Reactivation ofLatent HerpesSimplex VirusbyAdenovirus Recombinants Encoding MutantIE-OGeneProducts

Wehavepreviously shownthatadenovirus recombinants expressing functional ICPOreactivate latent herpes simplex virus type2(HSV-2) inan invitro latency system.Thisstudydemonstrated thatICPO,independent ofother HSVgeneproducts, issufficient toreactivate latent HSV-2inthis invitro system.To assess theeffects ofdefined mutations inthesequenceencoding ICPO(IE-0) on reactivation, seven in-frame insertion andthree in-frame deletion mutantswere movedinto an adenovirus expression vector. Eachrecombinant directed the synthesis ofstable ICPOofthecorrect size. Thetransactivation activity ofthemutatedsequences inthese recombinants was similar tothatwhenthey were tested inplasmids. Whenthese recombinants were examined fortheir ability toreactivate intheinvitro latency system, mutantswithdramatic defects intransactivation (Ad-0/125, Ad-0/89, Ad-0/2/7, andAd-0/88/93) were unabletoreactivate latent HSV-2independent ofthe multiplicity ofinfection. Anexception tothiscorrelation was thefinding thatAd-0/89, whichtransactivated poorly, was abletoreactivate latent virus after prolonged incubation whereas othertransactivation-deficient mutantscouldnot.Moreover, thepresence ofICP4didnotcompensatefortheinability ofany ofthe recombinants tested toreactivate HSV-2.Theseresults showthat(i) thetransactivation domainsofICPOare also usedinreactivation, (ii) thepresenceofanother essential HSVregulatory protein ICP4doesnotalter the pattern ofreactivation byICPO,and(iii) mutations insome regions ofIE-0previously showntoaffect viral growth andplaque formation didnotalter itsability toreactivate inthisinvitro system. Thelinear double-stranded DNA genome ofherpes simplexvirus type1(HSV-1) hasthecapacity toencode atleast 72unique proteins (29-31, 40). During thecourse ofproductiveinfection intissue culture, thesynthesis ofthese proteins iscoordinately regulated ina cascade fashion. This temporal program iscomposedofatleast three groups of virus proteins: immediate early (a.), early (13), andlate(-y)