Minute virus of mice initiator protein NS1 and a host KDWK family transcription factor must form a precise ternary complex with origin DNA for nicking to occur

Citation
J. Christensen et al., Minute virus of mice initiator protein NS1 and a host KDWK family transcription factor must form a precise ternary complex with origin DNA for nicking to occur, J VIROLOGY, 75(15), 2001, pp. 7009-7017
Citations number
34
Categorie Soggetti
Microbiology
Journal title
JOURNAL OF VIROLOGY
ISSN journal
0022538X → ACNP
Volume
75
Issue
15
Year of publication
2001
Pages
7009 - 7017
Database
ISI
SICI code
0022-538X(200108)75:15<7009:MVOMIP>2.0.ZU;2-P
Abstract
Parvoviral rolling hairpin replication generates palindromic genomic concat emers whose junctions are resolved to give unit-length genomes by a process involving DNA replication initiated at origins derived from each viral tel omere, The left-end origin of minute virus of mice (MVM), oriL, contains bi nding sites for the viral initiator nickase, NS1, and parvovirus initiation factor (PIF), a member of the emerging KDWK family of transcription factor s, oriL is generated as an active form, oriL(TC), and as an inactive form, oriL(GAA), which contains a single additional nucleotide inserted between t he NS1 and PIF sites. Here we examined the interactions on oriL(TC), which lead to activation of NS1 by PIF, The two subunits of PIF, p79 and p96, coo peratively bind two ACGT half-sites, which can be flexibly spaced. When coe xpressed from recombinant baculoviruses, the PIF subunits preferentially fo rm heterodimers which, in the presence of ATP, show cooperative binding wit h NS1 on oriL, but this interaction is preferentially enhanced on oriL(TC), compared to oriL(GAA). Without ATP, NS1 is unable to bind stably to its co gnate site, but PIF facilitates this interaction, rendering the NSI binding site, but not the nick site, resistant to DNase I. Varying the spacing of the PIF half-sites shows that the distance between the NS1 binding site and the NS1-proximal half-site is critical for nickase activation, whereas the position of the distal half-site is unimportant. When expressed separately , both PIF subunits form homodimers that bind site specifically to oriL, bu t only complexes containing p79 activate the NS1 nickase function.