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 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.