Differential mitotic phosphorylation of proteins of the nuclear pore complex.

During each cell cycle, the nucleus of higher eukaryotes undergoes a dramatic assembly and disassembly. These events can be faithfully reproduced in vitro using cell-free extracts derived from Xenopus eggs. Such extracts contain three major N-acetylglucosaminylated proteins, p200, p97, and p60. All three become assembled into reconstituted nuclear pores. Here we show that p200, p97, and p60 exist in eggs in soluble high molecular mass complexes of 1000, 450, and 600 kDA, respectively. The bulk of p60 is stably associated with proteins of 58 and 54 kDa, while p200 is associated with a fraction of p60 in a separate complex lacking p58 and p54. Upon examining the behavior of these proteins in the cell cycle, we find that p200 and p97 are highly phosphorylated at mitosis, both in vivo and in vitro. Moreover, in extracts that cycle between interphase and mitosis, p200 and p97 are specifically phosphorylated at mitosis. Corresponding with their mitotic phosphorylation, both p200 and p97 are specific substrates for purified mitotic Cdc2 kinase, whereas nucleoporin p60 is not. Analysis indicates that the size of the complexes containing the pore N-acetylglucosamine glycoproteins does not change during mitosis, suggesting that such complexes represent stable multicomponent modules into which the nucleus disassembles at mitosis.