A subset of FG-nucleoporins is necessary for efficient Msn5-mediated nuclear protein export

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• 作者：Erin M. Finn¹ ; ^{efinn87@gmail.com} ; Elise P. DeRoo ; George W. Clement² ; ^{gwclement@gmail.com} ; Sheila Rao³ ; ^{raoshe@gmail.com} ; Sarah E. Kruse⁴ ; ^{sarah.kruse.87@gmail.com} ; Kate M. Kokanovich⁵ ; ^{kkokanovich@gmail.com} ; Kenneth D. Belanger ; ^{kbelanger@colgate.edu}
• 关键词：Nuclear pore complex ; Nuclear export ; Nucleoporin ; Karyopherin ; Exportin
• 刊名：Biochimica et Biophysica Acta (BBA)/Molecular Cell Research
• 出版年：2013
• 出版时间：May, 2013
• 年：2013
• 卷：1833
• 期：5
• 页码：1096-1103
• 全文大小：732 K

文摘

The transport of proteins between the cytoplasm and nucleus requires interactions between soluble transport receptors (karyopherins) and phenylalanine-glycine (FG) repeat domains on nuclear pore complex proteins (nucleoporins). However, the role of specific FG repeat-containing nucleoporins in nuclear protein export has not been carefully investigated. We have developed a novel kinetic assay to investigate the relative export kinetics mediated by the karyopherin Msn5/Kap142 in yeast containing specific FG-Nup mutations. Using the Msn5 substrate Crz1 as a marker for Msn5-mediated protein export, we observe that deletions of NUP100 or NUP2 result in decreased rates of Crz1 export, while nup60¦¤ and nup42¦¤ mutants do not vary significantly from wild type. The decreased Msn5 export rate in nup100¦¤ was confirmed using Mig1-GFP as a transport substrate. A nup100¦¤GLFG mutant shows defects in nuclear export kinetics similar to a nup100¦¤ deletion. Removal of FG-repeats from Nsp1 also decreases export kinetics, while a loss of Nup1 FXFGs does not. To confirm that our export data reflected functional differences in protein localization, we performed Crz1 transcription activation assays using a CDRE::LacZ reporter gene that is upregulated upon increased transcription activation by Crz1 in vivo. We observe that expression from this reporter increases in nup100¦¤GLFG and nsp1¦¤FG¦¤FXFG strains that exhibit decreased Crz1 export kinetics but resembles wild-type levels in nup1¦¤FXFG strains that do not exhibit export defects. These data provide evidence that the export of Msn5 is likely mediated by a specific subset of FG-Nups and that the GLFG repeat domain of Nup100 is important for Msn5-mediated nuclear protein export.