title: The ULb' region of the human cytomegalovirus genome confers an increased requirement for the viral protein kinase UL97.
abstract: We report a requirement for the viral protein kinase UL97 in human cytomegalovirus (HCMV) replication that maps to the ULb' region of the viral genome. A UL97-null (Δ97) mutant of strain TB40/E, which encodes a full-length ULb' region, exhibited replication defects, particularly in production of cell-free virus, that were more severe than those seen with a Δ97 mutant of laboratory strain AD169, which harbors extensive deletions in its ULb' region. These differences were recapitulated with additional HCMV strains by treatment with a UL97 kinase inhibitor, 1-(β-L-ribofuranosyl)-2-isopropylamino-5,6-dichlorobenzimidazole (maribavir). We observed lower levels of viral DNA synthesis and an increased requirement for UL97 in viral late gene expression in strains with full-length ULb' regions. Analysis of UL97-deficient TB40/E infections by electron microscopy revealed fewer C-capsids in nuclei, unusual viral particles in the cytoplasmic assembly compartment, and defective viral nuclear egress. Partial inhibition of viral DNA synthesis caused defects in production of cell-free virus that were up to ≈ 100-fold greater than those seen with cell-associated virus in strains TB40/E and TR, suggesting that UL97-dependent defects in cell-free virus production in strains with full-length ULb' regions were secondary to DNA synthesis defects. Accordingly, a chimeric virus in which the ULb' region of TB40/E was replaced with that of AD169 showed reduced effects of UL97 inhibition on viral DNA synthesis, late gene expression, and production of cell-free virus compared to parental TB40/E. Together, these results argue that the ULb' region encodes a factor(s) which invokes an increased requirement for UL97 during viral DNA synthesis.
authors.affiliation: Department of Microbiology & Immunology and Center for Molecular and Tumor Virology, LSU Health Sciences Center, Shreveport, Louisiana, USA.
authors.forename: Depeng
authors.initials: D
authors.lastname: Wang
authors.forename: Gang
authors.initials: G
authors.lastname: Li
authors.forename: Martin
authors.initials: M
authors.lastname: Schauflinger
authors.forename: Christopher C
authors.initials: CC
authors.lastname: Nguyen
authors.forename: Ellie D
authors.initials: ED
authors.lastname: Hall
authors.forename: Andrew D
authors.initials: AD
authors.lastname: Yurochko
authors.forename: Jens
authors.initials: J
authors.lastname: von Einem
authors.forename: Jeremy P
authors.initials: JP
authors.lastname: Kamil
mesh_terms: D003587:Cytomegalovirus / Q000201:enzymology* / Q000235:genetics / Q000502:physiology; D003586:Cytomegalovirus Infections / Q000821:virology*; D015967:Gene Expression Regulation, Viral; D016679:Genome, Viral; D006801:Humans; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism; D014779:Virus Replication
chemical_list: D014764:Viral Proteins; D017853:Phosphotransferases (Alcohol Group Acceptor); C075365:ganciclovir kinase
references.citation: Virology. 1980 Feb;101(1):10-24
references.pmid: 6243816
references.citation: J Clin Invest. 2010 Sep;120(9):3191-208
references.pmid: 20679731
references.citation: Nature. 1995 Mar 9;374(6518):177-80
references.pmid: 7877691
references.citation: J Virol. 2013 May;87(9):5019-27
references.pmid: 23427156
references.citation: J Virol. 2011 Apr;85(8):3821-32
references.pmid: 21289123
references.citation: J Virol. 2003 Jan;77(2):905-14
references.pmid: 12502806
references.citation: Virology. 1988 Nov;167(1):87-96
references.pmid: 2847426
references.citation: Virology. 2006 Oct 10;354(1):69-79
references.pmid: 16872656
references.citation: Proc Natl Acad Sci U S A. 2003 Nov 25;100(24):14223-8
references.pmid: 14623981
references.citation: Methods Mol Biol. 2010;634:421-30
references.pmid: 20677001
references.citation: Clin Diagn Lab Immunol. 2001 Nov;8(6):1279-81
references.pmid: 11687477
references.citation: Biotechniques. 2006 Feb;40(2):191-7
references.pmid: 16526409
references.citation: J Virol. 2007 Oct;81(19):10659-68
references.pmid: 17634236
references.citation: Proc Natl Acad Sci U S A. 2003 Oct 14;100(21):12396-401
references.pmid: 14519856
references.citation: J Virol. 2010 Oct;84(20):10832-43
references.pmid: 20686030
references.citation: J Virol. 2006 Jan;80(2):710-22
references.pmid: 16378974
references.citation: J Virol. 1987 May;61(5):1358-67
references.pmid: 3033266
references.citation: Science. 2008 May 9;320(5877):797-9
references.pmid: 18467589
references.citation: J Virol. 2007 Mar;81(6):3042-8
references.pmid: 17192309
references.citation: Blood. 2008 Jun 1;111(11):5403-10
references.pmid: 18285548
references.citation: Arch Intern Med. 1993 Sep 13;153(17):2010-6
references.pmid: 8357286
references.citation: J Virol. 1975 Jan;15(1):71-80
references.pmid: 46282
references.citation: Biochem Biophys Res Commun. 1975 Sep 2;66(1):262-71
references.pmid: 169847
references.citation: J Virol. 1999 Jul;73(7):5663-70
references.pmid: 10364316
references.citation: Science. 1978 Sep 1;201(4358):819-21
references.pmid: 210500
references.citation: Antiviral Res. 2006 Sep;71(2-3):154-63
references.pmid: 16765457
references.citation: J Virol. 1992 May;66(5):3241-6
references.pmid: 1313929
references.citation: Proc Natl Acad Sci U S A. 2001 Feb 13;98(4):1895-900
references.pmid: 11172047
references.citation: Rev Med Virol. 2010 May;20(3):136-55
references.pmid: 20084641
references.citation: Am J Transplant. 2012 Nov;12(11):3021-30
references.pmid: 22947426
references.citation: Birth Defects Orig Artic Ser. 1984;20(1):305-24
references.pmid: 6329369
references.citation: J Virol. 1997 Jan;71(1):405-11
references.pmid: 8985364
references.citation: J Gen Virol. 2008 Feb;89(Pt 2):359-368
references.pmid: 18198366
references.citation: Curr Opin Virol. 2011 Dec;1(6):555-62
references.pmid: 22440913
references.citation: Virology. 2010 Sep 30;405(2):457-63
references.pmid: 20637486
references.citation: Lancet Infect Dis. 2011 Apr;11(4):284-92
references.pmid: 21414843
references.citation: J Virol. 1977 Feb;21(2):584-600
references.pmid: 189089
references.citation: Transpl Infect Dis. 2010 Dec;12(6):489-96
references.pmid: 20682012
references.citation: J Virol. 2008 May;82(10):5054-67
references.pmid: 18321963
references.citation: J Biol Chem. 2005 Sep 30;280(39):33357-67
references.pmid: 15975922
references.citation: Cell. 2006 Mar 24;124(6):1283-98
references.pmid: 16564017
references.citation: PLoS One. 2009 Aug 06;4(8):e6529
references.pmid: 19657394
references.citation: Proc Natl Acad Sci U S A. 2009 Sep 29;106(39):16823-8
references.pmid: 19805380
references.citation: J Virol. 2005 Aug;79(16):10330-8
references.pmid: 16051825
references.citation: J Virol. 2002 Sep;76(18):9551-5
references.pmid: 12186938
references.citation: PLoS Pathog. 2007 Jan;3(1):e6
references.pmid: 17206862
references.citation: J Gen Virol. 2010 Jun;91(Pt 6):1535-46
references.pmid: 20479471
references.citation: J Virol. 2005 Dec;79(24):15494-502
references.pmid: 16306620
references.citation: Cell Microbiol. 2012 May;14(5):644-55
references.pmid: 22329758
references.citation: Nature. 1996 Jul 4;382(6586):88-90
references.pmid: 8657311
references.citation: Virology. 1983 Oct 15;130(1):118-33
references.pmid: 6314643
references.citation: J Virol. 2011 Apr;85(7):3375-84
references.pmid: 21248036
references.citation: J Gen Virol. 2010 Mar;91(Pt 3):605-15
references.pmid: 19906940
references.citation: J Microsc. 2007 Oct;228(Pt 1):78-87
references.pmid: 17910700
references.citation: Proc Natl Acad Sci U S A. 2005 Dec 13;102(50):18153-8
references.pmid: 16319222
references.citation: Proc Natl Acad Sci U S A. 2003 Dec 9;100(25):14976-81
references.pmid: 14657367
references.citation: Arch Ophthalmol. 1998 Feb;116(2):178-85
references.pmid: 9488269
references.citation: PLoS Pathog. 2012;8(7):e1002789
references.pmid: 22792066
references.citation: J Virol. 2009 Mar;83(6):2480-90
references.pmid: 19116255
references.citation: Curr Opin Immunol. 2007 Oct;19(5):577-82
references.pmid: 17716883
references.citation: J Virol. 2005 Sep;79(18):11837-47
references.pmid: 16140760
references.citation: J Virol. 1996 Jan;70(1):78-83
references.pmid: 8523595
references.citation: J Gen Virol. 2006 Sep;87(Pt 9):2451-2460
references.pmid: 16894182
references.citation: Antimicrob Agents Chemother. 2002 Aug;46(8):2365-72
references.pmid: 12121906
references.citation: PLoS Pathog. 2009 Jan;5(1):e1000275
references.pmid: 19165338
references.citation: J Virol. 2000 Sep;74(17):7720-9
references.pmid: 10933677
references.citation: Mol Cell Biol. 2003 Mar;23(6):1885-95
references.pmid: 12612064
references.citation: Lancet Infect Dis. 2011 Apr;11(4):255-7
references.pmid: 21414844
references.citation: J Clin Virol. 2008 Mar;41(3):174-9
references.pmid: 18203656
references.citation: Blood. 2007 Aug 1;110(3):937-45
references.pmid: 17440050
references.citation: Curr Protoc Microbiol. 2010 Aug;Chapter 14:Unit 14E.3
references.pmid: 20812216
references.citation: Oncogene. 1991 Jul;6(7):1139-46
references.pmid: 1861863