Three-dimensional organization of micronuclei induced by colchicine in PtK1 cells

In PtK1 cells micronucleated by colchicine, we previously demonstrated that some micronuclei contain a single chromosome. Here, we investigated interphase chromosome organization in micronucleated PtK1 cells using conventional electron microscopy and three-dimensional computer reconstruction. The distribution of micronuclei was not always polarized, but in some cells they formed a ring. When this occurred, centrioles and Golgi apparatus were located inside the ring. On freeze-fracture replicas, we observed that nuclear pore distribution among the micronuclei was heterogeneous, and on thin sections some micronuclei displayed an incomplete nuclear envelope, with gaps in the double membrane and areas without lamina or condensed chromatin. By autoradiography, we showed that the fibrillar dots were not sites of active transcription. We applied three-dimensional reconstruction to one micronucleated cell containing 22 micronuclei whose size indicated that each micronucleus probably contained one chromosome. In this cell we demonstrated that only the smallest micronuclei had an incomplete nuclear envelope. The presence in micronuclei of either nucleoli or fibrillar dots was found to be mutually exclusive. These dots might constitute stores of nucleolar proteins which migrate into micronuclei possessing no ribosomal genes. In NOR-bearing micronuclei, the structural organization was similar to that of diploid nuclei: the nucleoli were attached to the nuclear membrane and a nucleolar canal was seen, even in single-chromosome spherical micronuclei. Taken together, these findings indicate that in the diploid nuclei of PtK1 cells, the three-dimensional organization of the nucleolar domain seems to be directly controlled by the X-chromosome.