Bromodeoxyuridine-DNA interactions associated with arrest of rat odontogenesis in vitro

To better characterize the molecular mechanism responsible for the bromodeoxyuridine (BrdU)-mediated arrest of mammalian odontogenesis in vitro, the nature of nuclear DNA-analogue interactions was determined. Bioactive doses of the radiolabelled analogue were added to the tissue culture medium of 16-day old embryonic rat incisor primordia. Control rudiments were similarly exposed to equimolar, radiolabelled thymidine. After 16–18 h, DNA was isolated and purified from the labelled organ cultures. Following sedimentation to equilibrium through neutral CsCl density gradients, [³H]-BrdU-labelled DNA revealed a buoyant density indicative of a 12–15 per cent level of substitution in place of thymidine. Furthermore, similar centrifugation of DNA through alkaline density gradients suggested that the substitution was localized predominantly within a single-strand. DNA-DNA reassociation kinetics subsequently revealed that disproportionately more radiolabelled BrdU was concentrated within repetitive DNA nucleotide sequences in contrast to the more random distribution of [³H]-thymidine moieties. Thus it is likely that BrdU exerts its inhibitory effects on odontogenic differentiation through a relatively small proportion of rat embryo nuclear DNA.