Nup2 requires a highly divergent partner, NupA, to fulfill functions at nuclear pore complexes and the mitotic chromatin region.
Bottom Line: These mitotic problems are not caused by overall defects in mitotic NPC disassembly-reassembly or general nuclear import.However, without Nup2 or NupA, although the SAC protein Mad1 locates to its mitotic locations, it fails to locate to NPCs normally in G1 after mitosis.Collectively the study provides new insight into the roles of Nup2 and NupA during mitosis and in a surveillance mechanism that regulates nucleokinesis when mitotic defects occur after SAC fulfillment.
Affiliation: Laboratory of Gene Regulation and Development, National Institute for Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892.Show MeSH
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Mentions: To investigate mitotic progression without Nup2 or NupA, we used GFP-tagged tubulin to monitor the dynamics of spindle formation (Ovechkina et al., 2003). We found that the time to complete mitosis was longer and overall more variable in nup2- and nupA-deleted cells, taking 10–11 min on average to complete versus 5.5 min for wild-type cells (Figure 6, A and B, and Supplemental Figure S3). To see whether the mitotic defects caused by the absence of Nup2 and NupA are monitored by the SAC, we asked whether the mitotic delays in these mutants are dependent on a functional SAC response (Li and Murray, 1991; Musacchio and Salmon, 2007). We found that in the absence of Mad2, the mitotic delays caused by nup2 or nupA deletion were abolished, and the average time of mitosis was similar to that of wild-type controls, with the variability in the duration of mitosis no longer apparent (Figure 6, A and B, and Supplemental Figure S3). We also followed Mad1-GFP in nup2- and nupA- mutants to monitor SAC activation, which causes the transition of Mad1-GFP from NPCs to mitotic kinetochores. As occurs in the wild type, in the mutant Mad1-GFP translocates to kinetochores during the first mitosis during spore germination. Mad1-GFP then stays associated with the kinetochore region, as defined by Ndc80-ChRFP, during the extended mitotic period (Figure 6C). Absence of Nup2 or NupA therefore causes defects in mitosis, which are monitored by the SAC.
Affiliation: Laboratory of Gene Regulation and Development, National Institute for Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892.