Role of the cytoplasmic and the intranuclear microtubules on the behavior of pronuclei during conjugation in Paramecium caudatum

Fertilization process of Paramecium caudatum includes a reciprocal exchange of migratory pronuclei across a temporary junction between conjugating cells. To know the role of microtubules on the behavior of pronuclei during conjugation in P. caudatum, the localization of microtubules was investigated by indirect immunofluorescence with a monoclonal anti-a-tubulin antibody. The immunofluorescence showed that the migratory pronucleus was surrounded by the cytoplasmic microtubules. The migratory pronucleus seems to be pushed to the cell junction by the cytoplasmic microtubules. Then the migratory pronucleus extended into the partner cell. An electron-microscopic observation showed that the cytoplasmic microtubules were assembled at the back of the migratory pronucleus, and the intranuclear microtubules of the pronucleus were aligned along the direction of the extension at the exact moment when the migratory pronucleus was crossing the cell junction. The transfer of migratory pronucleus was inhibited by injection of the monoclonal anti-a-tubulin antibody. These evidences suggest that both the cytoplasmic and the intranuclear microtubules are necessary for the dynamic behavior of gametic pronucleus across the cell junction of conjugating pair. We also suggest that microtubules are involved in the pronuclear migration after entering the partner cell.