THE MINUTE VIRUS OF MICE (MVM) NONSTRUCTURAL PROTEIN NS1 INDUCES NICKING OF MVM DNA AT A UNIQUE SITE OF THE RIGHT-END TELOMERE IN BOTH HAIRPIN AND DUPLEX CONFORMATIONS IN-VITRO

The right-end telomere of replicative form (RF) DNA of the autonomous parvovirus minute virus of mice (MVM) consists of a sequence that is s elf-complementary except for a th ree nucleotide loop around the axis of symmetry and an interior bulge of three unpaired nucleotides on one strand (designated the right-end 'bubble'). This right-end inverted r epeat can exist in the form of a folded-back strand (hairpin conformat ion) or in an extended form, base-paired to a copy strand (duplex conf ormation), We recently reported that the right-end telomere is process ed in an A9 cell extract supplemented with the MVM nonstructural prote in NS1. This processing is shown here to result from the NS1-dependent nicking of the complementary strand at a unique position 21 nt inboar d of the folded-back genomic 5' end, DNA species terminating in duplex or hairpin configurations, or in a mutated structure that has lost th e right-end bulge, are all cleaved in the presence of NS1, indicating that features distinguishing these structures are not prerequisites fo r nicking under the in vitro conditions tested, Cleavage of the hairpi n structure is followed by strand-displacement synthesis, generating t he right-end duplex conformation, while processing of the duplex struc ture leads to the release of free right-end telomeres, In the majority of molecules, displacement synthesis at the right terminus stops a fe w nucleotides before reaching the end of the template strand, possibly due to NS1 which is covalently bound to this end. A fraction of the r ight-end duplex product undergoes melting and re-folding into hairpin structures (formation of a 'rabbit-ear' structure).