title: The evolution of genes within genes and the control of DNA replication in microviruses.
abstract: Single-stranded DNA(ssDNA) viral life cycles must balance double-stranded DNA (dsDNA) and ssDNA biosynthesis. Previously published in vitro results suggest that microvirus C and host cell SSB proteins play antagonistic roles to achieve this balance. To investigate this in vivo, microvirus DNA replication was characterized in cells expressing cloned C or ssb genes, which would presumably alter the C:SSB protein ratios. Representatives of each microvirus clade (φX174, G4, and α3) were used in these studies. α3 DNA replication was significantly more complex. Results suggested that the recognized α3 C gene (C(S): small) is one of two C genes. A larger 5' extended gene could be translated from an upstream GTG start codon (C(B): big). Wild-type α3 acquired resistance to elevated SSB levels by mutations that exclusively frameshifted the C(B) reading frame, whereas mutations in the origin of replication conferred resistance to elevated C protein levels. Expression of either the cloned C(B) or C(S) gene complemented am(C) mutants, demonstrating functional redundancy. When the C(S) start codon was eliminated, strains were only viable if an additional amber mutation was placed in gene C and propagated in an informational suppressing host. Thus, C(B) protein likely reaches toxic levels in the absence of C(S) translation. This phenomenon may have driven the evolution of the C(S) gene within the larger C(B) gene and could constitute a unique mechanism of regulation. Furthermore, cross-complementation data suggested that interactions between the α3 C and other viral proteins have evolved enough specificity to biochemically isolate its DNA replication from G4 and φX174.
authors.affiliation: School of Plant Sciences and the BIO5 Institute, University of Arizona.
authors.forename: Sarah M
authors.initials: SM
authors.lastname: Doore
authors.affiliation: School of Plant Sciences and the BIO5 Institute, University of Arizona.
authors.forename: Cameron D
authors.initials: CD
authors.lastname: Baird
authors.affiliation: School of Plant Sciences and the BIO5 Institute, University of Arizona.
authors.forename: Aaron P
authors.initials: AP
authors.lastname: Roznowski
authors.affiliation: School of Plant Sciences and the BIO5 Institute, University of Arizona bfane@email.arizona.edu.
authors.forename: Bentley A
authors.initials: BA
authors.lastname: Fane
mesh_terms: D004261:DNA Replication; D004277:DNA, Single-Stranded / Q000378:metabolism; D004279:DNA, Viral / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004926:Escherichia coli / Q000378:metabolism* / Q000821:virology; D029968:Escherichia coli Proteins / Q000378:metabolism*; D019143:Evolution, Molecular; D005814:Genes, Viral; D017915:Microvirus / Q000145:classification / Q000235:genetics* / Q000254:growth & development*; D009154:Mutation; D010802:Phylogeny; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D004277:DNA, Single-Stranded; D004279:DNA, Viral; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C476430:SSB protein, E coli; D014764:Viral Proteins
keywords: DNA replication; microvirus; overlapping reading frames; phiX174