarian reserve defines the female reproductive lifespan, which in humans spans decades due to robust maintenance of meiotic arrest in oocytes residing in primordial follicles. Epigenetic reprogramming, including DNA demethylation, accompanies meiotic entry, but the chromatin changes that underpin the generation and preservation of ovarian reserves are poorly defined. We report that the Polycomb Repressive Complex 1 (PRC1) establishes repressive chromatin states in perinatal mouse oocytes that directly suppress the gene expression program of meiotic prophase-I and thereby enable the transition to dictyate arrest. PRC1 dysfuction causes depletion of the ovarian reserve and leads to premature ovarian failure. Our study demonstrates a fundamental role for PRC1-mediated gene silencing in female reproductive lifespan, and reveals a critical window of epigenetic programming required to establish ovarian reserve.Funding sources: National Institutes of Health grants R01GM122776 and R35GM141085 to S.H.N.Peer reviewe

Co-activation of macrophages and T cells contribute to chronic GVHD in human IL-6 transgenic humanised mouse model.

BACKGROUND: Graft-versus host disease (GVHD) is a complication of stem cell transplantation associated with significant morbidity and mortality. Non-specific immune-suppression, the mainstay of treatment, may result in immune-surveillance dysfunction and disease recurrence. METHODS: We created humanised mice model for chronic GVHD (cGVHD) by injecting cord blood (CB)-derived human CD34 FINDINGS: In cGVHD humanised mice, we found activation of T cells in the spleen, lung, liver, and skin, activation of macrophages in lung and liver, and loss of appendages in skin, obstruction of bronchioles in lung and portal fibrosis in liver recapitulating cGVHD. Acute GVHD humanised mice showed activation of T cells with skewed TCR repertoire without significant macrophage activation. INTERPRETATION: Using humanised mouse models, we demonstrated distinct immune mechanisms contributing acute and chronic GVHD. In cGVHD model, co-activation of human HSPC-derived macrophages and T cells educated in the recipient thymus contributed to delayed onset, multi-organ disease. In acute GVHD model, mature human T cells contained in the graft resulted in rapid disease progression. These humanised mouse models may facilitate future development of new molecular medicine targeting GVHD

DJ-1 depletion slows down immunoaging in T-cell compartments

Decline in immune function during aging increases susceptibility to different aging related diseases. However, the underlying molecular mechanisms, especially the genetic factors contributing to imbalance of naïve/memory T-cell subpopulations, still remain largely elusive. Here we show that loss of DJ-1 encoded by PARK7 /DJ-1, causing early-onset familial Parkinson’s disease (PD), unexpectedly delayed immunoaging in both human and mice. Compared with two gender-matched unaffected sibling carriers of similar ages, the index PD patient with DJ-1 deficiency showed a decline in many critical immunoaging features, including almost doubled frequencies of non-senescent T cells. The observation of a ‘younger’ immune system in the index patient was further consolidated by the results in aged DJ-1 knockout mice. Our data from bone marrow chimera models and adoptive transfer experiments demonstrated that DJ-1 regulates several immunoaging features via hematopoietic-intrinsic and naïve-CD8-intrinsic mechanisms. Our finding suggests an unrecognized critical role of DJ-1 in regulating immunoaging, discovering a potent target to interfere with immunoaging- and aging-associated diseases