Recent studies with adeno-associated virus (AAV) have shown that site-
specific integration is directed by DNA sequence motifs that are prese
nt in both the viral replication origin and the chromosomal preintegra
tion DNA and that specify binding and nicking sites for the viral regu
latory Rep protein. This finding raised the question as to whether oth
er parvovirus regulatory proteins might direct site-specific recombina
tion with DNA targets that contain origin sequences functionally equiv
alent to those described for AAV. To investigate this question, active
and inactive forms of the minute virus of mice (MVM) 3' replication o
rigin, derived from a replicative-form dimer-bridge intermediate, were
propagated in an Epstein-Barr virus-based shuttle vector which replic
ates as an episome in a cell-cycle-dependent manner in mammalian cells
. Upon MVM infection of these cells, the infecting genome integrated i
nto episomes containing the active-origin sequence reported to be effi
ciently nicked by the MVM regulatory protein NS1. In contrast, MVM did
not integrate into episomes containing either the inactive form of th
e origin sequence reported to be inefficiently nicked by NS1 or the ac
tive form from which the NS1 consensus nick site had been deleted. The
structure of the cloned MVM episomal recombinants displayed several f
eatures previously described for AAV episomal and chromosomal recombin
ants. The findings indicate that the rules which govern AAV site-speci
fic recombination also apply to MVM and suggest that site-specific chr
omosomal insertions may be achievable with different autonomous parvov
irus replicator proteins which recognize binding and nicking sites on
the target DNA.