Nucleocytoplasmic sorting of macromolecules following mitosis: fate of nuclear constituents after inhibition of pore complex function.
PtK2 cells in which pore complex-mediated transport is blocked by microinjection early in mitosis of a monoclonal antibody (specific for an Mr 68,000 pore complex glycoprotein) or of wheat germ agglutinin (WGA) complete cytokinesis. However, their nuclei remain stably arrested in a telophase-like organization characterized by highly condensed chromatin and the absence of nucleoli, indicating a requirement for pore-mediated transport for the reassembly of interphase nuclei. We have now examined this requirement more closely by monitoring the behavior of individual nuclear macromolecules in microinjected cells using immunofluorescence microscopy and have investigated the effect of microinjecting the antibody or WGA on cellular ultrastructure. The absence of nuclear transport did not affect the sequestration into daughter nuclei of components such as DNA, DNA topoisomerase I and the nucleolar protein fibrillarin that are carried through mitosis on chromosomes. On the other hand, lamins, snRNAs and the p68 pore complex glycoprotein, all cytoplasmic during mitosis, remained largely cytoplasmic in the telophase-arrested cells. Electron microscopy showed the nuclei to be surrounded by a double-layered membrane with some inserted pore complexes. In addition, however, a variety of membranous structures with associated pore complexes was regularly noted in the cytoplasm, suggesting that chromatin may not be essential for the postmitotic formation of pore complexes. We propose that cellular compartmentalization at telophase is a two-step process. First, a nuclear envelope tightly encloses the condensed chromosomes, excluding non-selectively all macromolecules not associated with the chromosomes. Interphase nuclear organization is then progressively restored by selective pore complex-mediated uptake of nuclear proteins from the cytoplasm.
Benavente
R
R
Institute of Zoology I, University of Würzburg, Federal Republic of Germany.
Scheer
U
U
Chaly
N
N
D000818:Animals; D001692:Biological Transport; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism* / Q000648:ultrastructure; D002868:Chromosomal Proteins, Non-Histone / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism* / Q000648:ultrastructure; D004247:DNA / Q000378:metabolism*; D004264:DNA Topoisomerases, Type I / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D034882:Lamins; D008845:Microinjections; D008854:Microscopy, Electron; D008938:Mitosis; D009687:Nuclear Proteins / Q000378:metabolism*; D012313:RNA / Q000378:metabolism*
D002868:Chromosomal Proteins, Non-Histone; D034882:Lamins; D009687:Nuclear Proteins; C047458:fibrillarin; D012313:RNA; D004247:DNA; D004264:DNA Topoisomerases, Type I