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. 2013 Jul 31;41(17):8266–8279. doi: 10.1093/nar/gkt675

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© The Author(s) 2013. Published by Oxford University Press.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.

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Figure 7. — A role for the Nup116-GLEBS domain in pre-60S subunit export. (A) Top: Gle2 does not co-enrich with export competent pre-60S subunits in mutant strain expressing nup116-glebs allele. TAP purifications using Arx1 as bait were performed in strains expressing WT (NUP116) and nup116-glebs mutant allele. The calmodulin-sepharose eluates were analyzed on NuPAGE 4–12% gradient gels followed by silver staining and western analyses. The large subunit ribosomal protein L35 served as loading control. Bottom: Gle2 protein is expressed at WT level in strain expressing nup116-glebs allele. Western analysis of whole-cell extracts (WCE) derived from indicated strains using the depicted antibodies. The large ribosomal subunit protein L35 served as loading control. (B) Genetic interaction between Nup116 and pre-60S export factors. The nup116-glebs mutant is synthetically lethal when combined with nmd3ΔNES1, mex67kraa, mtr2-33 and xpo1-1 mutants and synthetically enhanced with combined with arx1Δ and ecm1Δ. Strains carrying the WT and mutant alleles were spotted in 10-fold serial dilutions on 5-FOA (SD) plates and grown at 25–30°C for 3–5 days. (C) The arx1Δnup116-glebs and ecm1Δnup116-glebs strains are impaired in pre-60S subunit export. The indicated strains containing L5-GFP reporter were grown at 30°C till mid log phase. Localization of L5-GFP reporter was analyzed by fluorescence microscopy. Scale bar = 5 µm.