Nuclear pore protein NUP88 activates anaphase-promoting complex to promote aneuploidy
Ryan M. Naylor, … , Xiuqi Cao, Jan M. van Deursen
Ryan M. Naylor, … , Xiuqi Cao, Jan M. van Deursen
Published January 5, 2016
Citation Information: J Clin Invest. 2016;126(2):543-559. https://doi.org/10.1172/JCI82277.
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Nuclear pore protein NUP88 activates anaphase-promoting complex to promote aneuploidy

Abstract

The nuclear pore complex protein NUP88 is frequently elevated in aggressive human cancers and correlates with reduced patient survival; however, it is unclear whether and how NUP88 overexpression drives tumorigenesis. Here, we show that mice overexpressing NUP88 are cancer prone and form intestinal tumors. To determine whether overexpression of NUP88 drives tumorigenesis, we engineered transgenic mice with doxycycline-inducible expression of Nup88. Surprisingly, NUP88 overexpression did not alter global nuclear transport, but was a potent inducer of aneuploidy and chromosomal instability. We determined that NUP88 and the nuclear transport factors NUP98 and RAE1 comprise a regulatory network that inhibits premitotic activity of the anaphase-promoting complex/cyclosome (APC/C). When overexpressed, NUP88 sequesters NUP98-RAE1 away from APC/CCDH1, triggering proteolysis of polo-like kinase 1 (PLK1), a tumor suppressor and multitasking mitotic kinase. Premitotic destruction of PLK1 disrupts centrosome separation, causing mitotic spindle asymmetry, merotelic microtubule-kinetochore attachments, lagging chromosomes, and aneuploidy. These effects were replicated by PLK1 insufficiency, indicating that PLK1 is responsible for the mitotic defects associated with NUP88 overexpression. These findings demonstrate that the NUP88-NUP98-RAE1-APC/CCDH1 axis contributes to aneuploidy and suggest that it may be deregulated in the initiating stages of a broad spectrum of human cancers.

Authors

Ryan M. Naylor, Karthik B. Jeganathan, Xiuqi Cao, Jan M. van Deursen

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Figure 4

NUP88 sequesters NUP98-RAE1 away from APC/CCDH1 prior to mitotic entry when overexpressed.

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NUP88 sequesters NUP98-RAE1 away from APC/CCDH1 prior to mitotic entry w...
(A) Coprecipitation analysis of RO-3306–treated HeLa cells immunoprecipitated with an anti-NUP98 Ab. (B) Coprecipitation analysis of RO-3306–treated HeLa cells transduced with pTRIPZ–HA-Nup88 (NUP88) or pTRIPZ empty vector (EV) lentiviruses and immunoprecipitated with anti–HA affinity matrix. (C) Coprecipitation analysis of cells in B immunoprecipitated with an anti-NUP98 Ab. Values represent NUP88/EV protein quantification ratios. (D) Images of WT MEFs immunostained for APC6, CDC27, or CDH1. p-HH3 (Ser10) was used to identify cells in the G2 phase. (E) Western blot analysis of cytoplasmic (C) and nuclear (N) fractions from MEFs of the indicated genotypes. (F) Same as in E. (G) Coprecipitation analysis of cytoplasmic fractions from MEFs of the indicated genotypes immunoprecipitated with anti–HA affinity matrix. (H) Same as in G, except immunoprecipitated with an anti-NUP88 Ab. (I) Live-cell–imaging analysis of chromosome segregation errors in H2B-mRFP–positive MEFs of the indicated genotypes. (J) Mitotic checkpoint analysis of MEFs in I challenged with nocodazole. (K) Western blot of RO-3306–treated or (L) nocodazole-arrested MEFs of the indicated genotypes. Analysis in I was performed on 3 independent lines per genotype (25 cells/line). Analysis in J was performed on 3 independent lines per genotype (>10 cells/line). Data represent the mean ± SEM. Western blots are representative of 3 independent experiments. Cyclin B1, cyclin A2, and actin blots in K and L are identical to those in Figure 5, G and H. WT and NUP98 RAE1 values in I and J are identical to those in Figure 2, F and H. Statistical significance in I and J was determined using a 2-tailed, unpaired t test. *P < 0.05, **P < 0.01, and ***P < 0.001. Scale bar: 10 μm. WCE, whole-cell extract.