title: Regulation of transcription termination by glucosylated hydroxymethyluracil, base J, in Leishmania major and Trypanosoma brucei.
abstract: Base J, β-d-glucosyl-hydroxymethyluracil, is an epigenetic modification of thymine in the nuclear DNA of flagellated protozoa of the order Kinetoplastida. J is enriched at sites involved in RNA polymerase (RNAP) II initiation and termination. Reduction of J in Leishmania tarentolae via growth in BrdU resulted in cell death and indicated a role of J in the regulation of RNAP II termination. To further explore J function in RNAP II termination among kinetoplastids and avoid indirect effects associated with BrdU toxicity and genetic deletions, we inhibited J synthesis in Leishmania major and Trypanosoma brucei using DMOG. Reduction of J in L. major resulted in genome-wide defects in transcription termination at the end of polycistronic gene clusters and the generation of antisense RNAs, without cell death. In contrast, loss of J in T. brucei did not lead to genome-wide termination defects; however, the loss of J at specific sites within polycistronic gene clusters led to altered transcription termination and increased expression of downstream genes. Thus, J regulation of RNAP II transcription termination genome-wide is restricted to Leishmania spp., while in T. brucei it regulates termination and gene expression at specific sites within polycistronic gene clusters.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Georgia, Davison Life Sciences Building, 120 Green Street, Athens, GA 30602-7229, USA.
authors.forename: David
authors.initials: D
authors.lastname: Reynolds
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Georgia, Davison Life Sciences Building, 120 Green Street, Athens, GA 30602-7229, USA.
authors.forename: Laura
authors.initials: L
authors.lastname: Cliffe
authors.affiliation: Core Unit Systems Medicine, University of Wuerzburg, Wuerzburg 97080, Germany.
authors.forename: Konrad U
authors.initials: KU
authors.lastname: Förstner
authors.affiliation: Institut Pasteur, Unité Biologie Cellulaire du Parasitisme, Département Biologie cellulaire et infection, Paris 75015, France INSERM U786, Paris 75015, France.
authors.forename: Chung-Chau
authors.initials: CC
authors.lastname: Hon
authors.affiliation: Research Center for Infectious Diseases, University of Wuerzburg, Wuerzburg 97080, Germany.
authors.forename: T Nicolai
authors.initials: TN
authors.lastname: Siegel
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Georgia, Davison Life Sciences Building, 120 Green Street, Athens, GA 30602-7229, USA rsabatini@bmb.uga.edu.
authors.forename: Robert
authors.initials: R
authors.lastname: Sabatini
mesh_terms: D002460:Cell Line; D005786:Gene Expression Regulation; D005960:Glucosides; D018320:Leishmania major / Q000201:enzymology / Q000235:genetics*; D012319:RNA Polymerase II / Q000378:metabolism; D016053:RNA, Protozoan / Q000032:analysis; D061806:Transcription Termination, Genetic; D014346:Trypanosoma brucei brucei / Q000201:enzymology / Q000235:genetics*; D014498:Uracil / Q000031:analogs & derivatives* / Q000502:physiology
chemical_list: D005960:Glucosides; D016053:RNA, Protozoan; C084880:5-((glucopyranosyloxy)methyl)uracil; D014498:Uracil; D012319:RNA Polymerase II
references.citation: Mol Biochem Parasitol. 2000 Jul;109(2):133-45
references.pmid: 10960172
references.citation: RNA. 2013 Jul;19(7):863-75
references.pmid: 23704326
references.citation: Mol Cell Biol. 2011 Apr;31(8):1690-700
references.pmid: 21321080
references.citation: Exp Cell Res. 2001 May 15;266(1):53-63
references.pmid: 11339824
references.citation: Cell. 1986 Nov 21;47(4):527-35
references.pmid: 3022935
references.citation: Nucleic Acids Res. 1996 Jul 1;24(13):2476-82
references.pmid: 8692684
references.citation: BMC Genomics. 2012 Aug 27;13:427
references.pmid: 22925482
references.citation: Proc Natl Acad Sci U S A. 1998 Mar 3;95(5):2366-71
references.pmid: 9482891
references.citation: Science. 2005 Jul 15;309(5733):404-9
references.pmid: 16020724
references.citation: Bioinformatics. 2009 Aug 15;25(16):2078-9
references.pmid: 19505943
references.citation: J Neurosci. 2011 Oct 19;31(42):15205-17
references.pmid: 22016554
references.citation: Cell. 1993 Dec 17;75(6):1129-36
references.pmid: 8261512
references.citation: Mol Microbiol. 2002 Oct;46(1):37-47
references.pmid: 12366829
references.citation: Eukaryot Cell. 2011 Nov;10(11):1465-72
references.pmid: 21926332
references.citation: Mol Cell Biol. 1990 Mar;10(3):1084-94
references.pmid: 2304458
references.citation: J Biol Chem. 1999 May 7;274(19):13041-7
references.pmid: 10224055
references.citation: Microbes Infect. 2003 Nov;5(13):1231-40
references.pmid: 14623019
references.citation: Mol Cell. 2012 Nov 9;48(3):365-74
references.pmid: 23041286
references.citation: BMC Genomics. 2009 Apr 08;10:152
references.pmid: 19356248
references.citation: Mol Biochem Parasitol. 2007 Nov;156(1):24-31
references.pmid: 17706299
references.citation: Cell Cycle. 2009 Jun 1;8(11):1698-710
references.pmid: 19411852
references.citation: Cell. 1981 Dec;27(3 Pt 2):497-505
references.pmid: 6101223
references.citation: Mol Cell Biol. 2006 May;26(10):3986-96
references.pmid: 16648491
references.citation: Cell. 1983 Oct;34(3):901-9
references.pmid: 6313213
references.citation: PLoS Pathog. 2010 Sep 09;6(9):e1001090
references.pmid: 20838601
references.citation: Genes Dev. 1993 Jun;7(6):996-1007
references.pmid: 8504937
references.citation: Genes Dev. 1997 Dec 1;11(23):3232-41
references.pmid: 9389654
references.citation: Proc Natl Acad Sci U S A. 1998 Dec 8;95(25):14687-92
references.pmid: 9843950
references.citation: Cell. 2012 Aug 31;150(5):909-21
references.pmid: 22939620
references.citation: PLoS Negl Trop Dis. 2010 Apr 13;4(4):e658
references.pmid: 20404998
references.citation: Nucleic Acids Res. 2010 Jul;38(12):3923-35
references.pmid: 20215442
references.citation: Mol Cell. 2005 Feb 4;17(3):441-51
references.pmid: 15694344
references.citation: Science. 2009 May 15;324(5929):930-5
references.pmid: 19372391
references.citation: PLoS Genet. 2013;9(8):e1003722
references.pmid: 23966880
references.citation: J Biol Chem. 2008 Jun 6;283(23):15884-92
references.pmid: 18400752
references.citation: Annu Rev Microbiol. 2008;62:235-51
references.pmid: 18729733
references.citation: Nucleic Acids Res. 2009 Apr;37(5):1452-62
references.pmid: 19136460
references.citation: Biosci Biotechnol Biochem. 2002 Apr;66(4):906-9
references.pmid: 12036074
references.citation: Nat Methods. 2012 Mar 04;9(4):357-9
references.pmid: 22388286
references.citation: Gene. 1982 Dec;20(2):281-9
references.pmid: 7166234
references.citation: Cell. 1983 Oct;34(3):891-900
references.pmid: 6313212
references.citation: Science. 2005 Jul 15;309(5733):416-22
references.pmid: 16020726
references.citation: Nat Methods. 2008 Jul;5(7):621-8
references.pmid: 18516045
references.citation: Nucleic Acids Res. 2005 Mar 21;33(5):1699-709
references.pmid: 15781496
references.citation: J Biochem. 1999 Dec;126(6):1052-9
references.pmid: 10578056
references.citation: Exp Cell Res. 2002 Jun 10;276(2):174-84
references.pmid: 12027447
references.citation: Nucleic Acids Res. 2000 Aug 15;28(16):3017-21
references.pmid: 10931915
references.citation: J Biol Chem. 2012 Jun 8;287(24):19886-95
references.pmid: 22514282
references.citation: Genes Dev. 2009 May 1;23(9):1063-76
references.pmid: 19369410
references.citation: J Cell Biol. 1989 Feb;108(2):737-46
references.pmid: 2645304
references.citation: Mol Cell. 2003 May;11(5):1291-9
references.pmid: 12769852
references.citation: Cell. 1990 Jun 29;61(7):1157-60
references.pmid: 2142018
references.citation: FEBS J. 2010 Nov;277(21):4539-48
references.pmid: 21040474
references.citation: Open Biol. 2012 Apr;2(4):120033
references.pmid: 22724062
references.citation: EMBO J. 2002 Apr 15;21(8):1881-8
references.pmid: 11953307
references.citation: Nucleic Acids Res. 1982 Jun 25;10(12):3591-604
references.pmid: 6287414