title: Evolution of Sindbis virus with a low-methionine-resistant phenotype is dependent both on a pre-existing mutation and on the methionine concentration in the medium.
abstract: SVlm21 is a mutant of Sindbis virus which was isolated by serial passage of virus in mosquito cells maintained in low-methionine medium; it therefore has a low-methionine-resistant (LMR) phenotype. This phenotype requires mutations at nts 319 and 321; these mutations result in Arg to Leu and Ser to Cys changes at positions 87 and 88 respectively in the viral methyl transferase, nsP1. To better understand the genesis of SVlm21, we carried out serial passages of viruses having only one of these amino acid changes, but in mosquito cells maintained in normal methionine-medium. Whether the passage was begun with SV319 or with SV321, the dominant virus population which emerged always acquired the second SVlm21 amino acid change. However, when the passage was begun with virus having neither the nt 319 or the nt321 mutation, even after many passages neither of these mutations was seen in the passaged virus population. Virus with the LMR phenotype emerged earlier when the virus encoded a wild-type RDRP (passage 4) rather than the mutant RDRP encoded by SVpzf (passage 7). When the methionine concentration in the medium of mosquito cells was increased to 250 µM, more than 20 passages were required until the LMR phenotype predominated. Competition experiments were carried out to compare the relative fitness of SVlm21, SVwt, SV319 and SV321 to each other. Our results indicated that SVlm21 was dominant to SVwt, as well as to both SV319 and SV321. However, SV319 and SV321 were able to co-exist with SVwt implying that in these mixed infection the presence of SVwt inhibited the emergence of SVlm21. Finally, our experiments highlight how a virus population by mutation and selection can adapt to the intracellular concentration of a simple metabolite, S-adenosylmethionine.
authors.affiliation: Department of Biochemistry and Molecular Biology, UMDNJ-Robert Wood Johnson Medical School, Piscataway, New Jersey, United States of America.
authors.forename: Victor
authors.initials: V
authors.lastname: Stollar
authors.forename: Virginia
authors.initials: V
authors.lastname: Mensah
authors.forename: Sandra
authors.initials: S
authors.lastname: Adams
authors.forename: Mei-Ling
authors.initials: ML
authors.lastname: Li
mesh_terms: D000330:Aedes / Q000166:cytology / Q000187:drug effects / Q000821:virology; D000596:Amino Acids / Q000235:genetics; D000818:Animals; D003062:Codon / Q000235:genetics; D003470:Culture Media / Q000737:chemistry*; D019020:Directed Molecular Evolution; D024882:Drug Resistance, Viral / Q000187:drug effects / Q000235:genetics*; D008156:Luciferases / Q000378:metabolism; D008715:Methionine / Q000494:pharmacology*; D050505:Mutant Proteins / Q000378:metabolism; D009154:Mutation / Q000235:genetics*; D012436:S-Adenosylmethionine / Q000378:metabolism; D012692:Serial Passage; D012845:Sindbis Virus / Q000235:genetics*; D019562:Viral Load / Q000187:drug effects
chemical_list: D000596:Amino Acids; D003062:Codon; D003470:Culture Media; D050505:Mutant Proteins; D012436:S-Adenosylmethionine; D008715:Methionine; D008156:Luciferases
references.citation: J Virol. 2003 Nov;77(21):11555-62
references.pmid: 14557641
references.citation: Proc Natl Acad Sci U S A. 2003 Jun 10;100(12):7289-94
references.pmid: 12754380
references.citation: J Virol. 1998 Aug;72(8):6546-53
references.pmid: 9658098
references.citation: J Biol Chem. 1995 Dec 15;270(50):29690-7
references.pmid: 8530357
references.citation: Virology. 1989 Dec;173(2):408-14
references.pmid: 2596021
references.citation: Anal Biochem. 2003 May 1;316(1):1-10
references.pmid: 12694720
references.citation: Proc Natl Acad Sci U S A. 1995 Jan 17;92(2):507-11
references.pmid: 7831320
references.citation: Virology. 1985 Jul 30;144(2):529-33
references.pmid: 4060595
references.citation: Virology. 1989 Jun;170(2):385-91
references.pmid: 2728344
references.citation: Microbiol Mol Biol Rev. 2012 Jun;76(2):159-216
references.pmid: 22688811
references.citation: Virology. 1977 Jul 15;80(2):390-400
references.pmid: 196393
references.citation: Mol Biosyst. 2013 Jan 27;9(1):18-28
references.pmid: 23114495
references.citation: Virology. 1990 Oct;178(2):429-34
references.pmid: 2219702
references.citation: PLoS Pathog. 2012;8(8):e1002866
references.pmid: 22916018
references.citation: Virology. 2000 Jun 20;272(1):61-71
references.pmid: 10873749
references.citation: Proc Natl Acad Sci U S A. 2008 Jun 10;105(23):7899-906
references.pmid: 18524956
references.citation: J Virol. 1987 Dec;61(12):3809-19
references.pmid: 3479621
references.citation: Curr Top Microbiol Immunol. 2006;299:51-82
references.pmid: 16568896
references.citation: Proc Natl Acad Sci U S A. 2011 Sep 20;108(38):16038-43
references.pmid: 21896755