Semi-in vitro detection of Mg(2+)-dependent DNase that specifically digest mitochondrial nucleoids in the zygote of Physarum polycephalum

The maternal/uniparental inheritance of mitochondria is controlled by the selective elimination of paternal/uniparental mitochondria and digestion of their mitochondrial DNA (mtDNA). In isogamy, the selective digestion of mtDNA in uniparental mitochondria is initiated after mating and is completed prior to the elimination of mitochondria, but the molecular mechanism of the digestion of uniparental mtDNA remains unknown. In this study, we developed a semi-in vitro assay for DNase, wherein the digestion of mitochondrial nucleoids (mt-nucleoids) was microscopically observed using isolated mitochondria from Physarum polycephalum and the DNase involved in uniparental inheritance was characterized. When myxamoebae of AI35 and DP246 are crossed, mtDNA and mt-nucleoid from only the DP246 parent are digested. The digestion of mt-nucleoids was observed in zygotes 3 h after plating for mating. During the digestion of mt-nucleoids, mitochondrial membrane integrity was maintained. In the semi-in vitro assay, the digestion of mt-nucleoids was only observed in the presence of Mg(2+) at pH 7.5–9.0. Moreover, such Mg(2+)-dependent DNase activity was specifically detected in mitochondria isolated from zygotes 3 h after plating for mating. Therefore, Mg(2+)-dependent DNase is potentially involved in uniparental inheritance. Our findings provide insights into the DNase involved in uniparental inheritance and its regulatory mechanism.