A particular DNA structure is required for the function of a cis-acting component of the Epstein-Barr virus OriLyt origin of replication.
OriLyt, thecis-acting element of Epstein-Barr virus lytic origin of replication, consists of upstream and downstream components. The upstream component plays a dual role in transcription and replication. The downstream component contains a homopurine-homopyrimidine sequence which forms an H palindrome. We show that the downstream component can adopt a triple helix structure in vitro, that the 5' border of the homopyrimidine sequence is sensitive to P1 nuclease when carried by a supercoiled plasmid and that an oligonucleotide complementary to the homopyrimidine strand is taken up by a plasmid carrying the OriLyt H palindrome. We also show that all mutations which alter the H palindrome impair both oligonucleotide uptake and OriLyt-dependent replication. Interestingly, compensatory mutations which restore an H palindrome also restore oligonucleotide uptake by the mutated plasmids and their OriLyt-dependent replication. Thus, there is a strong correlation between the inability of the OriLyt H palindrome to form a non-B-DNA structure in vitro and impairment of OriLyt-dependent replication. This suggests that the presence of a non-B-DNA structure in the OriLyt downstream component is required for OriLyt-dependent replication.
Portes-Sentis
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Laboratoire de Virologie Humaine, U412 INSERM, ENS-Lyon, 46 Allée d'Italie, F-69364 Lyon cedex 07, France.
Sergeant
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Gruffat
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D001483:Base Sequence; D004261:DNA Replication; D004278:DNA, Superhelical / Q000378:metabolism; D004279:DNA, Viral / Q000737:chemistry* / Q000502:physiology; D004854:Herpesvirus 4, Human / Q000235:genetics* / Q000502:physiology; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009690:Nucleic Acid Conformation; D018741:Replication Origin; D015719:Single-Strand Specific DNA and RNA Endonucleases / Q000378:metabolism; D013329:Structure-Activity Relationship; D014407:Tumor Cells, Cultured; D014779:Virus Replication
D004278:DNA, Superhelical; D004279:DNA, Viral; D015719:Single-Strand Specific DNA and RNA Endonucleases