Evidence for Preferential Digestion of Male-derived Chloroplast DNA in Young Zygotes of Chlamydomonas reinhardtii by Histochemical Immunogold Electron Microscopy.

Abstract

The fate of chloroplast nuclei (cp-nuclei) was followed in young zygotes after mating of male and female gametes in Chlamydomonas reinhardtii using two methods; i.e., observation of a single field by both epifluorescent and electron microscope, and observation by immunogold electron microscopy. Before mating, gametes contain several cp-nuclei in the cup-shaped chloroplast. In the DAPI-stained thin sections of Technovit 7100 resin, a few cp-nuclei of a gamete are clearly observed in the chloroplast and large cp-nuclei are often associated with starches around the pyrenoid in a male gamete. One hour after mating, before the fusion of cell unclei, the preferential disappearance of all male-derived cp-nuclei occurs. Two hours after mating, after fusion of cell nuclei, the preferential disappearance of cp-nuclei in all zygotes is complete. By electron microscopy, the female-derived cp-nuclei in young zygotes, which emit strong fluorescence due to 4'-6-diamidino-2-phenylindole (DAPI) under fluorescence microscope, appear to be composed of a network of fine DNA-like fibrils. By contrast, a similar network of fibrils is never observed in the male-derived chloroplast in young zygotes when the cell nuclei begin to fuse together. Gold-conjugated secondary antibodies to the primary anti-DNA antibodies have been detected on the regions around the pyrenoids in the chloroplasts in young zygotes of both male and female origin immediately after mating of the gametes. In zygotes 2 hr after mating, the gold particles appear on the regions around the female-derived pyrenoid and not on the chloroplast from the male parent. The results suggest that the preferential disappearance of cp-nuclei is caused by the digestion of the cp-DNA molecules and therefore lend support to the hypothesis that maternal inheritance of cp-genome is due to the preferential digestion of male-derived cp-DNA in young zygotes.