Liver-specific and position-effect expression of a retinol-binding protein-lacZ fusion gene (RBP-lacZ) in transgenic mice☆

[摘要] Transgenicmicecarryingaliver-specificpromoterfusedtoanuclear-targetedlacZreportergeneweregenerated.Threeseparatelinesofmiceshowedliverexpressionintheadultandnoexpressionelsewhereintheanimal.Theseresultsshowthata1.7kb5′-flankingregionintheretinol-bindingproteingenecontainsnecessaryandsufficienttranscriptionalsignalsforexpressioninadultlivers.Afourthline(R197)didnotexpressthetransgeneintheliver;instead,constitutelacZexpressionwasseenduringpostimplantationstagesofdevelopmentfromDay9.5onwardsandappearedtobeassociatedwithsegmentedstructuresincludingsomites,branchialarches,andhindbrainrhombomeresuntillatefetalstages.Theβ-galactosidasepositivecellsinR197werelaterseentogiverisetofacialandflankmusculature,andtootherregion-specificsubpopulationsofthejaws,neocortex,andbrainstem.Northernblotanalysisforthehostretinol-bindingproteinRNAtranscriptdidnotshowoverlappingtissueexpressionwiththereportergeneandsuggeststhattransgenicphenotypeseeninsegmentedembryonicstructuresofR197andotherextra-hepaticsitesisfromnovelcis-actingtranscriptionalspecificity.RNaseprotectionassaysoftheR197mRNAindicatethatthelacZsequencesareappropriatelytranscribeddownstreamoftheRBPcanonicalTATAbox,eventhoughtheRBPpromoterisitselfsilent.ThisresultwouldsuggesthostflankingsequenceswithenhanceractivitymayhaveeitheractivatedthelacZreportergeneorcooperatedwiththeRBPpromotertocreatenovelregion-specifictranscription.Breedingexperimentshavesofarfailedtoproduceoffspringsthatarehomozygousforthetransgene,andmatingoftransgenicF1siblingsroutinelyproducereducedlittersizes.Embryosthatarehomozygousforthetransgeneappeartobeunabletosurvivebeyondtheeggcylinderstagesofdevelopment.Thus,disruptionofthehostgenomebyinsertionalmutationappearstobemanifestatanearlierstagethanwhenposition-effectexpressionofthetransgeneisfirstapparent.Theseexperimentsdemonstratethatthecomponentpartsofatransgenemaybesubjecttodifferentialactivationorsuppressionbyhostgenomicflankingsequencesandthatevenastrong,tissue-specificpromotermaybeoverriddenbyhostgenes.

[学科分类] 生物科学（综合）