Publications from Epigenetics

For a full list (GEO|SRA|Browser|Code) see below

  1. PROSER1 Mediates TET2 O-GlcNAcylation to Regulate DNA Demethylation on UTX-Dependent Enhancers and CpG Islands

    DNA methylation at enhancers and CpG islands usually leads to gene repression, which is counteracted by DNA demethylation through the TET protein family. However, how TET enzymes are recruited and regulated at these genomic loci is not fully understood. Furthermore, we provide evidence that PROSER1 acts as a more general regulator of OGT activity by controlling O-GlcNAcylation of multiple other chromatin signaling pathways. Taken together, this study describes for the first time a regulator of TET2 O-GlcNAcylation and its implications in mediating DNA demethylation at UTX-dependent enhancers and CpG islands and supports an important role for PROSER1 in regulating the function of various chromatin-associated proteins via OGT-mediated O-GlcNAcylation

    Wang Xiaokang*, Rosikiewicz Wojciech*, Sedkov Yurii*, Martinez Tanner, Hansen Baranda S., Schreiner Patrick, Christensen Jesper, Xu Beisi, Pruett-Miller Shondra M., Helin Kristian, Herz Hans-Martin

  2. Acute Depletion of CTCF Rewires Genome-Wide Chromatin Accessibility

    Background The transcription factor CTCF appears indispensable in defining topologically associated domain boundaries and maintaining chromatin loop structures within these domains, supported by numerous functional studies. However, acute depletion of CTCF globally reduces chromatin interactions but does not significantly alter transcription. Interestingly, many CTCF co-regulators that have alterations of their respective downstream gene expression do not show changes of their own expression levels across the multi-omics measurements upon acute CTCF loss, highlighting the strength of our system to discover hidden co-regulatory partners associated with CTCF-mediated transcription. Conclusions This study highlights that CTCF loss rewires genome-wide chromatin accessibility, which plays a critical role in transcriptional regulation

    Xu Beisi*#, Wang Hong*, Wright Shaela, Hyle Judith, Zhang Yang, Shao Ying, Niu Mingming, Fan Yiping, Rosikiewicz Wojciech, Djekidel Mohamed Nadhir, Peng Junmin, Lu Rui, Li Chunliang#

  3. Ybx1 Fine-Tunes PRC2 Activities to Control Embryonic Brain Development

    Chromatin modifiers affect spatiotemporal gene expression programs that underlie organismal development. The Polycomb repressive complex 2 (PRC2) is a crucial chromatin modifier in executing neurodevelopmental programs. In Ybx1-knockout NPCs, H3K27me3 reduction by PRC2 enzymatic inhibitor or genetic depletion partially rescues gene expression and NPC functions. Our findings suggest that Ybx1 fine-tunes PRC2 activities to regulate spatiotemporal gene expression in embryonic neural development and uncover a crucial epigenetic mechanism balancing forebrain–hindbrain lineages and self-renewal–differentiation choices in NPCs

    Evans Myron K., Matsui Yurika*, Xu Beisi*, Willis Catherine, Loome Jennifer, Milburn Luis, Fan Yiping, Pagala Vishwajeeth, Peng Jamy C.

  4. Targeting REGNASE-1 Programs Long-Lived Effector T Cells for Cancer Therapy

    Adoptive cell therapy represents a new paradigm in cancer immunotherapy, but it can be limited by the poor persistence and function of transferred T cells1. Here we use an in vivo pooled CRISPR–Cas9 mutagenesis screening approach to demonstrate that, by targeting REGNASE-1, CD8+ T cells are reprogrammed to long-lived effector cells with extensive accumulation, better persistence and robust effector function in tumours. By contrast, the targeting of additional signalling factors—including PTPN2 and SOCS1—improves the therapeutic efficacy of REGNASE-1-deficient CD8+ T cells. Our findings suggest that T cell persistence and effector function can be coordinated in tumour immunity and point to avenues for improving the efficacy of adoptive cell therapy for cancer

    Wei Jun, Long Lingyun, Zheng Wenting, Dhungana Yogesh, Lim Seon Ah, Guy Cliff, Wang Yanyan, Wang Yong-Dong, Qian Chenxi, Xu Beisi, Kc Anil, Saravia Jordy, Huang Hongling, Yu Jiyang, Doench John G., Geiger Terrence L., Chi Hongbo

  5. Nucleome Dynamics during Retinal Development

    Summary More than 8,000 genes are turned on or off as progenitor cells produce the 7 classes of retinal cell types during development. Thousands of enhancers are also active in the developing retinae, many having features of cell- and developmental stage-specific activity. Single-cell ATAC-seq and RNA-seq were integrated with our Hi-C and previous ChIP-seq data to identify cell- and developmental-stage-specific super-enhancers (SEs). We identified a bipolar neuron-specific core regulatory circuit SE upstream of Vsx2, whose deletion in mice led to the loss of bipolar neurons

    Norrie Jackie L.*, Lupo Marybeth S.*, Xu Beisi*, Al Diri Issam*, Valentine Marc*, Putnam Daniel*, Griffiths Lyra, Zhang Jiakun, Johnson Dianna, Easton John, Shao Ying, Honnell Victoria, Frase Sharon, Miller Shondra, Stewart Valerie, Zhou Xin, Chen Xiang, Dyer Michael A.

  6. Acute Depletion of CTCF Directly Affects MYC Regulation through Loss of Enhancer–Promoter Looping

    Numerous pieces of evidence support the complex, 3D spatial organization of the genome dictates gene expression. CTCF is essential to define topologically associated domain boundaries and to facilitate the formation of insulated chromatin loop structures. 8 Mb downstream. Notably, MYC expression was not profoundly affected upon CTCF loss in HUDEP-2 cells suggesting that CTCF could play a B-ALL cell line specific role in maintaining MYC expression

    Hyle Judith, Zhang Yang, Wright Shaela, Xu Beisi, Shao Ying, Easton John, Tian Liqing, Feng Ruopeng, Xu Peng, Li Chunliang

  7. Differentiation of Human Pluripotent Stem Cells into Neurons or Cortical Organoids Requires Transcriptional Co-Regulation by UTX and 53BP1

    UTX is a chromatin modifier required for development and neural lineage specification, but how it controls these biological processes is unclear. To determine the molecular mechanisms of UTX, we identified novel UTX protein interaction partners. 53BP1 promotes UTX chromatin binding, and in turn H3K27 modifications and gene activation, at a subset of genomic regions, including neurogenic genes. Overall, our data suggest that the 53BP1–UTX interaction supports the activation of key genes required for human neurodevelopment

    Yang Xiaoyang*, Xu Beisi*, Mulvey Brett, Evans Myron, Jordan Samuel, Wang Yong-Dong, Pagala Vishwajeeth, Peng Junmin, Fan Yiping, Patel Arishna, Peng Jamy C.

  8. Metabolic Heterogeneity Underlies Reciprocal Fates of T H 17 Cell Stemness and Plasticity

    A defining feature of adaptive immunity is the development of long-lived memory T cells to curtail infection. Recent studies have identified a unique stem-like T-cell subset amongst exhausted CD8-positive T cells in chronic infection1–3, but it remains unclear whether CD4-positive T-cell subsets with similar features exist in chronic inflammatory conditions. Single-cell RNA sequencing and experimental validation reveal heterogeneity in fate-mapped TH17 cells, and a developmental arrest in the TH1 transdifferentiation trajectory upon loss of mTORC1 activity or metabolic perturbation. Our results establish that the dichotomy of stemness and effector function underlies the heterogeneous TH17 responses and autoimmune pathogenesis, and point to previously unappreciated metabolic control of plasticity in helper T cells

    Karmaus Peer W. F., Chen Xiang, Lim Seon Ah, Herrada Andrés A., Nguyen Thanh-Long M., Xu Beisi, Dhungana Yogesh, Rankin Sherri, Chen Wenan, Rosencrance Celeste, Yang Kai, Fan Yiping, Cheng Yong, Easton John, Neale Geoffrey, Vogel Peter, Chi Hongbo

  9. Retinal Cell Type DNA Methylation and Histone Modifications Predict Reprogramming Efficiency and Retinogenesis in 3D Organoid Cultures

    Summary Diverse cell types can be reprogrammed into pluripotent stem cells by ectopic expression of Oct4 (Pou5f1), Klf4, Sox2, and Myc. Many of these induced pluripotent stem cells (iPSCs) retain memory, in terms of DNA methylation and histone modifications (epigenetic memory), of their cellular origins, and this may bias subsequent differentiation. Neurons with the lowest reprogramming efficiency produced iPSC lines with the best retinal differentiation and were more likely to retain epigenetic memory of their cellular origins. In addition, we identified biomarkers of iPSCs that are predictive of retinal differentiation

    Wang Lu*, Hiler Daniel*, Xu Beisi*, AlDiri Issam, Chen Xiang, Zhou Xin, Griffiths Lyra, Valentine Marc, Shirinifard Abbas, Sablauer András, Thiagarajan Suresh, Barabas Marie-Elizabeth, Zhang Jiakun, Johnson Dianna, Frase Sharon, Dyer Michael A.

  10. The Dynamic Epigenetic Landscape of the Retina During Development, Reprogramming, and Tumorigenesis

    In the developing retina, multipotent neural progenitors undergo unidirectional differentiation in a precise spatiotemporal order. Here we profile the epigenetic and transcriptional changes that occur during retinogenesis in mice and humans. The retinoblastoma epigenome mapped to the developmental stage when retinal progenitors switch from neurogenic to terminal patterns of cell division. The epigenome of retinoblastomas was more similar to that of the normal retina than that of retina-derived iPSCs, and we identified retina-specific epigenetic memory

    Aldiri Issam*, Xu Beisi*, Wang Lu, Chen Xiang, Hiler Daniel, Griffiths Lyra, Valentine Marc, Shirinifard Abbas, Thiagarajan Suresh, Sablauer Andras, Barabas Marie-Elizabeth, Zhang Jiakun, Johnson Dianna, Frase Sharon, Zhou Xin, Easton John, Zhang Jinghui, Mardis Elaine R., Wilson Richard K., Downing James R., Dyer Michael A.

 

Full List

*denotes equal contribution, #denotes corresponding

2022:1

  1. PROSER1 Mediates TET2 O-GlcNAcylation to Regulate DNA Demethylation on UTX-Dependent Enhancers and CpG Islands
    Wang Xiaokang*, Rosikiewicz Wojciech*, Sedkov Yurii*, Martinez Tanner, Hansen Baranda S., Schreiner Patrick, Christensen Jesper, Xu Beisi, Pruett-Miller Shondra M., Helin Kristian, Herz Hans-Martin
    Life Science Alliance 2022

    DNA methylation at enhancers and CpG islands usually leads to gene repression, which is counteracted by DNA demethylation through the TET protein family. However, how TET enzymes are recruited and regulated at these genomic loci is not fully understood. Here, we identify TET2, the glycosyltransferase OGT and a previously undescribed proline and serine rich protein, PROSER1 as interactors of UTX, a component of the enhancer-associated MLL3/4 complexes. We find that PROSER1 mediates the interaction between OGT and TET2, thus promoting TET2 O-GlcNAcylation and protein stability. In addition, PROSER1, UTX, TET1/2, and OGT colocalize on many genomic elements genome-wide. Loss of PROSER1 results in lower enrichment of UTX, TET1/2, and OGT at enhancers and CpG islands, with a concomitant increase in DNA methylation and transcriptional down-regulation of associated target genes and increased DNA hypermethylation encroachment at H3K4me1-predisposed CpG islands. Furthermore, we provide evidence that PROSER1 acts as a more general regulator of OGT activity by controlling O-GlcNAcylation of multiple other chromatin signaling pathways. Taken together, this study describes for the first time a regulator of TET2 O-GlcNAcylation and its implications in mediating DNA demethylation at UTX-dependent enhancers and CpG islands and supports an important role for PROSER1 in regulating the function of various chromatin-associated proteins via OGT-mediated O-GlcNAcylation.

2021:8

  1. Acute Depletion of CTCF Rewires Genome-Wide Chromatin Accessibility
    Xu Beisi*#, Wang Hong*, Wright Shaela, Hyle Judith, Zhang Yang, Shao Ying, Niu Mingming, Fan Yiping, Rosikiewicz Wojciech, Djekidel Mohamed Nadhir, Peng Junmin, Lu Rui, Li Chunliang#
    Genome Biol. 2021

    Background The transcription factor CTCF appears indispensable in defining topologically associated domain boundaries and maintaining chromatin loop structures within these domains, supported by numerous functional studies. However, acute depletion of CTCF globally reduces chromatin interactions but does not significantly alter transcription. Results Here, we systematically integrate multi-omics data including ATAC-seq, RNA-seq, WGBS, Hi-C, Cut&Run, and CRISPR-Cas9 survival dropout screens, and time-solved deep proteomic and phosphoproteomic analyses in cells carrying auxin-induced degron at endogenous CTCF locus. Acute CTCF protein degradation markedly rewires genome-wide chromatin accessibility. Increased accessible chromatin regions are frequently located adjacent to CTCF-binding sites at promoter regions and insulator sites associated with enhanced transcription of nearby genes. In addition, we use CTCF-associated multi-omics data to establish a combinatorial data analysis pipeline to discover CTCF co-regulatory partners. We successfully identify 40 candidates, including multiple established partners. Interestingly, many CTCF co-regulators that have alterations of their respective downstream gene expression do not show changes of their own expression levels across the multi-omics measurements upon acute CTCF loss, highlighting the strength of our system to discover hidden co-regulatory partners associated with CTCF-mediated transcription. Conclusions This study highlights that CTCF loss rewires genome-wide chromatin accessibility, which plays a critical role in transcriptional regulation.

  2. Enhancer Hijacking Drives Oncogenic BCL11B Expression in Lineage Ambiguous Stem Cell Leukemia
    Montefiori Lindsey E., Bendig Sonja, Gu Zhaohui, Chen Xiaolong, Polonen Petri, Ma Xiaotu, Murison Alex, Zeng Andy, Garcia-Prat Laura, Dickerson Kirsten, Iacobucci Ilaria, Abdelhamed Sherif, Hiltenbrand Ryan, Mead Paul E., Mehr Cyrus M., Xu Beisi, Cheng Zhongshan, Chang Ti-Cheng, Westover Tamara, Ma Jing, Stengel Anna, Kimura Shunsuke, Qu Chunxu, Valentine Marcus B., Rashkovan Marissa, Luger Selina, Litzow Mark R., Rowe Jacob M., Boer Monique L., Wang Victoria, Yin Jun, Kornblau Steven M., Hunger Stephen P., Loh Mignon L., Pui Ching-Hon, Yang Wenjian, Crews Kristine R., Roberts Kathryn G., Yang Jun J., Relling Mary V., Evans William E., Stock Wendy, Paietta Elisabeth M., Ferrando Adolfo A., Zhang Jinghui, Kern Wolfgang, Haferlach Torsten, Wu Gang, Dick John E., Klco Jeffery M., Haferlach Claudia, Mullighan Charles G.
    Cancer Discovery 2021

    Lineage ambiguous leukemias are high-risk malignancies of poorly understood genetic basis. Here, we describe a distinct subgroup of acute leukemia with expression of myeloid, T lymphoid and stem cell markers driven by aberrant allele-specific deregulation of BCL11B, a master transcription factor responsible for thymic T-lineage commitment and specification. Mechanistically, this deregulation was driven by chromosomal rearrangements that juxtapose BCL11B to super-enhancers active in hematopoietic progenitors, or focal amplifications that generate a super-enhancer from a non-coding element distal to BCL11B. Chromatin conformation analyses demonstrate long range interactions of rearranged enhancers with the expressed BCL11B allele, and association of BCL11B with activated hematopoietic progenitor cell cis-regulatory elements, suggesting BCL11B is aberrantly co-opted into a gene regulatory network that drives transformation by maintaining a progenitor state. These data support a role for ectopic BCL11B expression in primitive hematopoietic cells mediated by enhancer hijacking as an oncogenic driver of human lineage ambiguous leukemia.

  3. Foxp3 Enhancers Synergize to Maximize Regulatory T Cell Suppressive Capacity
    Zong Xinying, Hao Xiaolei, Xu Beisi, Crawford Jeremy Chase, Wright Shaela, Li Jun, Zhang Yang, Bai Lu, He Minghong, Jiang Menglin, Fan Yiping, Connelly Jon P., Pruett-Miller Shondra M., Berns Hartmut, Janke Laura, Li Chunliang, Feng Yongqiang
    Journal of Experimental Medicine 2021

    T reg cells bearing a diverse antigen receptor repertoire suppress pathogenic T cells and maintain immune homeostasis during their long lifespan. How their robust function is determined genetically remains elusive. Here, we investigate the regulatory space of the cis-regulatory elements of T reg lineage–specifying factor Foxp3. Foxp3 enhancers are known as distinct readers of environmental cues controlling T reg cell induction or lineage stability. However, their single deficiencies cause mild, if any, immune dysregulation, leaving the key transcriptional mechanisms determining Foxp3 expression and thereby T reg cell suppressive capacity uncertain. We examined the collective activities of Foxp3 enhancers and found that they coordinate to maximize T reg cell induction, Foxp3 expression level, or lineage stability through distinct modes and that ablation of synergistic enhancers leads to lethal autoimmunity in young mice. Thus, the induction and maintenance of a diverse, stable T reg cell repertoire rely on combinatorial Foxp3 enhancers, suggesting broad, stage-specific, synergistic activities of cell-intrinsic factors and cell-extrinsic cues in determining T reg cell suppressive capacity.

  4. A Distal Foxp3 Enhancer Enables Interleukin-2 Dependent Thymic Treg Cell Lineage Commitment for Robust Immune Tolerance
    Dikiy Stanislav, Li Jun, Bai Lu, Jiang Menglin, Janke Laura, Zong Xinying, Hao Xiaolei, Hoyos Beatrice, Wang Zhong-Min, Xu Beisi, Fan Yiping, Rudensky Alexander Y., Feng Yongqiang
    Immunity 2021

    Activation of the STAT5 transcription factor downstream of the Interleukin-2 receptor (IL-2R) induces expression of Foxp3, a critical step in the differentiation of regulatory T (Treg) cells. Due to the pleiotropic effects of IL-2R signaling, it is unclear how STAT5 acts directly on the Foxp3 locus to promote its expression. Here, we report that IL-2 – STAT5 signaling converged on an enhancer (CNS0) during Foxp3 induction. CNS0 facilitated the IL-2 dependent CD25+Foxp3– precursor to Treg cell transition in the thymus. Its deficiency resulted in impaired Treg cell generation in neonates, which was partially mitigated with age. While the thymic Treg cell paucity caused by CNS0 deficiency did not result in autoimmunity on its own, it exacerbated autoimmune manifestations caused by disruption of the Aire gene. Thus, CNS0 enhancer activity ensures robust Treg cell differentiation early in postnatal life and cooperatively with other tolerance mechanisms minimizes autoimmunity.

  5. Distinct Dynamics and Functions of H2AK119ub1 and H3K27me3 in Mouse Preimplantation Embryos
    Chen Zhiyuan, Djekidel Mohamed Nadhir, Zhang Yi
    Nature Genetics 2021

    Polycomb repressive complexes 1 and 2 (PRC1/2) maintain transcriptional silencing of developmental genes largely by catalyzing the formation of mono-ubiquitinated histone H2A at lysine 119 (H2AK119ub1) and trimethylated histone H3 at lysine 27 (H3K27me3), respectively. How Polycomb domains are reprogrammed during mammalian preimplantation development remains largely unclear. Here we show that, although H2AK119ub1 and H3K27me3 are highly colocalized in gametes, they undergo differential reprogramming dynamics following fertilization. H3K27me3 maintains thousands of maternally biased domains until the blastocyst stage, whereas maternally biased H2AK119ub1 distribution in zygotes is largely equalized at the two-cell stage. Notably, while maternal PRC2 depletion has a limited effect on global H2AK119ub1 in early embryos, it disrupts allelic H2AK119ub1 at H3K27me3 imprinting loci including Xist. By contrast, acute H2AK119ub1 depletion in zygotes does not affect H3K27me3 imprinting maintenance, at least by the four-cell stage. Importantly, loss of H2AK119ub1, but not H3K27me3, causes premature activation of developmental genes during zygotic genome activation (ZGA) and subsequent embryonic arrest. Thus, our study reveals distinct dynamics and functions of H3K27me3 and H2AK119ub1 in mouse preimplantation embryos.

  6. Promoter Switching in Response to Changing Environment and Elevated Expression of Protein-Coding Genes Overlapping at Their 5’ Ends
    Rosikiewicz Wojciech, Sikora Jarosław, Skrzypczak Tomasz, Kubiak Magdalena R., Makałowska Izabela
    Scientific Reports 2021

    Despite the number of studies focused on sense-antisense transcription, the key question of whether such organization evolved as a regulator of gene expression or if this is only a byproduct of other regulatory processes has not been elucidated to date. In this study, protein-coding sense-antisense gene pairs were analyzed with a particular focus on pairs overlapping at their 5’ ends. Analyses were performed in 73 human transcription start site libraries. The results of our studies showed that the overlap between genes is not a stable feature and depends on which TSSs are utilized in a given cell type. An analysis of gene expression did not confirm that overlap between genes causes downregulation of their expression. This observation contradicts earlier findings. In addition, we showed that the switch from one promoter to another, leading to genes overlap, may occur in response to changing environment of a cell or tissue. We also demonstrated that in transfected and cancerous cells genes overlap is observed more often in comparison with normal tissues. Moreover, utilization of overlapping promoters depends on particular state of a cell and, at least in some groups of genes, is not merely coincidental.

  7. Proteasome Stress in Skeletal Muscle Mounts a Long-Range Protective Response That Delays Retinal and Brain Aging
    Rai Mamta, Coleman Zane, Curley Michelle, Nityanandam Anjana, Platt Anna, Robles-Murguia Maricela, Jiao Jianqin, Finkelstein David, Wang Yong-Dong, Xu Beisi, Fan Yiping, Demontis Fabio
    Cell Metabolism 2021

    Neurodegeneration in the central nervous system (CNS) is a defining feature of organismal aging that is influenced by peripheral tissues. Clinical observations indicate that skeletal muscle influences CNS aging, but the underlying muscle-to-brain signaling remains unexplored. In Drosophila, we find that moderate perturbation of the proteasome in skeletal muscle induces compensatory preservation of CNS proteostasis during aging. Such long-range stress signaling depends on muscle-secreted Amyrel amylase. Mimicking stress-induced Amyrel upregulation in muscle reduces age-related accumulation of poly-ubiquitinated proteins in the brain and retina via chaperones. Preservation of proteostasis stems from the disaccharide maltose, which is produced via Amyrel amylase activity. Correspondingly, RNAi for SLC45 maltose transporters reduces expression of Amyrel-induced chaperones and worsens brain proteostasis during aging. Moreover, maltose preserves proteostasis and neuronal activity in human brain organoids challenged by thermal stress. Thus, proteasome stress in skeletal muscle hinders retinal and brain aging by mounting an adaptive response via amylase/maltose.

  8. MethylationToActivity: A Deep-Learning Framework That Reveals Promoter Activity Landscapes from DNA Methylomes in Individual Tumors
    Williams Justin, Xu Beisi, Putnam Daniel, Thrasher Andrew, Li Chunliang, Yang Jun, Chen Xiang
    Genome Biology 2021

    Although genome-wide DNA methylomes have demonstrated their clinical value as reliable biomarkers for tumor detection, subtyping, and classification, their direct biological impacts at the individual gene level remain elusive. Here we present MethylationToActivity (M2A), a machine learning framework that uses convolutional neural networks to infer promoter activities based on H3K4me3 and H3K27ac enrichment, from DNA methylation patterns for individual genes. Using publicly available datasets in real-world test scenarios, we demonstrate that M2A is highly accurate and robust in revealing promoter activity landscapes in various pediatric and adult cancers, including both solid and hematologic malignant neoplasms.

2020:9

  1. FBXO11-Mediated Proteolysis of BAHD1 Relieves PRC2-Dependent Transcriptional Repression in Erythropoiesis
    Xu Peng, Scott Daniel C., Xu Beisi, Yao Yu, Feng Ruopeng, Cheng Li, Mayberry Kalin, Bi Wenjian, Palmer Lance E, King Moeko T., Wang Hong, Li Yuxin, Fan Yiping, Alpi Arno F., Li Chunliang, Peng Junmin, Papizan James, Pruett-Miller Shondra M, Spallek Ria, Bassermann Florian, Cheng Yong, Schulman Brenda A., Weiss Mitchell J.
    Blood 2020

    The histone mark H3K27me3 and its reader/writer Polycomb repressive complex 2 (PRC2) mediate widespread transcriptional repression in stem and progenitor cells. Mechanisms that regulate this activity are critical for hematopoietic development but poorly understood. Here we show that the E3 ubiquitin ligase FBXO11 relieves PRC2-mediated repression during erythroid maturation by targeting its newly identified substrate BAHD1, an H3K27me3 reader that recruits transcriptional co-repressors. Erythroblasts lacking FBXO11 are developmentally delayed, with reduced expression of maturation-associated genes, most of which harbor bivalent histone marks (activating H3K4me3 and repressive H3K27me3), bind BAHD1, and fail to recruit the erythroid transcription factor GATA1. The BAHD1 complex interacts physically with PRC2 and depletion of either component restores FBXO11-deficient erythroid gene expression. Our studies identify BAHD1 as a novel effector of PRC2-mediated repression and reveal how a single E3 ubiquitin ligase eliminates PRC2 repression at developmentally poised bivalent genes during erythropoiesis.

  2. Functional Interrogation of HOXA9 Regulome in MLLr Leukemia via Reporter-Based CRISPR/Cas9 Screen
    Zhang Hao, Zhang Yang, Zhou Xinyue, Wright Shaela, Hyle Judith, Zhao Lianzhong, An Jie, Zhao Xujie, Shao Ying, Xu Beisi, Lee Hyeong-Min, Chen Taosheng, Zhou Yang, Chen Xiang, Lu Rui, Li Chunliang
    eLife 2020

    Aberrant HOXA9 expression is a hallmark of most aggressive acute leukemias, notably those with KMT2A (MLL) gene rearrangements. HOXA9 overexpression not only predicts poor diagnosis and outcome but also plays a critical role in leukemia transformation and maintenance. However, our current understanding of HOXA9 regulation in leukemia is limited, hindering development of therapeutic strategies. Here, we generated the HOXA9-mCherry knock-in reporter cell lines to dissect HOXA9 regulation. By utilizing the reporter and CRISPR/Cas9 screens, we identified transcription factors controlling HOXA9 expression, including a novel regulator, USF2, whose depletion significantly down-regulated HOXA9 expression and impaired MLLr leukemia cell proliferation. Ectopic expression of Hoxa9 rescued impaired leukemia cell proliferation upon USF2 loss. Cut and Run analysis revealed the direct occupancy of USF2 at HOXA9 promoter in MLLr leukemia cells. Collectively, the HOXA9 reporter facilitated the functional interrogation of the HOXA9 regulome and has advanced our understanding of the molecular regulation network in HOXA9-driven leukemia.

  3. UTX/KDM6A Suppresses AP-1 and a Gliogenesis Program during Neural Differentiation of Human Pluripotent Stem Cells
    Xu Beisi*, Mulvey Brett*, Salie Muneeb, Yang Xiaoyang, Matsui Yurika, Nityanandam Anjana, Fan Yiping, Peng Jamy C.
    Epigenetics & Chromatin 2020

    Background UTX/KDM6A is known to interact and influence multiple different chromatin modifiers to promote an open chromatin environment to facilitate gene activation, but its molecular activities in developmental gene regulation remain unclear. Results We report that in human neural stem cells, UTX binding correlates with both promotion and suppression of gene expression. These activities enable UTX to modulate neural stem cell self-renewal, promote neurogenesis, and suppress gliogenesis. In neural stem cells, UTX has a less influence over histone H3 lysine 27 and lysine 4 methylation but more predominantly affects histone H3 lysine 27 acetylation and chromatin accessibility. Furthermore, UTX suppresses components of AP-1 and, in turn, a gliogenesis program. Conclusions Our findings revealed that UTX coordinates dualistic gene regulation to govern neural stem cell properties and neurogenesis–gliogenesis switch.

  4. Ybx1 Fine-Tunes PRC2 Activities to Control Embryonic Brain Development
    Evans Myron K., Matsui Yurika*, Xu Beisi*, Willis Catherine, Loome Jennifer, Milburn Luis, Fan Yiping, Pagala Vishwajeeth, Peng Jamy C.
    Nature Communications 2020

    Chromatin modifiers affect spatiotemporal gene expression programs that underlie organismal development. The Polycomb repressive complex 2 (PRC2) is a crucial chromatin modifier in executing neurodevelopmental programs. Here, we find that PRC2 interacts with the nucleic acid–binding protein Ybx1. In the mouse embryo in vivo, Ybx1 is required for forebrain specification and restricting mid-hindbrain growth. In neural progenitor cells (NPCs), Ybx1 controls self-renewal and neuronal differentiation. Mechanistically, Ybx1 highly overlaps PRC2 binding genome-wide, controls PRC2 distribution, and inhibits H3K27me3 levels. These functions are consistent with Ybx1-mediated promotion of genes involved in forebrain specification, cell proliferation, or neuronal differentiation. In Ybx1-knockout NPCs, H3K27me3 reduction by PRC2 enzymatic inhibitor or genetic depletion partially rescues gene expression and NPC functions. Our findings suggest that Ybx1 fine-tunes PRC2 activities to regulate spatiotemporal gene expression in embryonic neural development and uncover a crucial epigenetic mechanism balancing forebrain–hindbrain lineages and self-renewal–differentiation choices in NPCs.

  5. TET2 Deficiency Reprograms the Germinal Center B Cell Epigenome and Silences Genes Linked to Lymphomagenesis
    Rosikiewicz Wojciech*, Chen Xiaowen*, Dominguez Pilar M.*, Ghamlouch Hussein, Aoufouchi Said, Bernard Olivier A., Melnick Ari, Li Sheng
    Science Advances 2020

    The TET2 DNA hydroxymethyltransferase is frequently disrupted by somatic mutations in diffuse large B cell lymphomas (DLBCLs), a tumor that originates from germinal center (GC) B cells. Here, we show that TET2 deficiency leads to DNA hypermethylation of regulatory elements in GC B cells, associated with silencing of the respective genes. This hypermethylation affects the binding of transcription factors including those involved in exit from the GC reaction and involves pathways such as B cell receptor, antigen presentation, CD40, and others. Normal GC B cells manifest a typical hypomethylation signature, which is caused by AID, the enzyme that mediates somatic hypermutation. However, AID-induced demethylation is markedly impaired in TET2-deficient GC B cells, suggesting that AID epigenetic effects are partially dependent on TET2. Last, we find that TET2 mutant DLBCLs also manifest the aberrant TET2-deficient GC DNA methylation signature, suggesting that this epigenetic pattern is maintained during and contributes to lymphomagenesis. TET2 deficiency reprograms B cell epigenome linked to lymphomagenesis. TET2 deficiency reprograms B cell epigenome linked to lymphomagenesis.

  6. MYCN Amplification and ATRX Mutations Are Incompatible in Neuroblastoma
    Zeineldin Maged, Federico Sara, Chen Xiang, Fan Yiping, Xu Beisi, Stewart Elizabeth, Zhou Xin, Jeon Jongrye, Griffiths Lyra, Nguyen Rosa, Norrie Jackie, Easton John, Mulder Heather, Yergeau Donald, Liu Yanling, Wu Jianrong, Van Ryn Collin, Naranjo Arlene, Hogarty Michael D., Kamiński Marcin M., Valentine Marc, Pruett-Miller Shondra M., Pappo Alberto, Zhang Jinghui, Clay Michael R., Bahrami Armita, Vogel Peter, Lee Seungjae, Shelat Anang, Sarthy Jay F., Meers Michael P., George Rani E., Mardis Elaine R., Wilson Richard K., Henikoff Steven, Downing James R., Dyer Michael A.
    Nature Communications 2020

    Aggressive cancers often have activating mutations in growth-controlling oncogenes and inactivating mutations in tumor-suppressor genes. In neuroblastoma, amplification of the MYCN oncogene and inactivation of the ATRX tumor-suppressor gene correlate with high-risk disease and poor prognosis. Here we show that ATRX mutations and MYCN amplification are mutually exclusive across all ages and stages in neuroblastoma. Using human cell lines and mouse models, we found that elevated MYCN expression and ATRX mutations are incompatible. Elevated MYCN levels promote metabolic reprogramming, mitochondrial dysfunction, reactive-oxygen species generation, and DNA-replicative stress. The combination of replicative stress caused by defects in the ATRX–histone chaperone complex, and that induced by MYCN-mediated metabolic reprogramming, leads to synthetic lethality. Therefore, ATRX and MYCN represent an unusual example, where inactivation of a tumor-suppressor gene and activation of an oncogene are incompatible. This synthetic lethality may eventually be exploited to improve outcomes for patients with high-risk neuroblastoma.

  7. From Influenza Virus Infections to Lupus: Synchronous Estrogen Receptor α and RNA Polymerase II Binding Within the Immunoglobulin Heavy Chain Locus
    Jones Bart G., Sealy Robert E., Penkert Rhiannon R., Surman Sherri L., Birshtein Barbara K., Xu Beisi, Neale Geoffrey, Maul Robert W., Gearhart Patricia J., Hurwitz Julia L.
    Viral Immunology 2020

    Males and females respond to pathogens differently and exhibit significantly different frequencies of autoimmune disease. For example, vaccinated adult females control influenza virus better than males, but females suffer systemic lupus erythematosus at a 9:1 frequency compared to males. Numerous explanations have been offered for these sex differences, but most have involved indirect mechanisms by which estrogen, a nuclear hormone, modifies cell barriers or immunity. In search of a direct mechanism, we examined the binding of estrogen receptor α (ERα), a class I nuclear hormone receptor, to the immunoglobulin heavy chain locus. Here, we show that in purified murine B cells, ERα and RNA polymerase II (RNA Pol II) exhibit extraordinarily similar DNA binding patterns. We further demonstrate that ERα preferentially binds adenosine–cytidine (AC)-repeats in the immunoglobulin heavy chain locus when supplemental estrogen is added to purified, lipopolysaccharide-activated B cells. Based on these and previous data, we hypothesize that (i) estrogen guides the binding of ERα and its RNA Pol II partner within the locus, which in turn instructs sterile transcription and class switch recombination (CSR), (ii) ERα binding to AC-repeats modifies the DNA architecture and loops associated with CSR, and (iii) by these mechanisms, estrogen instructs antibody expression. By targeting ERα-DNA interactions in the immunoglobulin heavy chain locus, clinicians may ultimately enhance antibody responses in the context of infectious diseases and reduce antibody responses in the context of allergic or autoimmune reactions.

  8. An ABC Transporter Drives Medulloblastoma Pathogenesis by Regulating Sonic Hedgehog Signaling
    Wijaya Juwina, Vo BaoHan T., Liu Jingjing, Xu Beisi, Wu Gang, Wang Yao, Peng Junmin, Zhang Jin, Janke Laura J., Orr Brent A., Yu Jiyang, Roussel Martine F., Schuetz John D.
    Cancer Research 2020

    Mutations in Sonic hedgehog (SHH) signaling promote aberrant proliferation and tumor growth. SHH medulloblastoma (MB) is among the most frequent brain tumors in children less than three years of age. While key components of the SHH pathway are well-known, we hypothesized that new disease-modifying targets of SHH MB might be identified from large-scale bioinformatics and systems biology analyses. Using a data-driven systems biology approach, we built a medulloblastoma-specific interactome. The ATP-binding cassette transporter ABCC4 was identified as a modulator of SHH-MB. Accordingly, increased ABCC4 expression correlated with poor overall survival in SHH MB patients. Knockdown of ABCC4 expression markedly blunted the constitutive activation of the SHH pathway secondary to Ptch1 or Sufu insufficiency. In human tumor cell lines, ABCC4 knockdown and inhibition reduced full-length GLI3 levels. In a clinically relevant murine SHH MB model, targeted ablation of Abcc4 in primary tumors significantly reduced tumor burden and extended the lifespan of tumor-bearing mice. These studies reveal ABCC4 as a potent SHH pathway regulator and a new candidate to target with the potential to improve SHH MB therapy.

  9. Muscle-Derived Dpp Regulates Feeding Initiation via Endocrine Modulation of Brain Dopamine Biosynthesis
    Robles-Murguia Maricela, Rao Deepti, Finkelstein David, Xu Beisi, Fan Yiping, Demontis Fabio
    Genes & Development 2020

    In animals, the brain regulates feeding behavior in response to local energy demands of peripheral tissues, which secrete orexigenic and anorexigenic hormones. Although skeletal muscle is a key peripheral tissue, it remains unknown whether muscle-secreted hormones regulate feeding. In Drosophila, we found that decapentaplegic (dpp), the homolog of human bone morphogenetic proteins BMP2 and BMP4, is a muscle-secreted factor (a myokine) that is induced by nutrient sensing and that circulates and signals to the brain. Muscle-restricted dpp RNAi promotes foraging and feeding initiation, whereas dpp overexpression reduces it. This regulation of feeding by muscle-derived Dpp stems from modulation of brain tyrosine hydroxylase (TH) expression and dopamine biosynthesis. Consistently, Dpp receptor signaling in dopaminergic neurons regulates TH expression and feeding initiation via the downstream transcriptional repressor Schnurri. Moreover, pharmacologic modulation of TH activity rescues the changes in feeding initiation due to modulation of dpp expression in muscle. These findings indicate that muscle-to-brain endocrine signaling mediated by the myokine Dpp regulates feeding behavior.

2019:10

  1. Matters of Life and Death: How Estrogen and Estrogen Receptor Binding to the Immunoglobulin Heavy Chain Locus May Influence Outcomes of Infection, Allergy, and Autoimmune Disease
    Jones Bart G., Penkert Rhiannon R., Surman Sherri L., Sealy Robert E., Pelletier Stephane, Xu Beisi, Neale Geoff, Maul Robert W., Gearhart Patricia J., Hurwitz J. L.
    Cellular Immunology 2019

    Sex hormones are best known for their influences on reproduction, but they also have profound influences on the immune response. Examples of sex-specific differences include: (i) the relatively poor control of influenza virus infections in males compared to females, (ii) allergic asthma, an IgE-associated hypersensitivity reaction that is exacerbated in adolescent females compared to males, and (iii) systemic lupus erythematosus, a life-threatening autoimmune disease with a 9:1 female:male bias. Here we consider how estrogen and estrogen receptor α (ERα) may influence the immune response by modifying class switch recombination (CSR) and immunoglobulin expression patterns. We focus on ERα binding to enhancers (Eμ and the 3’ regulatory region) and switch sites (S\textmu and Sε) in the immunoglobulin heavy chain locus. Our preliminary data from ChIP-seq analyses of purified, activated B cells show estrogen-mediated changes in the positioning of ERα binding within and near S\textmu and Sε. In the presence of estrogen, ERα is bound not only to estrogen response elements (ERE), but also to adenosine-cytidine (AC)-repeats and poly adenosine (poly A) sequences, in some cases within constant region gene introns. We propose that by binding these sites, estrogen and ERα directly participate in the DNA loop formation required for CSR. We further suggest that estrogen regulates immunoglobulin expression patterns and can thereby influence life-and-death outcomes of infection, hypersensitivity, and autoimmune disease.

  2. Targeting REGNASE-1 Programs Long-Lived Effector T Cells for Cancer Therapy
    Wei Jun, Long Lingyun, Zheng Wenting, Dhungana Yogesh, Lim Seon Ah, Guy Cliff, Wang Yanyan, Wang Yong-Dong, Qian Chenxi, Xu Beisi, Kc Anil, Saravia Jordy, Huang Hongling, Yu Jiyang, Doench John G., Geiger Terrence L., Chi Hongbo
    Nature 2019

    Adoptive cell therapy represents a new paradigm in cancer immunotherapy, but it can be limited by the poor persistence and function of transferred T cells1. Here we use an in vivo pooled CRISPR–Cas9 mutagenesis screening approach to demonstrate that, by targeting REGNASE-1, CD8+ T cells are reprogrammed to long-lived effector cells with extensive accumulation, better persistence and robust effector function in tumours. REGNASE-1-deficient CD8+ T cells show markedly improved therapeutic efficacy against mouse models of melanoma and leukaemia. By using a secondary genome-scale CRISPR–Cas9 screening, we identify BATF as the key target of REGNASE-1 and as a rheostat that shapes antitumour responses. Loss of BATF suppresses the increased accumulation and mitochondrial fitness of REGNASE-1-deficient CD8+ T cells. By contrast, the targeting of additional signalling factors—including PTPN2 and SOCS1—improves the therapeutic efficacy of REGNASE-1-deficient CD8+ T cells. Our findings suggest that T cell persistence and effector function can be coordinated in tumour immunity and point to avenues for improving the efficacy of adoptive cell therapy for cancer.

  3. Nucleome Dynamics during Retinal Development
    Norrie Jackie L.*, Lupo Marybeth S.*, Xu Beisi*, Al Diri Issam*, Valentine Marc*, Putnam Daniel*, Griffiths Lyra, Zhang Jiakun, Johnson Dianna, Easton John, Shao Ying, Honnell Victoria, Frase Sharon, Miller Shondra, Stewart Valerie, Zhou Xin, Chen Xiang, Dyer Michael A.
    Neuron 2019

    Summary More than 8,000 genes are turned on or off as progenitor cells produce the 7 classes of retinal cell types during development. Thousands of enhancers are also active in the developing retinae, many having features of cell- and developmental stage-specific activity. We studied dynamic changes in the 3D chromatin landscape important for precisely orchestrated changes in gene expression during retinal development by ultra-deep in situ Hi-C analysis on murine retinae. We identified developmental-stage-specific changes in chromatin compartments and enhancer-promoter interactions. We developed a machine learning-based algorithm to map euchromatin and heterochromatin domains genome-wide and overlaid it with chromatin compartments identified by Hi-C. Single-cell ATAC-seq and RNA-seq were integrated with our Hi-C and previous ChIP-seq data to identify cell- and developmental-stage-specific super-enhancers (SEs). We identified a bipolar neuron-specific core regulatory circuit SE upstream of Vsx2, whose deletion in mice led to the loss of bipolar neurons.

  4. Acute Depletion of CTCF Directly Affects MYC Regulation through Loss of Enhancer–Promoter Looping
    Hyle Judith, Zhang Yang, Wright Shaela, Xu Beisi, Shao Ying, Easton John, Tian Liqing, Feng Ruopeng, Xu Peng, Li Chunliang
    Nucleic Acids Research 2019

    Numerous pieces of evidence support the complex, 3D spatial organization of the genome dictates gene expression. CTCF is essential to define topologically associated domain boundaries and to facilitate the formation of insulated chromatin loop structures. To understand CTCF’s direct role in global transcriptional regulation, we integrated the miniAID-mClover3 cassette to the endogenous CTCF locus in a human pediatric B-ALL cell line, SEM, and an immortal erythroid precursor cell line, HUDEP-2, to allow for acute depletion of CTCF protein by the auxin-inducible degron system. In SEM cells, CTCF loss notably disrupted intra-TAD loops and TAD integrity in concurrence with a reduction in CTCF-binding affinity, while showing no perturbation to nuclear compartment integrity. Strikingly, the overall effect of CTCF’s loss on transcription was minimal. Whole transcriptome analysis showed hundreds of genes differentially expressed in CTCF-depleted cells, among which MYC and a number of MYC target genes were specifically downregulated. Mechanically, acute depletion of CTCF disrupted the direct interaction between the MYC promoter and its distal enhancer cluster residing ∼1.8 Mb downstream. Notably, MYC expression was not profoundly affected upon CTCF loss in HUDEP-2 cells suggesting that CTCF could play a B-ALL cell line specific role in maintaining MYC expression.

  5. Long-Read Sequencing Unveils IGH-DUX4 Translocation into the Silenced IGH Allele in B-Cell Acute Lymphoblastic Leukemia
    Tian Liqing, Shao Ying, Nance Stephanie, Dang Jinjun, Xu Beisi, Ma Xiaotu, Li Yongjin, Ju Bensheng, Dong Li, Newman Scott, Zhou Xin, Schreiner Patrick, Tseng Elizabeth, Hon Ting, Ashby Meredith, Li Chunliang, Easton John, Gruber Tanja A., Zhang Jinghui
    Nature Communications 2019

    IGH@ proto-oncogene translocation is a common oncogenic event in lymphoid lineage cancers such as B-ALL, lymphoma and multiple myeloma. Here, to investigate the interplay between IGH@ proto-oncogene translocation and IGH allelic exclusion, we perform long-read whole-genome and transcriptome sequencing along with epigenetic and 3D genome profiling of Nalm6, an IGH-DUX4 positive B-ALL cell line. We detect significant allelic imbalance on the wild-type over the IGH-DUX4 haplotype in expression and epigenetic data, showing IGH-DUX4 translocation occurs on the silenced IGH allele. In vitro, this reduces the oncogenic stress of DUX4 high-level expression. Moreover, patient samples of IGH-DUX4 B-ALL have similar expression profile and IGH breakpoints as Nalm6, suggesting a common mechanism to allow optimal dosage of non-toxic DUX4 expression.

  6. H3.3 K27M Depletion Increases Differentiation and Extends Latency of Diffuse Intrinsic Pontine Glioma Growth in Vivo
    Silveira André B., Kasper Lawryn H., Fan Yiping, Jin Hongjian, Wu Gang, Shaw Timothy I., Zhu Xiaoyan, Larson Jon D., Easton John, Shao Ying, Yergeau Donald A., Rosencrance Celeste, Boggs Kristy, Rusch Michael C., Ding Liang, Zhang Junyuan, Finkelstein David, Noyes Rachel M., Russell Brent L., Xu Beisi, Broniscer Alberto, Wetmore Cynthia, Pounds Stanley B., Ellison David W., Zhang Jinghui, Baker Suzanne J.
    Acta Neuropathologica 2019

    Histone H3 K27M mutation is the defining molecular feature of the devastating pediatric brain tumor, diffuse intrinsic pontine glioma (DIPG). The prevalence of histone H3 K27M mutations indicates a critical role in DIPGs, but the contribution of the mutation to disease pathogenesis remains unclear. We show that knockdown of this mutation in DIPG xenografts restores K27M-dependent loss of H3K27me3 and delays tumor growth. Comparisons of matched DIPG xenografts with and without K27M knockdown allowed identification of mutation-specific effects on the transcriptome and epigenome. The resulting transcriptional changes recapitulate expression signatures from K27M primary DIPG tumors and are strongly enriched for genes associated with nervous system development. Integrated analysis of ChIP-seq and expression data showed that genes upregulated by the mutation are overrepresented in apparently bivalent promoters. Many of these targets are associated with more immature differentiation states. Expression profiles indicate K27M knockdown decreases proliferation and increases differentiation within lineages represented in DIPG. These data suggest that K27M-mediated loss of H3K27me3 directly regulates a subset of genes by releasing poised promoters, and contributes to tumor phenotype and growth by limiting differentiation. The delayed tumor growth associated with knockdown of H3 K27M provides evidence that this highly recurrent mutation is a relevant therapeutic target.

  7. Complex Sex-Biased Antibody Responses: Estrogen Receptors Bind Estrogen Response Elements Centered within Immunoglobulin Heavy Chain Gene Enhancers
    Jones Bart G, Sealy Robert E, Penkert Rhiannon R, Surman Sherri L, Maul Robert W, Neale Geoff, Xu Beisi, Gearhart Patricia J, Hurwitz Julia L
    International Immunology 2019

    Nuclear hormone receptors including the estrogen receptor (ERα) and the retinoic acid receptor regulate a plethora of biological functions including reproduction, circulation and immunity. To understand how estrogen and other nuclear hormones influence antibody production, we characterized total serum antibody isotypes in female and male mice of C57BL/6J, BALB/cJ and C3H/HeJ mouse strains. Antibody levels were higher in females compared to males in all strains and there was a female preference for IgG2b production. Sex-biased patterns were influenced by vitamin levels, and by antigen specificity toward influenza virus or pneumococcus antigens. To help explain sex biases, we examined the direct effects of estrogen on immunoglobulin heavy chain sterile transcript production among purified, lipopolysaccharide-stimulated B cells. Supplemental estrogen in B-cell cultures significantly increased immunoglobulin heavy chain sterile transcripts. Chromatin immunoprecipitation analyses of activated B cells identified significant ERα binding to estrogen response elements (EREs) centered within enhancer elements of the immunoglobulin heavy chain locus, including the E\textmu enhancer and hypersensitive site 1,2 (HS1,2) in the 3’ regulatory region. The ERE in HS1,2 was conserved across animal species, and in humans marked a site of polymorphism associated with the estrogen-augmented autoimmune disease, lupus. Taken together, the results highlight: (i) the important targets of ERα in regulatory regions of the immunoglobulin heavy chain locus that influence antibody production, and (ii) the complexity of mechanisms by which estrogen instructs sex-biased antibody production profiles.

  8. Differentiation of Human Pluripotent Stem Cells into Neurons or Cortical Organoids Requires Transcriptional Co-Regulation by UTX and 53BP1
    Yang Xiaoyang*, Xu Beisi*, Mulvey Brett, Evans Myron, Jordan Samuel, Wang Yong-Dong, Pagala Vishwajeeth, Peng Junmin, Fan Yiping, Patel Arishna, Peng Jamy C.
    Nature Neuroscience 2019

    UTX is a chromatin modifier required for development and neural lineage specification, but how it controls these biological processes is unclear. To determine the molecular mechanisms of UTX, we identified novel UTX protein interaction partners. Here we show that UTX and 53BP1 directly interact and co-occupy promoters in human embryonic stem cells and differentiating neural progenitor cells. Human 53BP1 contains a UTX-binding site that diverges from its mouse homolog by 41%, and disruption of the 53BP1–UTX interaction abrogated human, but not mouse, neurogenesis in vitro. The 53BP1–UTX interaction is required to upregulate key neurodevelopmental genes during the differentiation of human embryonic stem cells into neurons or into cortical organoids. 53BP1 promotes UTX chromatin binding, and in turn H3K27 modifications and gene activation, at a subset of genomic regions, including neurogenic genes. Overall, our data suggest that the 53BP1–UTX interaction supports the activation of key genes required for human neurodevelopment.

  9. Histone H3.3 K27M Accelerates Spontaneous Brainstem Glioma and Drives Restricted Changes in Bivalent Gene Expression
    Larson Jon D., Kasper Lawryn H., Paugh Barbara S., Jin Hongjian, Wu Gang, Kwon Chang-Hyuk, Fan Yiping, Shaw Timothy I., Silveira André B., Qu Chunxu, Xu Raymond, Zhu Xiaoyan, Zhang Junyuan, Russell Helen R., Peters Jennifer L., Finkelstein David, Xu Beisi, Lin Tong, Tinkle Christopher L., Patay Zoltan, Onar-Thomas Arzu, Pounds Stanley B., McKinnon Peter J., Ellison David W., Zhang Jinghui, Baker Suzanne J.
    Cancer Cell 2019

    Summary Diffuse intrinsic pontine gliomas (DIPGs) are incurable childhood brainstem tumors with frequent histone H3 K27M mutations and recurrent alterations in PDGFRA and TP53. We generated genetically engineered inducible mice and showed that H3.3 K27M enhanced neural stem cell self-renewal while preserving regional identity. Neonatal induction of H3.3 K27M cooperated with activating platelet-derived growth factor receptor α (PDGFRα) mutant and Trp53 loss to accelerate development of diffuse brainstem gliomas that recapitulated human DIPG gene expression signatures and showed global changes in H3K27 posttranslational modifications, but relatively restricted gene expression changes. Genes upregulated in H3.3 K27M tumors were enriched for those associated with neural development where H3K27me3 loss released the poised state of apparently bivalent promoters, whereas downregulated genes were enriched for those encoding homeodomain transcription factors.

  10. Metabolic Heterogeneity Underlies Reciprocal Fates of T H 17 Cell Stemness and Plasticity
    Karmaus Peer W. F., Chen Xiang, Lim Seon Ah, Herrada Andrés A., Nguyen Thanh-Long M., Xu Beisi, Dhungana Yogesh, Rankin Sherri, Chen Wenan, Rosencrance Celeste, Yang Kai, Fan Yiping, Cheng Yong, Easton John, Neale Geoffrey, Vogel Peter, Chi Hongbo
    Nature 2019

    A defining feature of adaptive immunity is the development of long-lived memory T cells to curtail infection. Recent studies have identified a unique stem-like T-cell subset amongst exhausted CD8-positive T cells in chronic infection1–3, but it remains unclear whether CD4-positive T-cell subsets with similar features exist in chronic inflammatory conditions. Amongst helper T cells, TH17 cells have prominent roles in autoimmunity and tissue inflammation and are characterized by inherent plasticity4–7, although how such plasticity is regulated is poorly understood. Here we demonstrate that TH17 cells in a mouse model of autoimmune disease are functionally and metabolically heterogeneous; they contain a subset with stemness-associated features but lower anabolic metabolism, and a reciprocal subset with higher metabolic activity that supports transdifferentiation into TH1-like cells. These two TH17-cell subsets are defined by selective expression of the transcription factors TCF-1 and T-bet, and by discrete levels of CD27 expression. We also identify signalling via the kinase complex mTORC1 as a central regulator of TH17-cell fate decisions by coordinating metabolic and transcriptional programmes. TH17 cells with disrupted mTORC1 signalling or anabolic metabolism fail to induce autoimmune neuroinflammation or to develop into TH1-like cells, but instead upregulate TCF-1 expression and acquire stemness-associated features. Single-cell RNA sequencing and experimental validation reveal heterogeneity in fate-mapped TH17 cells, and a developmental arrest in the TH1 transdifferentiation trajectory upon loss of mTORC1 activity or metabolic perturbation. Our results establish that the dichotomy of stemness and effector function underlies the heterogeneous TH17 responses and autoimmune pathogenesis, and point to previously unappreciated metabolic control of plasticity in helper T cells.

2018:6

  1. The Hippo Pathway Prevents YAP/TAZ-Driven Hypertranscription and Controls Neural Progenitor Number
    Lavado Alfonso, Park Jun Young, Paré Joshua, Finkelstein David, Pan Haitao, Xu Beisi, Fan Yiping, Kumar Ram Parikshan, Neale Geoffrey, Kwak Young Don, McKinnon Peter J., Johnson Randy L., Cao Xinwei
    Developmental Cell 2018

    Summary The Hippo pathway controls the activity of YAP/TAZ transcriptional coactivators through a kinase cascade. Despite the critical role of this pathway in tissue growth and tumorigenesis, it remains unclear how YAP/TAZ-mediated transcription drives proliferation. By analyzing the effects of inactivating LATS1/2 kinases, the direct upstream inhibitors of YAP/TAZ, on mouse brain development and applying cell-number-normalized transcriptome analyses, we discovered that YAP/TAZ activation causes a global increase in transcription activity, known as hypertranscription, and upregulates many genes associated with cell growth and proliferation. In contrast, conventional read-depth-normalized RNA-sequencing analysis failed to detect the scope of the transcriptome shift and missed most relevant gene ontologies. Following a transient increase in proliferation, however, hypertranscription in neural progenitors triggers replication stress, DNA damage, and p53 activation, resulting in massive apoptosis. Our findings reveal a significant impact of YAP/TAZ activation on global transcription activity and have important implications for understanding YAP/TAZ function.

  2. The Genetic Basis and Cell of Origin of Mixed Phenotype Acute Leukaemia
    Alexander Thomas B., Gu Zhaohui, Iacobucci Ilaria, Dickerson Kirsten, Choi John K., Xu Beisi, Payne-Turner Debbie, Yoshihara Hiroki, Loh Mignon L., Horan John, Buldini Barbara, Basso Giuseppe, Elitzur Sarah, de Haas Valerie, Zwaan C. Michel, Yeoh Allen, Reinhardt Dirk, Tomizawa Daisuke, Kiyokawa Nobutaka, Lammens Tim, De Moerloose Barbara, Catchpoole Daniel, Hori Hiroki, Moorman Anthony, Moore Andrew S., Hrusak Ondrej, Meshinchi Soheil, Orgel Etan, Devidas Meenakshi, Borowitz Michael, Wood Brent, Heerema Nyla A., Carrol Andrew, Yang Yung-Li, Smith Malcolm A., Davidsen Tanja M., Hermida Leandro C., Gesuwan Patee, Marra Marco A., Ma Yussanne, Mungall Andrew J., Moore Richard A., Jones Steven J. M., Valentine Marcus, Janke Laura J., Rubnitz Jeffrey E., Pui Ching-Hon, Ding Liang, Liu Yu, Zhang Jinghui, Nichols Kim E., Downing James R., Cao Xueyuan, Shi Lei, Pounds Stanley, Newman Scott, Pei Deqing, Guidry Auvil Jaime M., Gerhard Daniela S., Hunger Stephen P., Inaba Hiroto, Mullighan Charles G.
    Nature 2018

    Mixed phenotype acute leukaemia (MPAL) is a high-risk subtype of leukaemia with myeloid and lymphoid features, limited genetic characterization, and a lack of consensus regarding appropriate therapy. Here we show that the two principal subtypes of MPAL, T/myeloid (T/M) and B/myeloid (B/M), are genetically distinct. Rearrangement of ZNF384 is common in B/M MPAL, and biallelic WT1 alterations are common in T/M MPAL, which shares genomic features with early T-cell precursor acute lymphoblastic leukaemia. We show that the intratumoral immunophenotypic heterogeneity characteristic of MPAL is independent of somatic genetic variation, that founding lesions arise in primitive haematopoietic progenitors, and that individual phenotypic subpopulations can reconstitute the immunophenotypic diversity in vivo. These findings indicate that the cell of origin and founding lesions, rather than an accumulation of distinct genomic alterations, prime tumour cells for lineage promiscuity. Moreover, these findings position MPAL in the spectrum of immature leukaemias and provide a genetically informed framework for future clinical trials of potential treatments for MPAL.

  3. Identification of Therapeutic Targets in Rhabdomyosarcoma through Integrated Genomic, Epigenomic, and Proteomic Analyses
    Stewart Elizabeth, McEvoy Justina, Wang Hong, Chen Xiang, Honnell Victoria, Ocarz Monica, Gordon Brittney, Dapper Jason, Blankenship Kaley, Yang Yanling, Li Yuxin, Shaw Timothy I., Cho Ji-Hoon, Wang Xusheng, Xu Beisi, Gupta Pankaj, Fan Yiping, Liu Yu, Rusch Michael, Griffiths Lyra, Jeon Jongrye, Freeman Burgess B., Clay Michael R., Pappo Alberto, Easton John, Shurtleff Sheila, Shelat Anang, Zhou Xin, Boggs Kristy, Mulder Heather, Yergeau Donald, Bahrami Armita, Mardis Elaine R., Wilson Richard K., Zhang Jinghui, Peng Junmin, Downing James R., Dyer Michael A.
    Cancer Cell 2018

    Summary Personalized cancer therapy targeting somatic mutations in patient tumors is increasingly being incorporated into practice. Other therapeutic vulnerabilities resulting from changes in gene expression due to tumor specific epigenetic perturbations are progressively being recognized. These genomic and epigenomic changes are ultimately manifest in the tumor proteome and phosphoproteome. We integrated transcriptomic, epigenomic, and proteomic/phosphoproteomic data to elucidate the cellular origins and therapeutic vulnerabilities of rhabdomyosarcoma (RMS). We discovered that alveolar RMS occurs further along the developmental program than embryonal RMS. We also identified deregulation of the RAS/MEK/ERK/CDK4/6, G2/M, and unfolded protein response pathways through our integrated analysis. Comprehensive preclinical testing revealed that targeting the WEE1 kinase in the G2/M pathway is the most effective approach in vivo for high-risk RMS.

  4. Mouse Medulloblastoma Driven by CRISPR Activation of Cellular Myc
    Vo BaoHan T., Kwon Jin Ah, Li Chunliang, Finkelstein David, Xu Beisi, Orr Brent A., Sherr Charles J., Roussel Martine F.
    Scientific Reports 2018

    MYC-driven Group 3 (G3) medulloblastoma (MB) is the most aggressive of four molecular subgroups classified by transcriptome, genomic landscape and clinical outcomes. Mouse models that recapitulate human G3 MB all rely on retroviral vector-induced Myc expression driven by viral regulatory elements (Retro-Myc tumors). We used nuclease-deficient CRISPR/dCas9-based gene activation with combinatorial single guide RNAs (sgRNAs) to enforce transcription of endogenous Myc in Trp53-null neurospheres that were orthotopically transplanted into the brains of naïve animals. Three combined sgRNAs linked to dCas9-VP160 induced cellular Myc expression and large cell anaplastic MBs (CRISPR-Myc tumors) which recapitulated the molecular characteristics of mouse and human G3 MBs. The BET inhibitor JQ1 suppressed MYC expression in a human G3 MB cell line (HD-MB03) and CRISPR-Myc, but not in Retro-Myc MBs. This G3 MB mouse model in which Myc expression is regulated by its own promoter will facilitate pre-clinical studies with drugs that regulate Myc transcription.

  5. MYC Drives a Subset of High-Risk Pediatric Neuroblastomas and Is Activated through Mechanisms Including Enhancer Hijacking and Focal Enhancer Amplification
    Zimmerman Mark W., Liu Yu, He Shuning, Durbin Adam D., Abraham Brian J., Easton John, Shao Ying, Xu Beisi, Zhu Shizhen, Zhang Xiaoling, Li Zhaodong, Weichert-Leahey Nina, Young Richard A., Zhang Jinghui, Look A. Thomas
    Cancer Discovery 2018

    The amplified MYCN gene serves as an oncogenic driver in approximately 20% of high-risk pediatric neuroblastomas. Here, we show that the family member MYC is a potent transforming gene in a separate subset of high-risk neuroblastoma cases (∼10%), based on (i) its upregulation by focal enhancer amplification or genomic rearrangements leading to enhancer hijacking, and (ii) its ability to transform neuroblastoma precursor cells in a transgenic animal model. The aberrant regulatory elements associated with oncogenic MYC activation include focally amplified distal enhancers and translocation of highly active enhancers from other genes to within topologically associating domains containing the MYC gene locus. The clinical outcome for patients with high levels of MYC expression is virtually identical to that of patients with amplification of the MYCN gene, a known high-risk feature of this disease. Together, these findings establish MYC as a bona fide oncogene in a clinically significant group of high-risk childhood neuroblastomas. Significance: Amplification of the MYCN oncogene is a recognized hallmark of high-risk pediatric neuroblastoma. Here, we demonstrate that MYC is also activated as a potent oncogene in a distinct subset of neuroblastoma cases through either focal amplification of distal enhancers or enhancer hijacking mediated by chromosomal translocation. Cancer Discov; 8(3); 320–35. \textcopyright 2017 AACR. This article is highlighted in the In This Issue feature, p. 253

  6. Retinal Cell Type DNA Methylation and Histone Modifications Predict Reprogramming Efficiency and Retinogenesis in 3D Organoid Cultures
    Wang Lu*, Hiler Daniel*, Xu Beisi*, AlDiri Issam, Chen Xiang, Zhou Xin, Griffiths Lyra, Valentine Marc, Shirinifard Abbas, Sablauer András, Thiagarajan Suresh, Barabas Marie-Elizabeth, Zhang Jiakun, Johnson Dianna, Frase Sharon, Dyer Michael A.
    Cell Reports 2018

    Summary Diverse cell types can be reprogrammed into pluripotent stem cells by ectopic expression of Oct4 (Pou5f1), Klf4, Sox2, and Myc. Many of these induced pluripotent stem cells (iPSCs) retain memory, in terms of DNA methylation and histone modifications (epigenetic memory), of their cellular origins, and this may bias subsequent differentiation. Neurons are difficult to reprogram, and there has not been a systematic side-by-side characterization of reprogramming efficiency or epigenetic memory across different neuronal subtypes. Here, we compare reprogramming efficiency of five different retinal cell types at two different stages of development. Retinal differentiation from each iPSC line was measured using a quantitative standardized scoring system called STEM-RET and compared to the epigenetic memory. Neurons with the lowest reprogramming efficiency produced iPSC lines with the best retinal differentiation and were more likely to retain epigenetic memory of their cellular origins. In addition, we identified biomarkers of iPSCs that are predictive of retinal differentiation.

2017:2

  1. Orthotopic Patient-Derived Xenografts of Paediatric Solid Tumours
    Stewart Elizabeth, Federico Sara M., Chen Xiang, Shelat Anang A., Bradley Cori, Gordon Brittney, Karlstrom Asa, Twarog Nathaniel R., Clay Michael R., Bahrami Armita, Iii Burgess B. Freeman, Xu Beisi, Zhou Xin, Wu Jianrong, Honnell Victoria, Ocarz Monica, Blankenship Kaley, Dapper Jason, Mardis Elaine R., Wilson Richard K., Downing James, Zhang Jinghui, Easton John, Pappo Alberto, Dyer Michael A.
    Nature 2017

    Preclinical models of paediatric solid tumours that could help identify predictive biomarkers of a patient’s sensitivity to therapy have been lacking. Over five years, the authors have developed an open access collection of orthotopic xenografts of 12 types of paediatric tumour. Genomic and epigenetic characterization reveals that xenografts retain characteristics of the tumour of origin. A high-throughput drug screen provides a resource for the community to identify potentially efficacious drug combinations.

  2. The Dynamic Epigenetic Landscape of the Retina During Development, Reprogramming, and Tumorigenesis
    Aldiri Issam*, Xu Beisi*, Wang Lu, Chen Xiang, Hiler Daniel, Griffiths Lyra, Valentine Marc, Shirinifard Abbas, Thiagarajan Suresh, Sablauer Andras, Barabas Marie-Elizabeth, Zhang Jiakun, Johnson Dianna, Frase Sharon, Zhou Xin, Easton John, Zhang Jinghui, Mardis Elaine R., Wilson Richard K., Downing James R., Dyer Michael A.
    Neuron 2017

    In the developing retina, multipotent neural progenitors undergo unidirectional differentiation in a precise spatiotemporal order. Here we profile the epigenetic and transcriptional changes that occur during retinogenesis in mice and humans. Although some progenitor genes and cell cycle genes were epigenetically silenced during retinogenesis, the most dramatic change was derepression of cell-type-specific differentiation programs. We identified developmental-stage-specific super-enhancers and showed that most epigenetic changes are conserved in humans and mice. To determine how the epigenome changes during tumorigenesis and reprogramming, we performed integrated epigenetic analysis of murine and human retinoblastomas and induced pluripotent stem cells (iPSCs) derived from murine rod photoreceptors. The retinoblastoma epigenome mapped to the developmental stage when retinal progenitors switch from neurogenic to terminal patterns of cell division. The epigenome of retinoblastomas was more similar to that of the normal retina than that of retina-derived iPSCs, and we identified retina-specific epigenetic memory.

2016:3

  1. Deregulation of DUX4 and ERG in Acute Lymphoblastic Leukemia
    Zhang Jinghui, McCastlain Kelly, Yoshihara Hiroki, Xu Beisi, Chang Yunchao, Churchman Michelle L., Wu Gang, Li Yongjin, Wei Lei, Iacobucci Ilaria, Liu Yu, Qu Chunxu, Wen Ji, Edmonson Michael, Payne-Turner Debbie, Kaufmann Kerstin B., Takayanagi Shin-ichiro, Wienholds Erno, Waanders Esmé, Ntziachristos Panagiotis, Bakogianni Sofia, Wang Jingjing, Aifantis Iannis, Roberts Kathryn G., Ma Jing, Song Guangchun, Easton John, Mulder Heather L., Chen Xiang, Newman Scott, Ma Xiaotu, Rusch Michael, Gupta Pankaj, Boggs Kristy, Vadodaria Bhavin, Dalton James, Liu Yanling, Valentine Marcus L., Ding Li, Lu Charles, Fulton Robert S., Fulton Lucinda, Tabib Yashodhan, Ochoa Kerri, Devidas Meenakshi, Pei Deqing, Cheng Cheng, Yang Jun, Evans William E., Relling Mary V., Pui Ching-Hon, Jeha Sima, Harvey Richard C., Chen I.-Ming L., Willman Cheryl L., Marcucci Guido, Bloomfield Clara D., Kohlschmidt Jessica, Mrózek Krzysztof, Paietta Elisabeth, Tallman Martin S., Stock Wendy, Foster Matthew C., Racevskis Janis, Rowe Jacob M., Luger Selina, Kornblau Steven M., Shurtleff Sheila A., Raimondi Susana C., Mardis Elaine R., Wilson Richard K., Dick John E., Hunger Stephen P., Loh Mignon L., Downing James R., Mullighan Charles G., Project the St Jude Children’s Research Hospital-Washington University Pediatric Cancer Genome
    Nature Genetics 2016

    Chromosomal rearrangements deregulating hematopoietic transcription factors are common in acute lymphoblastic leukemia (ALL). Here we show that deregulation of the homeobox transcription factor gene DUX4 and the ETS transcription factor gene ERG is a hallmark of a subtype of B-progenitor ALL that comprises up to 7% of B-ALL. DUX4 rearrangement and overexpression was present in all cases and was accompanied by transcriptional deregulation of ERG, expression of a novel ERG isoform, ERGalt, and frequent ERG deletion. ERGalt uses a non-canonical first exon whose transcription was initiated by DUX4 binding. ERGalt retains the DNA-binding and transactivation domains of ERG, but it inhibits wild-type ERG transcriptional activity and is transforming. These results illustrate a unique paradigm of transcription factor deregulation in leukemia in which DUX4 deregulation results in loss of function of ERG, either by deletion or induced expression of an isoform that is a dominant-negative inhibitor of wild-type ERG function.

  2. Binding of Estrogen Receptors to Switch Sites and Regulatory Elements in the Immunoglobulin Heavy Chain Locus of Activated B Cells Suggests a Direct Influence of Estrogen on Antibody Expression
    Jones Bart G., Penkert Rhiannon R., Xu Beisi, Fan Yiping, Neale Geoff, Gearhart Patricia J., Hurwitz Julia L.
    Molecular Immunology 2016

    Females and males differ in antibody isotype expression patterns and in immune responses to foreign- and self-antigens. For example, systemic lupus erythematosus is a condition that associates with the production of isotype-skewed anti-self antibodies, and exhibits a 9:1 female:male disease ratio. To explain differences between B cell responses in males and females, we sought to identify direct interactions of the estrogen receptor (ER) with the immunoglobulin heavy chain locus. This effort was encouraged by our previous identification of estrogen response elements (ERE) in heavy chain switch (S) regions. We conducted a full-genome chromatin immunoprecipitation analysis (ChIP-seq) using DNA from LPS-activated B cells and an ERα-specific antibody. Results revealed ER binding to a wide region of DNA, spanning sequences from the JH cluster to Cδ, with peaks in Eμ and Sμ sites. Additional peaks of ERα binding were coincident with hs1,2 and hs4 sites in the 3’ regulatory region (3’RR) of the heavy chain locus. This first demonstration of direct binding of ER to key regulatory elements in the immunoglobulin locus supports our hypothesis that estrogen and other nuclear hormone receptors and ligands may directly influence antibody expression and class switch recombination (CSR). Our hypothesis encourages the conduct of new experiments to evaluate the consequences of ER binding. A better understanding of ER:DNA interactions in the immunoglobulin heavy chain locus, and respective mechanisms, may ultimately translate to better control of antibody expression, better protection against pathogens, and prevention of pathologies caused by auto-immune disease.

  3. Hotspots for Vitamin–Steroid–Thyroid Hormone Response Elements Within Switch Regions of Immunoglobulin Heavy Chain Loci Predict a Direct Influence of Vitamins and Hormones on B Cell Class Switch Recombination
    Hurwitz Julia L., Penkert Rhiannon R., Xu Beisi, Fan Yiping, Partridge Janet F., Maul Robert W., Gearhart Patricia J.
    Viral Immunology 2016

    Vitamin A deficiencies are common throughout the world and have a significant negative influence on immune protection against viral infections. Mouse models demonstrate that the production of IgA, a first line of defense against viruses at mucosal sites, is inhibited in the context of vitamin A deficiency. In vitro, the addition of vitamin A to activated B cells can enhance IgA expression, but downregulate IgE. Previous reports have demonstrated that vitamin A modifies cytokine patterns, and in so doing may influence antibody isotype expression by an indirect mechanism. However, we have now discovered hundreds of potential response elements among Sμ, S\varepsilon, and Sα switch sites within immunoglobulin heavy chain loci. These hotspots appear in both mouse and human loci and include targets for vitamin receptors and related proteins (e.g., estrogen receptors) in the nuclear receptor superfamily. Full response elements with direct repeats are relatively infrequent or absent in Sγ regions although half-sites are present. Based on these results, we pose a hypothesis that nuclear receptors have a direct effect on the immunoglobulin heavy chain class switch recombination event. We propose that vitamin A may alter S site accessibility to activation-induced deaminase and nonhomologous end-joining machinery, thereby influencing the isotype switch, antibody production, and protection against viral infections at mucosal sites.

2015:3

  1. Brg1 Coordinates Multiple Processes during Retinogenesis and Is a Tumor Suppressor in Retinoblastoma
    Aldiri Issam, Ajioka Itsuki, Xu Beisi, Zhang Jiakun, Chen Xiang, Benavente Claudia, Finkelstein David, Johnson Dianna, Akiyama Jennifer, Pennacchio Len A., Dyer Michael A.
    Development 2015

    Skip to Next Section Retinal development requires precise temporal and spatial coordination of cell cycle exit, cell fate specification, cell migration and differentiation. When this process is disrupted, retinoblastoma, a developmental tumor of the retina, can form. Epigenetic modulators are central to precisely coordinating developmental events, and many epigenetic processes have been implicated in cancer. Studying epigenetic mechanisms in development is challenging because they often regulate multiple cellular processes; therefore, elucidating the primary molecular mechanisms involved can be difficult. Here we explore the role of Brg1 (Smarca4) in retinal development and retinoblastoma in mice using molecular and cellular approaches. Brg1 was found to regulate retinal size by controlling cell cycle length, cell cycle exit and cell survival during development. Brg1 was not required for cell fate specification but was required for photoreceptor differentiation and cell adhesion/polarity programs that contribute to proper retinal lamination during development. The combination of defective cell differentiation and lamination led to retinal degeneration in Brg1-deficient retinae. Despite the hypocellularity, premature cell cycle exit, increased cell death and extended cell cycle length, retinal progenitor cells persisted in Brg1-deficient retinae, making them more susceptible to retinoblastoma. ChIP-Seq analysis suggests that Brg1 might regulate gene expression through multiple mechanisms.

  2. The Glucose-Sensing Transcription Factor MLX Promotes Myogenesis via Myokine Signaling
    Hunt Liam C., Xu Beisi, Finkelstein David, Fan Yiping, Carroll Patrick A., Cheng Pei-Feng, Eisenman Robert N., Demontis Fabio
    Genes & Development 2015

    Metabolic stress and changes in nutrient levels modulate many aspects of skeletal muscle function during aging and disease. Growth factors and cytokines secreted by skeletal muscle, known as myokines, are important signaling factors, but it is largely unknown whether they modulate muscle growth and differentiation in response to nutrients. Here, we found that changes in glucose levels increase the activity of the glucose-responsive transcription factor MLX (Max-like protein X), which promotes and is necessary for myoblast fusion. MLX promotes myogenesis not via an adjustment of glucose metabolism but rather by inducing the expression of several myokines, including insulin-like growth factor 2 (IGF2), whereas RNAi and dominant-negative MLX reduce IGF2 expression and block myogenesis. This phenotype is rescued by conditioned medium from control muscle cells and by recombinant IGF2, which activates the myogenic kinase Akt. Importantly, MLX-null mice display decreased IGF2 induction and diminished muscle regeneration in response to injury, indicating that the myogenic function of MLX is manifested in vivo. Thus, glucose is a signaling molecule that regulates myogenesis and muscle regeneration via MLX/IGF2/Akt signaling.

  3. Quantification of Retinogenesis in 3D Cultures Reveals Epigenetic Memory and Higher Efficiency in iPSCs Derived from Rod Photoreceptors
    Hiler Daniel, Chen Xiang, Hazen Jennifer, Kupriyanov Sergey, Carroll Patrick A., Qu Chunxu, Xu Beisi, Johnson Dianna, Griffiths Lyra, Frase Sharon, Rodriguez Alberto R., Martin Greg, Zhang Jiakun, Jeon Jongrye, Fan Yiping, Finkelstein David, Eisenman Robert N., Baldwin Kristin, Dyer Michael A.
    Cell Stem Cell 2015

    Summary Cell-based therapies to treat retinal degeneration are now being tested in clinical trials. However, it is not known whether the source of stem cells is important for the production of differentiated cells suitable for transplantation. To test this, we generated induced pluripotent stem cells (iPSCs) from murine rod photoreceptors (r-iPSCs) and scored their ability to make retinae by using a standardized quantitative protocol called STEM-RET. We discovered that r-iPSCs more efficiently produced differentiated retinae than did embryonic stem cells (ESCs) or fibroblast-derived iPSCs (f-iPSCs). Retinae derived from f-iPSCs had fewer amacrine cells and other inner nuclear layer cells. Integrated epigenetic analysis showed that DNA methylation contributes to the defects in f-iPSC retinogenesis and that rod-specific CTCF insulator protein-binding sites may promote r-iPSC retinogenesis. Together, our data suggest that the source of stem cells is important for producing retinal neurons in three-dimensional (3D) organ cultures.