Regulation of H2B Monoubiquitination Pathway in Breast cancer

H2B monoubiquitination (H2Bub1) regulation via CDK9-WAC-RNF20/40 axis has been well illustrated. It is interesting to determine the upstream regulators which dictate the process of monoubiquitination of H2B. On further investigation, histone chaperone SUPT6H known to bind P-Ser2 RNAPII was observed to regulate ERα-dependent signaling through H2Bub1 pathway. Perturbation of SUPT6H led to a decrease in H2Bub1 resulting in impaired estrogen-dependent signaling and mesenchymal stem cell differentiation due to increase in the H3K27me3 repressive mark on the promoters of the genes. Moreover, SUPT6H levels were decreased with tumor progression. Together, these data identify SUPT6H as a new epigenetic regulator of ERα activity and cellular differentiation. Further upstream regulator of CDK9, BRD4 was also examined. For the first time, we showed the connection between BRD4 and H2Bub1 pathway. They both regulated the gene expression in a similar fashion and had gene expression-dependent occupancy on the genes. Interruption of this pathway by BRD4 or H2Bub1 depletion resulted in acquisition of EMT and stem cell-like phenotype in mammary epithelial cells. The preliminary data for conditional RNF40 KO in mammary gland also showed increased mammary branching further emphasizing the critical role of H2Bub1 as a tumor suppressor. These important findings could help to harness these epigenetic regulators for anti-cancer therapy.

[1]  Xin Hu,et al.  Requirement of the histone demethylase LSD1 in Snai1-mediated transcriptional repression during epithelial-mesenchymal transition , 2010, Oncogene.

[2]  R. Weinberg,et al.  Transitions between epithelial and mesenchymal states: acquisition of malignant and stem cell traits , 2009, Nature Reviews Cancer.

[3]  Nevan J. Krogan,et al.  COMPASS: A complex of proteins associated with a trithorax-related SET domain protein , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[4]  T. Beißbarth,et al.  The histone H2B monoubiquitination regulatory pathway is required for differentiation of multipotent stem cells. , 2012, Molecular cell.

[5]  B. O’Malley,et al.  Minireview: steroid receptor coactivator-3: a multifarious coregulator in mammary gland metastasis. , 2011, Endocrinology.

[6]  Lee E. Edsall,et al.  Distinct epigenomic landscapes of pluripotent and lineage-committed human cells. , 2010, Cell stem cell.

[7]  A. Ciechanover,et al.  The ubiquitin-proteasome proteolytic pathway: destruction for the sake of construction. , 2002, Physiological reviews.

[8]  R. Spang,et al.  Gene Expression Signatures for Tumor Progression, Tumor Subtype, and Tumor Thickness in Laser-Microdissected Melanoma Tissues , 2007, Clinical Cancer Research.

[9]  F. Winston,et al.  Evidence That Spt6p Controls Chromatin Structure by a Direct Interaction with Histones , 1996, Science.

[10]  C. Korch,et al.  Global Decrease of Histone H3K27 Acetylation in ZEB1-Induced Epithelial to Mesenchymal Transition in Lung Cancer Cells , 2013, Cancers.

[11]  Jerard Hurwitz,et al.  A Mammalian Bromodomain Protein, Brd4, Interacts with Replication Factor C and Inhibits Progression to S Phase , 2002, Molecular and Cellular Biology.

[12]  I. Haviv,et al.  Epigenetic Regulation of Cell Type–Specific Expression Patterns in the Human Mammary Epithelium , 2011, PLoS genetics.

[13]  P. D. Dal Cin,et al.  BRD4 bromodomain gene rearrangement in aggressive carcinoma with translocation t(15;19). , 2001, The American journal of pathology.

[14]  Chi-Hung Huang,et al.  Bmi1 is essential in Twist1-induced epithelial–mesenchymal transition , 2010, Nature Cell Biology.

[15]  N. Pedersen,et al.  Analysis of the epidermal growth factor receptor specific transcriptome: Effect of receptor expression level and an activating mutation , 2005, Journal of cellular biochemistry.

[16]  D. Reinberg,et al.  Human Spt6 Stimulates Transcription Elongation by RNA Polymerase II In Vitro , 2004, Molecular and Cellular Biology.

[17]  Piotr J. Balwierz,et al.  Sox4 is a master regulator of epithelial-mesenchymal transition by controlling Ezh2 expression and epigenetic reprogramming. , 2013, Cancer cell.

[18]  Julia Schüler,et al.  The EMT-activator ZEB1 promotes tumorigenicity by repressing stemness-inhibiting microRNAs , 2009, Nature Cell Biology.

[19]  James A. Cuff,et al.  A Bivalent Chromatin Structure Marks Key Developmental Genes in Embryonic Stem Cells , 2006, Cell.

[20]  Sun-Mi Park,et al.  The miR-200 family determines the epithelial phenotype of cancer cells by targeting the E-cadherin repressors ZEB1 and ZEB2. , 2008, Genes & development.

[21]  C. Antonescu,et al.  Midline carcinoma of children and young adults with NUT rearrangement. , 2004, Journal of clinical oncology : official journal of the American Society of Clinical Oncology.

[22]  Shwu‐Yuan Wu,et al.  Brd4 links chromatin targeting to HPV transcriptional silencing. , 2006, Genes & development.

[23]  S. Sif.  ATP‐dependent nucleosome remodeling complexes: Enzymes tailored to deal with chromatin , 2004, Journal of cellular biochemistry.

[24]  A. Wolffe,et al.  Acetylation of general transcription factors by histone acetyltransferases , 1997, Current Biology.

[25]  D. Raleigh,et al.  Histone H2B ubiquitylation disrupts local and higher order chromatin compaction , 2010, Nature chemical biology.

[26]  M. Osley,et al.  H2B ubiquitylation plays a role in nucleosome dynamics during transcription elongation. , 2008, Molecular cell.

[27]  M. Parker,et al.  The antiestrogen ICI 182780 disrupts estrogen receptor nucleocytoplasmic shuttling. , 1993, Journal of cell science.

[28]  B. Peterlin,et al.  Controlling the elongation phase of transcription with P-TEFb. , 2006, Molecular cell.

[29]  Patrick Cramer,et al.  Structure and in vivo requirement of the yeast Spt6 SH2 domain. , 2009, Journal of molecular biology.

[30]  P. Pandolfi,et al.  The oncogenic microRNA miR-22 targets the TET2 tumor suppressor to promote hematopoietic stem cell self-renewal and transformation. , 2013, Cell stem cell.

[31]  R. Roeder,et al.  The human homolog of yeast BRE1 functions as a transcriptional coactivator through direct activator interactions. , 2005, Molecular cell.

[32]  P. Argos,et al.  Human oestrogen receptor cDNA: sequence, expression and homology to v-erb-A , 1986, Nature.

[33]  C. Glass,et al.  Functional roles of enhancer RNAs for oestrogen-dependent transcriptional activation , 2013, Nature.

[34]  R. Tibshirani,et al.  Gene expression patterns of breast carcinomas distinguish tumor subclasses with clinical implications , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[35]  A. Ruifrok,et al.  Quantification of histochemical staining by color deconvolution. , 2001, Analytical and quantitative cytology and histology.

[36]  S. Johnsen.  The enigmatic role of H2Bub1 in cancer , 2012, FEBS letters.

[37]  William B. Smith,et al.  Selective inhibition of BET bromodomains , 2010, Nature.

[38]  Ming-Ming Zhou,et al.  Structure and mechanisms of lysine methylation recognition by the chromodomain in gene transcription. , 2011, Biochemistry.

[39]  Hiroshi Handa,et al.  P-TEFb-mediated phosphorylation of hSpt5 C-terminal repeats is critical for processive transcription elongation. , 2006, Molecular cell.

[40]  P. Chambon,et al.  Modulation of transcriptional activation by ligand‐dependent phosphorylation of the human oestrogen receptor A/B region. , 1993, The EMBO journal.

[41]  D. Faller,et al.  SIRT1 induces EMT by cooperating with EMT transcription factors and enhances prostate cancer cell migration and metastasis , 2012, Oncogene.

[42]  H. Huber,et al.  A crucial function of PDGF in TGF-β-mediated cancer progression of hepatocytes , 2006, Oncogene.

[43]  C. Peterson,et al.  The SWI-SNF complex: a chromatin remodeling machine? , 1995, Trends in biochemical sciences.

[44]  Chawnshang Chang,et al.  Androgen Receptor Interacts with the Positive Elongation Factor P-TEFb and Enhances the Efficiency of Transcriptional Elongation* , 2001, The Journal of Biological Chemistry.

[45]  K. Korach,et al.  Estrogen receptor transcription and transactivation Estrogen receptor knockout mice: what their phenotypes reveal about mechanisms of estrogen action , 2000 .

[46]  S. Johnsen,et al.  Insights into the function of the human P-TEFb component CDK9 in the regulation of chromatin modifications and co-transcriptional mRNA processing , 2009, Cell cycle.

[47]  Robert A. Weinberg,et al.  Poised Chromatin at the ZEB1 Promoter Enables Breast Cancer Cell Plasticity and Enhances Tumorigenicity , 2013, Cell.

[48]  G. Almouzni,et al.  Histone chaperones, a supporting role in the limelight. , 2004, Biochimica et biophysica acta.

[49]  K. Helin,et al.  Polycomb Complex 2 Is Required for E-cadherin Repression by the Snail1 Transcription Factor , 2008, Molecular and Cellular Biology.

[50]  M. Hochstrasser.  Ubiquitin-dependent protein degradation. , 1996, Annual review of genetics.

[51]  Heike Brand,et al.  Estrogen Receptor-α Directs Ordered, Cyclical, and Combinatorial Recruitment of Cofactors on a Natural Target Promoter , 2003, Cell.

[52]  J. Lippincott-Schwartz,et al.  A Bromodomain Protein, MCAP, Associates with Mitotic Chromosomes and Affects G2-to-M Transition , 2000, Molecular and Cellular Biology.

[53]  D. Faller,et al.  You bet-cha: a novel family of transcriptional regulators. , 2001, Frontiers in bioscience : a journal and virtual library.

[54]  N. Avvakumov,et al.  Histone chaperones: modulators of chromatin marks. , 2011, Molecular cell.

[55]  T. Fujita,et al.  Inhibition of histone deacetylase activity suppresses epithelial-to-mesenchymal transition induced by TGF-beta1 in human renal epithelial cells. , 2007, Journal of the American Society of Nephrology : JASN.

[56]  V. Theodorou,et al.  GATA3 acts upstream of FOXA1 in mediating ESR1 binding by shaping enhancer accessibility , 2013, Genome research.

[57]  Wenlin Huang,et al.  The polycomb group protein Bmi-1 represses the tumor suppressor PTEN and induces epithelial-mesenchymal transition in human nasopharyngeal epithelial cells. , 2009, The Journal of clinical investigation.

[58]  H. Cedar,et al.  Linking DNA methylation and histone modification: patterns and paradigms , 2009, Nature Reviews Genetics.

[59]  Leighton J. Core,et al.  Postrecruitment Regulation of RNA Polymerase II Directs Rapid Signaling Responses at the Promoters of Estrogen Target Genes , 2008, Molecular and Cellular Biology.

[60]  P. Kaufman.  Nucleosome assembly: the CAF and the HAT. , 1996, Current opinion in cell biology.

[61]  A. Link,et al.  Inhibition of P-TEFb (CDK9/Cyclin T) kinase and RNA polymerase II transcription by the coordinated actions of HEXIM1 and 7SK snRNA. , 2003, Molecular cell.

[62]  A. G. Herreros,et al.  The transcription factor Snail is a repressor of E-cadherin gene expression in epithelial tumour cells , 2000, Nature Cell Biology.

[63]  Adam A. Margolin,et al.  The Cancer Cell Line Encyclopedia enables predictive modeling of anticancer drug sensitivity , 2012, Nature.

[64]  S. Berger,et al.  The histone H2B-specific ubiquitin ligase RNF20/hBRE1 acts as a putative tumor suppressor through selective regulation of gene expression. , 2008, Genes & development.

[65]  A. Dimmler,et al.  The transcriptional repressor ZEB1 promotes metastasis and loss of cell polarity in cancer. , 2008, Cancer research.

[66]  M. Grunstein.  Histone acetylation in chromatin structure and transcription , 1997, Nature.

[67]  M. Ittmann,et al.  SRC-3 is required for prostate cancer cell proliferation and survival. , 2005, Cancer research.

[68]  L. Tora,et al.  The Tightly Controlled Deubiquitination Activity of the Human SAGA Complex Differentially Modifies Distinct Gene Regulatory Elements , 2011, Molecular and Cellular Biology.

[69]  T. Kouzarides.  Chromatin Modifications and Their Function , 2007, Cell.

[70]  J. Taylor‐Papadimitriou,et al.  Mammosphere culture of metastatic breast cancer cells enriches for tumorigenic breast cancer cells , 2008, Breast Cancer Research.

[71]  Pier Paolo Di Fiore,et al.  Human USP3 Is a Chromatin Modifier Required for S Phase Progression and Genome Stability , 2007, Current Biology.

[72]  J. Gustafsson,et al.  Role of estrogen receptor beta in estrogen action. , 2001, Annual review of physiology.

[73]  Jun Yao,et al.  G9a interacts with Snail and is critical for Snail-mediated E-cadherin repression in human breast cancer. , 2012, The Journal of clinical investigation.

[74]  V. Sartorelli,et al.  The histone chaperone Spt6 coordinates histone H3K27 demethylation and myogenesis , 2013, The EMBO journal.

[75]  R. Shiekhattar,et al.  Requirement for SNAPC1 in Transcriptional Responsiveness to Diverse Extracellular Signals , 2012, Molecular and Cellular Biology.

[76]  Gang Li,et al.  Regulation of p53 by ING family members in suppression of tumor initiation and progression , 2011, Cancer and Metastasis Reviews.

[77]  A. Postigo,et al.  ZEB1 represses E-cadherin and induces an EMT by recruiting the SWI/SNF chromatin-remodeling protein BRG1 , 2010, Oncogene.

[78]  T. Jensen,et al.  Telomerase expression extends the proliferative life-span and maintains the osteogenic potential of human bone marrow stromal cells , 2002, Nature Biotechnology.

[79]  J. Crabtree,et al.  Bidirectional transcriptional activity of PGK‐neomycin and unexpected embryonic lethality in heterozygote chimeric knockout mice , 2001, Genesis.

[80]  A. Giordano,et al.  Abrogation of signal-dependent activation of the cdk9/cyclin T2a complex in human RD rhabdomyosarcoma cells , 2007, Cell Death and Differentiation.

[81]  Vladimir Benes,et al.  Transient cyclical methylation of promoter DNA , 2008, Nature.

[82]  M. Korpal,et al.  The miR-200 Family Inhibits Epithelial-Mesenchymal Transition and Cancer Cell Migration by Direct Targeting of E-cadherin Transcriptional Repressors ZEB1 and ZEB2* , 2008, Journal of Biological Chemistry.

[83]  Kevin Truong,et al.  Cadherins in embryonic and neural morphogenesis , 2000, Nature Reviews Molecular Cell Biology.

[84]  N. Crawford,et al.  Bromodomain 4 activation predicts breast cancer survival , 2008, Proceedings of the National Academy of Sciences.

[85]  R. Evans,et al.  The Spt6 SH2 domain binds Ser2-P RNAPII to direct Iws1-dependent mRNA splicing and export. , 2007, Genes & development.

[86]  R. Weinberg,et al.  Cancer stem cells and epithelial-mesenchymal transition: concepts and molecular links. , 2012, Seminars in cancer biology.

[87]  Kevin Struhl,et al.  Evidence for Eviction and Rapid Deposition of Histones upon Transcriptional Elongation by RNA Polymerase II , 2004, Molecular and Cellular Biology.

[88]  J. Aster,et al.  BRD–NUT oncoproteins: a family of closely related nuclear proteins that block epithelial differentiation and maintain the growth of carcinoma cells , 2008, Oncogene.

[89]  Olivier Poch,et al.  Identification of genes associated with tumorigenesis and metastatic potential of hypopharyngeal cancer by microarray analysis , 2004, Oncogene.

[90]  K. Korach,et al.  Receptor null mice reveal contrasting roles for estrogen receptor α and β in reproductive tissues , 2000, The Journal of Steroid Biochemistry and Molecular Biology.

[91]  S. Lowe,et al.  RNAi screen identifies Brd4 as a therapeutic target in acute myeloid leukaemia , 2011, Nature.

[92]  U. K. Laemmli,et al.  Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4 , 1970, Nature.

[93]  T. Wada,et al.  SPT genes: key players in the regulation of transcription, chromatin structure and other cellular processes. , 2001, Journal of biochemistry.

[94]  J. Qin,et al.  A Proteomic Analysis of Ataxia Telangiectasia-mutated (ATM)/ATM-Rad3-related (ATR) Substrates Identifies the Ubiquitin-Proteasome System as a Regulator for DNA Damage Checkpoints* , 2007, Journal of Biological Chemistry.

[95]  V. Orlando.  Polycomb, Epigenomes, and Control of Cell Identity , 2003, Cell.

[96]  S. Johnsen.  CDK9 and H2B Monoubiquitination: A Well-Choreographed Dance , 2012, PLoS genetics.

[97]  Van Trung Nguyen,et al.  Binding of the 7SK snRNA turns the HEXIM1 protein into a P‐TEFb (CDK9/cyclin T) inhibitor , 2004, The EMBO journal.

[98]  I B Dawid,et al.  The bromodomain: a conserved sequence found in human, Drosophila and yeast proteins. , 1992, Nucleic acids research.

[99]  K. Korach,et al.  Estrogen receptor knockout mice: phenotypes in the female reproductive tract , 2003, Gynecological endocrinology : the official journal of the International Society of Gynecological Endocrinology.

[100]  I. Ellis,et al.  Expression of luminal and basal cytokeratins in human breast carcinoma , 2004, The Journal of pathology.

[101]  Paula D. Bos,et al.  Modeling metastasis in the mouse. , 2010, Current opinion in pharmacology.

[102]  Vinay Varadan,et al.  DNA methylation patterns in luminal breast cancers differ from non‐luminal subtypes and can identify relapse risk independent of other clinical variables , 2011, Molecular oncology.

[103]  R. Young,et al.  BET Bromodomain Inhibition as a Therapeutic Strategy to Target c-Myc , 2011, Cell.

[104]  I. Issaeva,et al.  UTX and JMJD3 are histone H3K27 demethylases involved in HOX gene regulation and development , 2007, Nature.

[105]  Clifford A. Meyer,et al.  Cistrome: an integrative platform for transcriptional regulation studies , 2011, Genome Biology.

[106]  D. Reinberg,et al.  Histone H2B Monoubiquitination Functions Cooperatively with FACT to Regulate Elongation by RNA Polymerase II , 2006, Cell.

[107]  T. Jenuwein,et al.  The many faces of histone lysine methylation. , 2002, Current opinion in cell biology.

[108]  K. Pantel,et al.  Micrometastasis in breast cancer and other solid tumors. , 2004, Journal of biological regulators and homeostatic agents.

[109]  D. Bartel.  MicroRNAs Genomics, Biogenesis, Mechanism, and Function , 2004, Cell.

[110]  J. Lees,et al.  Identification of a conserved region required for hormone dependent transcriptional activation by steroid hormone receptors. , 1992, The EMBO journal.

[111]  F. Winston,et al.  Noncanonical Tandem SH2 Enables Interaction of Elongation Factor Spt6 with RNA Polymerase II* , 2010, The Journal of Biological Chemistry.

[112]  Clifford A. Meyer,et al.  Genome-wide analysis of estrogen receptor binding sites , 2006, Nature Genetics.

[113]  D. Armant,et al.  Epigenetics: Definition, Mechanisms and Clinical Perspective , 2009, Seminars in reproductive medicine.

[114]  P. Eilers,et al.  E-cadherin transcriptional downregulation by promoter methylation but not mutation is related to epithelial-to-mesenchymal transition in breast cancer cell lines , 2006, British Journal of Cancer.

[115]  M. Montano,et al.  The breast cell growth inhibitor, estrogen down regulated gene 1, modulates a novel functional interaction between estrogen receptor alpha and transcriptional elongation factor cyclin T1 , 2005, Oncogene.

[116]  D. Picard,et al.  Activation of the unliganded estrogen receptor by EGF involves the MAP kinase pathway and direct phosphorylation. , 1996, The EMBO journal.

[117]  M. Colombo,et al.  Inhibiting interactions of lysine demethylase LSD1 with snail/slug blocks cancer cell invasion. , 2013, Cancer research.

[118]  Philippe Soriano,et al.  Widespread recombinase expression using FLPeR (Flipper) mice , 2000, Genesis.

[119]  A. Rice,et al.  Antiapoptotic Function of Cdk9 (TAK/P-TEFb) in U937 Promonocytic Cells , 2001, Journal of Virology.

[120]  I. Weissman,et al.  Stem cells, cancer, and cancer stem cells , 2001, Nature.

[121]  A. Lustig,et al.  Coupling of Glucose Deprivation with Impaired Histone H2B Monoubiquitination in Tumors , 2012, PloS one.

[122]  D. Teng,et al.  BRG1, a component of the SWI-SNF complex, is mutated in multiple human tumor cell lines. , 2000, Cancer research.

[123]  R. Beddington,et al.  Growth and Early Postimplantation Defects in Mice Deficient for the Bromodomain-Containing Protein Brd4 , 2002, Molecular and Cellular Biology.

[124]  Pengbo Liu,et al.  Sex-determining region Y box 4 is a transforming oncogene in human prostate cancer cells. , 2006, Cancer research.

[125]  Shridar Ganesan,et al.  X chromosomal abnormalities in basal-like human breast cancer. , 2006, Cancer cell.

[126]  Bing Li,et al.  The Role of Chromatin during Transcription , 2007, Cell.

[127]  B. Peterlin,et al.  Dynamics of Human Immunodeficiency Virus Transcription: P-TEFb Phosphorylates RD and Dissociates Negative Effectors from the Transactivation Response Element , 2004, Molecular and Cellular Biology.

[128]  C. Bountra,et al.  Epigenetic protein families: a new frontier for drug discovery , 2012, Nature Reviews Drug Discovery.

[129]  Kou-Juey Wu,et al.  Interplay between HDAC3 and WDR5 is essential for hypoxia-induced epithelial-mesenchymal transition. , 2011, Molecular cell.

[130]  L. Hennighausen,et al.  Cre-mediated gene deletion in the mammary gland. , 1997, Nucleic acids research.

[131]  Daniel Chourrout,et al.  Genome Regulation by Polycomb and Trithorax Proteins , 2007, Cell.

[132]  R. Jaenisch,et al.  TET1 plays an essential oncogenic role in MLL-rearranged leukemia , 2013, Proceedings of the National Academy of Sciences.

[133]  Carlos S. Moreno,et al.  MTA3, a Mi-2/NuRD Complex Subunit, Regulates an Invasive Growth Pathway in Breast Cancer , 2003, Cell.

[134]  F. Jeanmougin,et al.  The bromodomain revisited. , 1997, Trends in biochemical sciences.

[135]  R. Kingston,et al.  The Core of the Polycomb Repressive Complex Is Compositionally and Functionally Conserved in Flies and Humans , 2002, Molecular and Cellular Biology.

[136]  R. Kingston,et al.  Chromatin Compaction by a Polycomb Group Protein Complex , 2004, Science.

[137]  Renato Paro,et al.  Epigenetic regulation of cellular memory by the Polycomb and Trithorax group proteins. , 2004, Annual review of genetics.

[138]  T. Kiss,et al.  Regulation of Polymerase II Transcription by 7SK snRNA: Two Distinct RNA Elements Direct P-TEFb and HEXIM1 Binding , 2006, Molecular and Cellular Biology.

[139]  M. Parker,et al.  Molecular mechanisms of steroid hormone action , 1998 .

[140]  V. Corces,et al.  Enhancer function: new insights into the regulation of tissue-specific gene expression , 2011, Nature Reviews Genetics.

[141]  B. Zhou,et al.  The SNAG domain of Snail1 functions as a molecular hook for recruiting lysine‐specific demethylase 1 , 2010, The EMBO journal.

[142]  H. Stunnenberg,et al.  A TFTC/STAGA module mediates histone H2A and H2B deubiquitination, coactivates nuclear receptors, and counteracts heterochromatin silencing. , 2008, Molecular cell.

[143]  H. Towbin,et al.  Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. , 1979, Proceedings of the National Academy of Sciences of the United States of America.

[144]  M. Meguid,et al.  Estrogen receptor expression in benign breast epithelium and breast cancer risk. , 1998, Journal of the National Cancer Institute.

[145]  Tom Misteli,et al.  The double bromodomain protein Brd4 binds to acetylated chromatin during interphase and mitosis , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[146]  A. Gill,et al.  The tumor suppressor CDC73 interacts with the ring finger proteins RNF20 and RNF40 and is required for the maintenance of histone 2B monoubiquitination. , 2012, Human molecular genetics.

[147]  P. Angrand,et al.  Targeted insertion results in a Rhombomere 2‐specific Hoxa2 knockdown and ectopic activation of Hoxa1 expression , 2002, Developmental dynamics : an official publication of the American Association of Anatomists.

[148]  Yi Zhang,et al.  Tudor, MBT and chromo domains gauge the degree of lysine methylation , 2006, EMBO reports.

[149]  Qiang Zhou,et al.  Recruitment of P-TEFb for stimulation of transcriptional elongation by the bromodomain protein Brd4. , 2005, Molecular cell.

[150]  H. Tauchi,et al.  Regulation of homologous recombination by RNF20-dependent H2B ubiquitination. , 2011, Molecular cell.

[151]  J. Brown,et al.  R-loops and genomic instability in Bre1 (RNF20/40)-deficient cells , 2012, Cell cycle.

[152]  R Ohba,et al.  Yeast Gcn5 functions in two multisubunit complexes to acetylate nucleosomal histones: characterization of an Ada complex and the SAGA (Spt/Ada) complex. , 1997, Genes & development.

[153]  F. Pépin,et al.  Stromal gene expression predicts clinical outcome in breast cancer , 2008, Nature Medicine.

[154]  C. Allis,et al.  Translating the Histone Code , 2001, Science.

[155]  W. Birchmeier,et al.  Cadherin expression in carcinomas: role in the formation of cell junctions and the prevention of invasiveness. , 1994, Biochimica et biophysica acta.

[156]  A. Muñoz,et al.  The expression levels of the transcriptional regulators p300 and CtBP modulate the correlations between SNAIL, ZEB1, E‐cadherin and vitamin D receptor in human colon carcinomas , 2006, International journal of cancer.

[157]  M. Karin,et al.  A conserved C-terminal sequence that is deleted in v-ErbA is essential for the biological activities of c-ErbA (the thyroid hormone receptor) , 1993, Molecular and cellular biology.

[158]  X. Chen,et al.  Identification and analysis of the human and murine putative chromatin structure regulator SUPT6H and Supt6h. , 1996, Genomics.

[159]  S. Gygi,et al.  Profiling of UV-induced ATM/ATR signaling pathways , 2007, Proceedings of the National Academy of Sciences.

[160]  S. Amente,et al.  The histone LSD1 demethylase in stemness and cancer transcription programs. , 2013, Biochimica et biophysica acta.

[161]  S. Sleijfer,et al.  Decreased expression of EZH2 is associated with upregulation of ER and favorable outcome to tamoxifen in advanced breast cancer , 2010, Breast Cancer Research and Treatment.

[162]  S. Gygi,et al.  The histone H 3 Lys 27 demethylase JMJD 3 regulates gene expression by impacting transcriptional elongation , 2012 .

[163]  Lei Zhou,et al.  The Malignant Brain Tumor (MBT) Domain Protein SFMBT1 Is an Integral Histone Reader Subunit of the LSD1 Demethylase Complex for Chromatin Association and Epithelial-to-mesenchymal Transition* , 2013, The Journal of Biological Chemistry.

[164]  F. Winston,et al.  Evidence that Spt4, Spt5, and Spt6 control transcription elongation by RNA polymerase II in Saccharomyces cerevisiae. , 1998, Genes & development.

[165]  Stephan Sauer,et al.  Chromatin signatures of pluripotent cell lines , 2006, Nature Cell Biology.

[166]  B. Stillman,et al.  Stepwise assembly of chromatin during DNA replication in vitro. , 1991, The EMBO journal.

[167]  A. Shilatifard,et al.  RAD6-Mediated Transcription-Coupled H2B Ubiquitylation Directly Stimulates H3K4 Methylation in Human Cells , 2009, Cell.

[168]  Claude C. Warzecha,et al.  The putative cancer stem cell marker USP22 is a subunit of the human SAGA complex required for activated transcription and cell-cycle progression. , 2008, Molecular cell.

[169]  H. Neumann,et al.  The H2B ubiquitin ligase RNF40 cooperates with SUPT16H to induce dynamic changes in chromatin structure during DNA double-strand break repair , 2011, Cell cycle.

[170]  Wenjun Guo,et al.  The Epithelial-Mesenchymal Transition Generates Cells with Properties of Stem Cells , 2008, Cell.

[171]  M. Parker.  Structure and function of estrogen receptors. , 1995, Vitamins and hormones.

[172]  Danny Reinberg,et al.  Histones: annotating chromatin. , 2009, Annual review of genetics.

[173]  Cun-Yu Wang,et al.  Histone Demethylase KDM6B Promotes Epithelial-Mesenchymal Transition* , 2012, The Journal of Biological Chemistry.

[174]  Shona Murphy,et al.  Cracking the RNA polymerase II CTD code. , 2008, Trends in genetics : TIG.

[175]  S. Fawell,et al.  Identification of two transactivation domains in the mouse oestrogen receptor. , 1989, Nucleic acids research.

[176]  J. Pink,et al.  HEXIM1 is a critical determinant of the response to tamoxifen , 2011, Oncogene.

[177]  Danila Coradini,et al.  Isolation and in vitro propagation of tumorigenic breast cancer cells with stem/progenitor cell properties. , 2005, Cancer research.

[178]  Jianguo Song,et al.  Histone deacetylase 1 is required for transforming growth factor-beta1-induced epithelial-mesenchymal transition. , 2010, The international journal of biochemistry & cell biology.

[179]  Yan-Long Liu,et al.  USP22 Acts as an Oncogene by the Activation of BMI-1-Mediated INK4a/ARF Pathway and Akt Pathway , 2011, Cell Biochemistry and Biophysics.

[180]  B. O’Malley,et al.  Molecular mechanisms of action of steroid/thyroid receptor superfamily members. , 1994, Annual review of biochemistry.

[181]  Dawn R. Chin,et al.  Cyclin-dependent Kinase 9 Is Required for Tumor Necrosis Factor-α-stimulated Matrix Metalloproteinase-9 Expression in Human Lung Adenocarcinoma Cells* , 2005, Journal of Biological Chemistry.

[182]  Simak Ali,et al.  Estrogen Receptor Alpha in Human Breast Cancer: Occurrence and Significance , 2000, Journal of Mammary Gland Biology and Neoplasia.

[183]  Michael F. Clarke,et al.  Downregulation of miRNA-200c Links Breast Cancer Stem Cells with Normal Stem Cells , 2009, Cell.

[184]  J. Brady,et al.  The bromodomain protein Brd4 is a positive regulatory component of P-TEFb and stimulates RNA polymerase II-dependent transcription. , 2005, Molecular cell.

[185]  N. Webster,et al.  The human estrogen receptor has two independent nonacidic transcriptional activation functions , 1989, Cell.

[186]  Laurence Florens,et al.  Histone Crosstalk between H2B Monoubiquitination and H3 Methylation Mediated by COMPASS , 2007, Cell.

[187]  J. Herman,et al.  Aberrant patterns of DNA methylation, chromatin formation and gene expression in cancer. , 2001, Human molecular genetics.

[188]  W. Plunkett,et al.  Transcription inhibition by flavopiridol: mechanism of chronic lymphocytic leukemia cell death. , 2005, Blood.

[189]  N. Krogan,et al.  Histone H2B Ubiquitylation Is Associated with Elongating RNA Polymerase II , 2005, Molecular and Cellular Biology.

[190]  Morag Park,et al.  Pc2-mediated Sumoylation of Smad-interacting Protein 1 Attenuates Transcriptional Repression of E-cadherin* , 2005, Journal of Biological Chemistry.

[191]  Thomas Waerner,et al.  Expression profiling of epithelial plasticity in tumor progression , 2003, Oncogene.

[192]  D. Hammond,et al.  Deubiquitinase Activities Required for Hepatocyte Growth Factor-Induced Scattering of Epithelial Cells , 2009, Current Biology.

[193]  G. Zajicek.  A new cancer hypothesis. , 1996, Medical hypotheses.

[194]  C. Palii,et al.  UTX mediates demethylation of H3K27me3 at muscle-specific genes during myogenesis , 2010, The EMBO journal.

[195]  Anna Frolov,et al.  Role of SRC-1 in the promotion of prostate cancer cell growth and tumor progression. , 2005, Cancer research.

[196]  Jianming Xu,et al.  The steroid receptor coactivator-1 regulates twist expression and promotes breast cancer metastasis. , 2009, Cancer research.

[197]  J. Gustafsson,et al.  Cloning of a novel receptor expressed in rat prostate and ovary. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[198]  K. Ozato,et al.  Brd4 marks select genes on mitotic chromatin and directs postmitotic transcription. , 2009, Molecular biology of the cell.

[199]  S. Morrison,et al.  Prospective identification of tumorigenic breast cancer cells , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[200]  K. Korach,et al.  Estrogen receptors and human disease. , 2006, The Journal of clinical investigation.

[201]  Ming-Ming Zhou,et al.  Brd4 Coactivates Transcriptional Activation of NF-κB via Specific Binding to Acetylated RelA , 2008, Molecular and Cellular Biology.

[202]  Matthew B. Wilson,et al.  Sustained induction of epithelial to mesenchymal transition activates DNA methylation of genes silenced in basal-like breast cancers , 2008, Proceedings of the National Academy of Sciences.

[203]  Dongmei Cheng,et al.  Analysis of the Large Inactive P-TEFb Complex Indicates That It Contains One 7SK Molecule, a Dimer of HEXIM1 or HEXIM2, and Two P-TEFb Molecules Containing Cdk9 Phosphorylated at Threonine 186* , 2005, Journal of Biological Chemistry.

[204]  M. T. McCabe,et al.  Cancer DNA Methylation: Molecular Mechanisms and Clinical Implications , 2009, Clinical Cancer Research.

[205]  T. Beißbarth,et al.  Estrogen-dependent gene transcription in human breast cancer cells relies upon proteasome-dependent monoubiquitination of histone H2B. , 2011, Cancer research.

[206]  G. Kreiman,et al.  Widespread transcription at neuronal activity-regulated enhancers , 2010, Nature.

[207]  Junmin Peng,et al.  Cyclin K Functions as a CDK9 Regulatory Subunit and Participates in RNA Polymerase II Transcription* , 1999, The Journal of Biological Chemistry.

[208]  E. Domany,et al.  RNF20 and USP44 regulate stem cell differentiation by modulating H2B monoubiquitylation. , 2012, Molecular cell.

[209]  Absence of p300 induces cellular phenotypic changes characteristic of epithelial to mesenchyme transition , 2006, British Journal of Cancer.

[210]  M. Churchill,et al.  The histone shuffle: histone chaperones in an energetic dance. , 2010, Trends in biochemical sciences.

[211]  M. Montano,et al.  HEXIM1 regulates 17beta-estradiol/estrogen receptor-alpha-mediated expression of cyclin D1 in mammary cells via modulation of P-TEFb. , 2008, Cancer research.

[212]  Eran Segal,et al.  Monoubiquitinated H2B is associated with the transcribed region of highly expressed genes in human cells , 2008, Nature Cell Biology.

[213]  P. V. van Diest,et al.  Poorly differentiated breast carcinoma is associated with increased expression of the human polycomb group EZH2 gene. , 2003, Neoplasia.

[214]  Charles Boone,et al.  A conserved RING finger protein required for histone H2B monoubiquitination and cell size control. , 2003, Molecular cell.

[215]  A. Shukla,et al.  Functional Analysis of H2B-Lys-123 Ubiquitination in Regulation of H3-Lys-4 Methylation and Recruitment of RNA Polymerase II at the Coding Sequences of Several Active Genes in Vivo* , 2006, Journal of Biological Chemistry.

[216]  P. Pelicci,et al.  Biological and Molecular Heterogeneity of Breast Cancers Correlates with Their Cancer Stem Cell Content , 2010, Cell.

[217]  X. de la Cruz,et al.  Do protein motifs read the histone code? , 2005, BioEssays : news and reviews in molecular, cellular and developmental biology.

[218]  Ali Shilatifard,et al.  Transcriptional activation via sequential histone H2B ubiquitylation and deubiquitylation, mediated by SAGA-associated Ubp8. , 2003, Genes & development.

[219]  J. Milton,et al.  Identification of multiple cyclin subunits of human P-TEFb. , 1998, Genes & development.

[220]  Bernhard Kuster,et al.  Requirement of ATM-dependent monoubiquitylation of histone H2B for timely repair of DNA double-strand breaks. , 2011, Molecular cell.

[221]  B. Williams,et al.  Mapping and quantifying mammalian transcriptomes by RNA-Seq , 2008, Nature Methods.

[222]  S. Frye,et al.  Targeting Chromatin Readers , 2013, Clinical pharmacology and therapeutics.

[223]  Y. Shang,et al.  Epigenetic control of epithelial-to-mesenchymal transition and cancer metastasis. , 2013, Experimental cell research.

[224]  D. Gottschling,et al.  Ubp10/Dot4p Regulates the Persistence of Ubiquitinated Histone H2B: Distinct Roles in Telomeric Silencing and General Chromatin , 2005, Molecular and Cellular Biology.

[225]  Y. Kanno,et al.  Selective recognition of acetylated histones by bromodomain proteins visualized in living cells. , 2004, Molecular cell.

[226]  J. Astola,et al.  Systematic bioinformatic analysis of expression levels of 17,330 human genes across 9,783 samples from 175 types of healthy and pathological tissues , 2008, Genome Biology.

[227]  Tae-Young Roh,et al.  Functional elements demarcated by histone modifications in breast cancer cells. , 2012, Biochemical and biophysical research communications.

[228]  Xiaochun Yu,et al.  WAC, a functional partner of RNF20/40, regulates histone H2B ubiquitination and gene transcription. , 2011, Molecular cell.

[229]  Shwu‐Yuan Wu,et al.  The Double Bromodomain-containing Chromatin Adaptor Brd4 and Transcriptional Regulation* , 2007, Journal of Biological Chemistry.

[230]  G. Berx,et al.  SIP1/ZEB2 induces EMT by repressing genes of different epithelial cell–cell junctions , 2005, Nucleic acids research.

[231]  M. Oren,et al.  CDK9 directs H2B monoubiquitination and controls replication‐dependent histone mRNA 3′‐end processing , 2009, EMBO reports.

[232]  M. Vidal,et al.  Role of histone H2A ubiquitination in Polycomb silencing , 2004, Nature.

[233]  Paul Tempst,et al.  Monoubiquitination of human histone H2B: the factors involved and their roles in HOX gene regulation. , 2005, Molecular cell.

[234]  G. Glinsky,et al.  Microarray analysis identifies a death-from-cancer signature predicting therapy failure in patients with multiple types of cancer. , 2005, The Journal of clinical investigation.

[235]  Peter A. Jones,et al.  The Epigenomics of Cancer , 2007, Cell.

[236]  T. Ley,et al.  Long-range disruption of gene expression by a selectable marker cassette. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[237]  John R Yates,et al.  Deubiquitination of Histone H2B by a Yeast Acetyltransferase Complex Regulates Transcription* , 2004, Journal of Biological Chemistry.

[238]  Chi Wang,et al.  Interaction with Suv39H1 is Critical for Snail-mediated E-cadherin Repression in Breast Cancer , 2012, Oncogene.

[239]  J. Fletcher,et al.  BRD4-NUT fusion oncogene: a novel mechanism in aggressive carcinoma. , 2003, Cancer research.

[240]  Z. Ling,et al.  Tumor development is associated with decrease of TET gene expression and 5-methylcytosine hydroxylation , 2013, Oncogene.

[241]  Johan Eide,et al.  Expression of enhancer of zeste homologue 2 is significantly associated with increased tumor cell proliferation and is a marker of aggressive breast cancer. , 2006, Clinical cancer research : an official journal of the American Association for Cancer Research.

[242]  Xin-shu Dong,et al.  Aberrant expression of USP22 is associated with liver metastasis and poor prognosis of colorectal cancer , 2011, Journal of surgical oncology.

[243]  A. Ciechanover,et al.  The ubiquitin system. , 1998, Annual review of biochemistry.

[244]  K. Korach,et al.  Estrogen receptor null mice: what have we learned and where will they lead us? , 1999, Endocrine reviews.

[245]  B. N. Devaiah,et al.  BRD4 is an atypical kinase that phosphorylates Serine2 of the RNA Polymerase II carboxy-terminal domain , 2012, Proceedings of the National Academy of Sciences.

[246]  Yi Zhang,et al.  Bre1, an E3 ubiquitin ligase required for recruitment and substrate selection of Rad6 at a promoter. , 2003, Molecular cell.

[247]  W. Huber,et al.  which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. MAnorm: a robust model for quantitative comparison of ChIP-Seq data sets , 2011 .

[248]  Hsien-Da Huang,et al.  TET1 suppresses cancer invasion by activating the tissue inhibitors of metalloproteinases. , 2012, Cell reports.

[249]  Lidong Sun,et al.  Methyl‐H3K9‐binding protein MPP8 mediates E‐cadherin gene silencing and promotes tumour cell motility and invasion , 2010, The EMBO journal.

[250]  A. Klug,et al.  Structure of the nucleosome core particle at 7 Å resolution , 1984, Nature.

[251]  George Reid,et al.  Transcription in four dimensions: nuclear receptor‐directed initiation of gene expression , 2006, EMBO reports.

[252]  H Clevers,et al.  The chromatin remodelling factor Brg‐1 interacts with β‐catenin to promote target gene activation , 2001, The EMBO journal.

[253]  E. Jensen,et al.  Estrogen-Receptor Interaction , 1973, Science.

[254]  S. Pileri,et al.  CDK9/CYCLIN T1 expression during normal lymphoid differentiation and malignant transformation , 2004, The Journal of pathology.

[255]  Craig D. Kaplan,et al.  Transcription Elongation Factors Repress Transcription Initiation from Cryptic Sites , 2003, Science.

[256]  Kevin W Eliceiri,et al.  NIH Image to ImageJ: 25 years of image analysis , 2012, Nature Methods.

[257]  E. Liu,et al.  An Oestrogen Receptor α-bound Human Chromatin Interactome , 2009, Nature.

[258]  Su Chen,et al.  Histone H2B lysine 120 monoubiquitination is required for embryonic stem cell differentiation , 2012, Cell Research.

[259]  Z. Klase,et al.  Bromodomain Protein Brd4 Regulates Human Immunodeficiency Virus Transcription through Phosphorylation of CDK9 at Threonine 29 , 2008, Journal of Virology.