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Journal Article

Identification of CRM1-dependent nuclear export cargos using quantitative mass spectrometry.

MPS-Authors
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Karaca,  S.
Research Group of Bioanalytical Mass Spectrometry, MPI for biophysical chemistry, Max Planck Society;

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Urlaub,  H.
Research Group of Bioanalytical Mass Spectrometry, MPI for biophysical chemistry, Max Planck Society;

Fulltext (public)

1737856.pdf
(Publisher version), 4MB

Supplementary Material (public)

1737856_Suppl_1.pdf
(Supplementary material), 148KB

1737856_Suppl_2.pdf
(Supplementary material), 338KB

1737856_Suppl_3.pdf
(Supplementary material), 131KB

1737856_Suppl_4.pdf
(Supplementary material), 79KB

1737856_Suppl_5.pdf
(Supplementary material), 13KB

1737856_Suppl_6.pdf
(Supplementary material), 4MB

1737856_Suppl_7.xls
(Supplementary material), 6MB

1737856_Suppl_8.xls
(Supplementary material), 20MB

Citation

Thakar, K., Karaca, S., Port, S. A., Urlaub, H., & Kehlenbach, R. H. (2013). Identification of CRM1-dependent nuclear export cargos using quantitative mass spectrometry. Molecular and Cellular Proteomics, 12, 664-678. doi:10.1074/mcp.M112.024877.


Cite as: http://hdl.handle.net/11858/00-001M-0000-000E-F57F-D
Abstract
Chromosome region maintenance 1/exportin1/Exp1/Xpo1 (CRM1) is the major transport receptor for the export of proteins from the nucleus. It binds to nuclear export signals (NESs) that are rich in leucines and other hydrophobic amino acids. The prediction of NESs is difficult because of the extreme recognition flexibility of CRM1. Furthermore, proteins can be exported upon binding to an NES-containing adaptor protein. Here we present an approach for identifying targets of the CRM1-export pathway via quantitative mass spectrometry using stable isotope labeling with amino acids in cell culture. With this approach, we identified >100 proteins from HeLa cells that were depleted from cytosolic fractions and/or enriched in nuclear fractions in the presence of the selective CRM1-inhibitor leptomycin B. Novel and validated substrates are the polyubiquitin-binding protein sequestosome 1, the cancerous inhibitor of protein phosphatase 2A (PP2A), the guanine nucleotide-binding protein-like 3-like protein, the programmed cell death protein 2-like protein, and the cytosolic carboxypeptidase 1 (CCP1). We identified a functional NES in CCP1 that mediates direct binding to the export receptor CRM1. The method will be applicable to other nucleocytoplasmic transport pathways, as well as to the analysis of nucleocytoplasmic shuttling proteins under different growth conditions.