The Nup107-160 Nucleoporin Complex Promotes Mitotic Events via Control of the Localization State of the Chromosome Passenger Complex

Abstract

The human Nup107-160 nucleoporin complex plays a major role in formation of the nuclear pore complex and is localized to kinetochores in mitosis. Here we report that Seh1, a component of the Nup107-160 complex, functions in chromosome alignment and segregation by regulating the centromeric localization of Aurora B and other chromosome passenger complex proteins. Localization of CENP-E is not affected by Seh1 depletion and analysis by electron microscopy showed that microtubule kinetochore attachments are intact. Seh1-depleted cells show impaired Aurora B localization, which results in severe defects in biorientation and organization of the spindle midzone and midbody. Our results indicate that a major function of the Nup107 complex in mitosis is to ensure the proper localization of the CPC at the centromere. INTRODUCTIONThe nuclear envelope (NE) forms the interface between the nucleus and the cytoplasm of the interphase eukaryotic cell and is essential to maintain the unique identity of each compartment. Transport between the two compartments takes place via the nuclear pore complexes (NPCs) of which there are several thousand in vertebrate somatic cells (Allen et al., 2000;Conti and Izaurralde, 2001). Each NPC contains multiple subunits of ϳ30 proteins called nucleoporins (Nups;Cronshaw et al., 2002). In higher eukaryotes NPCs are stable throughout interphase (Daigle et al., 2001), but during mitosis both the nuclear envelope and NPC undergo major structural reorganization. Starting early in prometaphase, breakdown of the nuclear envelope occurs, including disassembly of the nuclear lamina and NPCs. The nuclear envelope membrane proteins and the transmembrane nucleoporins relocalize to the endoplasmic reticulum (ER), whereas the rest of the nuclear envelope and NPC components become distributed throughout the mitotic cytoplasm (Antonin et al., 2008).During mitosis, two complete sets of chromosomes are delivered to a pair of daughter cells. Segregation of each sister chromatid pair is achieved by a highly orchestrated process that requires attachment of the sister kinetochores of each chromosome to microtubules emanating from opposite spindle poles. Kinetochores are protein structures that assemble on chromosome regions known as centromeres and mediate microtubule attachment, mitotic checkpoint signaling, and force generation (Maiato et al., 2004;Tanaka et al., 2005;Cheeseman and Desai, 2008). Electron microscopy (EM) has provided information regarding the structure of vertebrate kinetochores and has led to the division of kinetochores into three distinct regions: the inner kinetochore that associates with chromatin, the outer kinetochore that interacts with spindle microtubules, and the less dense middle kinetochore region (McEwen et al., 2007). Multiple different MT-associated proteins function at kinetochores to form a core attachment site between kinetochore and MTs, with the KMN network (KNL-1/Mis12/NDC80 complex) being the major structural component (Cheeseman et al., 2006). Functional analysis of the NDC8...

The nucleoporin Seh1 forms a complex with Mio and serves an essential tissue-specific function in Drosophila oogenesis

“…In Xenopus egg extracts and mammalian tissue culture cells, a fraction of the Nup107 complex that includes Seh1 targets to kinetochores, spindles and/or spindle poles from early prometaphase through early anaphase (Belgareh et al, 2001;Orjalo et al, 2006). Moreover, reducing the levels of Nup107-160 components disrupts spindle assembly and cytokinesis (Aitchison et al, 1995;Bai et al, 2004;Mishra et al, 2010;Orjalo et al, 2006;Platani et al, 2009;Zuccolo et al, 2007). Importantly, the specific depletion of seh1 results in the failure of the Nup107 complex to target to kinetochores at mitosis and results in multiple mitotic defects (Zuccolo et al, 2007;Platani et al, 2009).…”

“…4D-F). In mammalian cells, reducing the levels of Seh1, or other Nup107-160 complex members, results in spindle defects and mitotic delay (Mishra et al, 2010;Platani et al, 2009;Zuccolo et al, 2007). Additionally, Xenopus egg extracts depleted of Nup107-160 complex members have a reduced ability to assemble bipolar spindles (Mishra et al, 2010;Orjalo et al, 2006).…”

“…Although present on the nuclear envelope in interphase, the entire complex targets to kinetochores, spindles and spindle poles to varying extents during mitosis (Loiodice et al, 2004;Orjalo et al, 2006). Consistent with a mitotic function, depleting components of the Nup107-160 complex results in cell cycle abnormalities, including defects in mitotic spindle formation, chromosome segregation and cytokinesis (Orjalo et al, 2006;Platani et al, 2009). Moreover, recent evidence indicates that in HeLa cells and Xenopus egg extracts, the Nup107-160 complex mediates microtubule nucleation at kinetochores via its interaction with the -TuRC complex (Mishra et al, 2010).…”

IQGAP1 is associated with nuclear envelope reformation and completion of abscission

“…Its dephosphorylation and subsequent deactivation presumably allows the abscission machinery to facilitate abscission. 6,16,17 Depletion of other NPC components including the protein ELYS, a putative transcription factor, or members of the Nup107/160 complex (Nup107, Seh1 and Nup133) also results in an increase in cytokinetic cells, 18,19 however it is unknown if this delayed abscission is due to incomplete NPC reassembly and persistent aurora B activation.…”

Nucleoporin gene expression in Xenopus tropicalis embryonic development

“…This specific role for nup107 may explain its apparently stronger expression in our in situs, since it plays more than a structural role in a single subcomplex. In another study, seh1 was shown to be necessary for correct chromosome alignment in mitosis and for completion of cytokinesis (Platani et al, 2009). This suggests that in addition to the role that the 107-160 subcomplex has as a unit, individual members of the subcomplex also have unique functions.…”