tRNA genes protect a reporter gene from epigenetic silencing in mouse cells

It is a well-established fact that the tRNA genes in yeast can function as chromatin barrier elements. However, so far there is no experimental evidence that tRNA and other Pol III-transcribed genes exhibit barrier activity in mammals. This study utilizes a recently developed reporter gene assay to test a set of Pol III-transcribed genes and gene clusters with variable promoter and intergenic regions for their ability to prevent heterochromatin-mediated reporter gene silencing in mouse cells. The results show that functional copies of mouse tRNA genes are effective barrier elements. The number of tRNA genes as well as their orientation influence barrier function. Furthermore, the DNA sequence composition of intervening and flanking regions affects barrier activity of tRNA genes. Barrier activity was maintained for much longer time when the intervening and flanking regions of tRNA genes were replaced by AT-rich sequences, suggesting a negative role of DNA methylation in the establishment of a functional barrier. Thus, our results suggest that tRNA genes are essential elements in establishment and maintenance of chromatin domain architecture in mammalian cells.

[1]  S. Ramachandran,et al.  Expansion Mechanisms and Functional Divergence of the Glutathione S-Transferase Family in Sorghum and Other Higher Plants , 2010, DNA research : an international journal for rapid publication of reports on genes and genomes.

[2]  M. Oshimura,et al.  Human Artificial Chromosome with a Conditional Centromere for Gene Delivery and Gene Expression , 2010, DNA research : an international journal for rapid publication of reports on genes and genomes.

[3]  V. Praz,et al.  Defining the RNA polymerase III transcriptome: Genome-wide localization of the RNA polymerase III transcription machinery in human cells. , 2010, Genome research.

[4]  Z. Weng,et al.  Genomic Binding Profiles of Functionally Distinct RNA Polymerase III Transcription Complexes in Human Cells , 2010, Nature Structural &Molecular Biology.

[5]  Suresh Cuddapah,et al.  Pol II and its associated epigenetic marks are present at pol III-transcribed non-coding RNA genes , 2010, Nature Structural &Molecular Biology.

[6]  M. Gerstein,et al.  Close association of RNA polymerase II and many transcription factors with Pol III genes , 2010, Proceedings of the National Academy of Sciences.

[7]  R. Kamakaka,et al.  tRNA insulator function: Insight into inheritance of transcription states? , 2010, Epigenetics.

[8]  S. Grewal,et al.  Centromeric Localization of Dispersed Pol III Genes in Fission Yeast , 2010, Molecular biology of the cell.

[9]  Christopher D. Brown,et al.  A Comprehensive Map of Insulator Elements for the Drosophila Genome , 2010, PLoS genetics.

[10]  David Haussler,et al.  The UCSC Genome Browser database: update 2010 , 2009, Nucleic Acids Res..

[11]  Robert J. White,et al.  Regulation of TFIIIB during F9 cell differentiation , 2010, BMC Molecular Biology.

[12]  N. Dhillon,et al.  Transcription Independent Insulation at TFIIIC-Dependent Insulators , 2009, Genetics.

[13]  T. Hibi,et al.  Chromatin remodeling at Alu repeats by epigenetic treatment activates silenced microRNA-512-5p with downregulation of Mcl-1 in human gastric cancer cells , 2009, Oncogene.

[14]  V. Noskov,et al.  Human gamma-satellite DNA maintains open chromatin structure and protects a transgene from epigenetic silencing. , 2009, Genome research.

[15]  D. J. Edwards,et al.  TFIIIC Binding Sites Function as both Heterochromatin Barriers and Chromatin Insulators in Saccharomyces cerevisiae , 2008, Eukaryotic Cell.

[16]  Robert J. White,et al.  Recruitment of RNA polymerase III in vivo , 2008, Nucleic acids research.

[17]  V. Noskov,et al.  Inactivation of a Human Kinetochore by Specific Targeting of Chromatin Modifiers , 2008, Developmental cell.

[18]  N. Saitou,et al.  Possible involvement of SINEs in mammalian-specific brain formation , 2008, Proceedings of the National Academy of Sciences.

[19]  M. Teichmann,et al.  The expanding RNA polymerase III transcriptome. , 2007, Trends in genetics : TIG.

[20]  V. Corces,et al.  The role of insulator elements in large-scale chromatin structure in interphase. , 2007, Seminars in cell & developmental biology.

[21]  Gratien G. Prefontaine,et al.  Developmentally Regulated Activation of a SINE B2 Repeat as a Domain Boundary in Organogenesis , 2007, Science.

[22]  T. Mikkelsen,et al.  Systematic discovery of regulatory motifs in conserved regions of the human genome, including thousands of CTCF insulator sites , 2007, Proceedings of the National Academy of Sciences.

[23]  Michael Q. Zhang,et al.  Analysis of the Vertebrate Insulator Protein CTCF-Binding Sites in the Human Genome , 2007, Cell.

[24]  Tao Pan,et al.  Tissue-Specific Differences in Human Transfer RNA Expression , 2006, PLoS genetics.

[25]  G. Felsenfeld,et al.  Insulators: exploiting transcriptional and epigenetic mechanisms , 2006, Nature Reviews Genetics.

[26]  C. M. Hart,et al.  The Drosophila Boundary Element-Associated Factors BEAF-32A and BEAF-32B Affect Chromatin Structure , 2006, Genetics.

[27]  S. Grewal,et al.  A Role for TFIIIC Transcription Factor Complex in Genome Organization , 2006, Cell.

[28]  M. Aladjem,et al.  Preventing gene silencing with human replicators , 2006, Nature Biotechnology.

[29]  Kristin C. Scott,et al.  A Heterochromatin Barrier Partitions the Fission Yeast Centromere into Discrete Chromatin Domains , 2006, Current Biology.

[30]  M. Batzer,et al.  From the margins of the genome: mobile elements shape primate evolution , 2005, BioEssays : news and reviews in molecular, cellular and developmental biology.

[31]  Vetle I. Torvik,et al.  Mammalian microRNAs derived from genomic repeats. , 2005, Trends in genetics : TIG.

[32]  C. Lundberg,et al.  Dynamics of transgene expression in a neural stem cell line transduced with lentiviral vectors incorporating the cHS4 insulator. , 2004, Experimental cell research.

[33]  Michael Black,et al.  Role of transposable elements in heterochromatin and epigenetic control , 2004, Nature.

[34]  H. Kazazian.  Mobile Elements: Drivers of Genome Evolution , 2004, Science.

[35]  Xin Bi,et al.  Formation of Boundaries of Transcriptionally Silent Chromatin by Nucleosome-Excluding Structures , 2004, Molecular and Cellular Biology.

[36]  Jerilyn A. Walker,et al.  Genome-wide analysis of the human Alu Yb-lineage , 2004, Human Genomics.

[37]  Robert J. White,et al.  TFIIIB is phosphorylated, disrupted and selectively released from tRNA promoters during mitosis in vivo , 2003, The EMBO journal.

[38]  G. Glazko,et al.  Origin of a substantial fraction of human regulatory sequences from transposable elements. , 2003, Trends in genetics : TIG.

[39]  David I. K. Martin,et al.  Transcriptional Interference by Independently Regulated Genes Occurs in Any Relative Arrangement of the Genes and Is Influenced by Chromosomal Integration Position , 2002, Molecular and Cellular Biology.

[40]  E. Geiduschek,et al.  The RNA polymerase III transcription apparatus. , 2001, Journal of molecular biology.

[41]  J. V. Moran,et al.  Initial sequencing and analysis of the human genome. , 2001, Nature.

[42]  R. Kamakaka,et al.  RNA polymerase III and RNA polymerase II promoter complexes are heterochromatin barriers in Saccharomyces cerevisiae , 2001, The EMBO journal.

[43]  S. Fiering,et al.  Position Effects Are Influenced by the Orientation of a Transgene with Respect to Flanking Chromatin , 2001, Molecular and Cellular Biology.

[44]  S. Fiering,et al.  Site-specific chromosomal integration in mammalian cells: highly efficient CRE recombinase-mediated cassette exchange. , 1999, Journal of molecular biology.

[45]  Robert J White,et al.  p53 is a general repressor of RNA polymerase III transcription , 1998, The EMBO journal.

[46]  T. Eickbush.  Telomerase and Retrotransposons: Which Came First? , 1997, Science.

[47]  Arian F. A. Smit,et al.  MIRs are classic, tRNA-derived SINEs that amplified before the mammalian radiation , 1995, Nucleic Acids Res..

[48]  J. Nielsen,et al.  Characterization of 5S rRNA genes from mouse. , 1994, Gene.

[49]  C. Schmid,et al.  Alu transcripts: cytoplasmic localisation and regulation by DNA methylation. , 1994, Nucleic acids research.

[50]  C. Schmid.  Human Alu subfamilies and their methylation revealed by blot hybridization. , 1991, Nucleic acids research.

[51]  T. Stewart,et al.  Saturation mutagenesis of the Drosophila tRNA(Arg) gene B-Box intragenic promoter element: requirements for transcription activation and stable complex formation. , 1990, Nucleic acids research.

[52]  E. Geiduschek,et al.  Transcription by RNA polymerase III. , 1988, Annual review of biochemistry.

[53]  John D. Harding,et al.  Nucleotide sequence of a mouse tRNALeu gene , 1986, Nucleic Acids Res..

[54]  G Bernardi,et al.  The mosaic genome of warm-blooded vertebrates. , 1985, Science.

[55]  J. Harding,et al.  Structure and evolution of a mouse tRNA gene cluster encoding tRNAAsp, tRNAGly and tRNAGlu and an unlinked, solitary gene encoding tRNAAsp. , 1983, Nucleic acids research.