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• W2034774438 endingPage "24549" @default.
• W2034774438 startingPage "24540" @default.
• W2034774438 abstract "The human importin (Imp)-β family consists of 21 nucleocytoplasmic transport carrier proteins, which transport thousands of proteins (cargoes) across the nuclear envelope through nuclear pores in specific directions. To understand the nucleocytoplasmic transport in a physiological context, the specificity of cargoes for their cognate carriers should be determined; however, only a limited number of nuclear proteins have been linked to specific carriers. To address this biological question, we recently developed a novel method to identify carrier-specific cargoes. This method includes the following three steps: (i) the cells are labeled by stable isotope labeling by amino acids in cell culture (SILAC); (ii) the labeled cells are permeabilized, and proteins in the unlabeled cell extracts are transported into the nuclei of the permeabilized cells by a particular carrier; and (iii) the proteins in the nuclei are quantitatively identified by LC-MS/MS. The effectiveness of this method was demonstrated by the identification of transportin (Trn)-specific cargoes. Here, we applied this method to identify cargo proteins specific for Imp-β, which is a predominant carrier that exclusively utilizes Imp-α as an adapter for cargo binding. We identified candidate cargoes, which included previously reported and potentially novel Imp-β cargoes. In in vitro binding assays, most of the candidate cargoes bound to Imp-β in one of three binding modes: directly, via Imp-α, or via other cargoes. Thus, our method is effective for identifying a variety of Imp-β cargoes. The identified Imp-β and Trn cargoes were compared, ensuring the carrier specificity of the method and illustrating the complexity of these transport pathways." @default.
• W2034774438 created "2016-06-24" @default.
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• W2034774438 creator A5083738682 @default.
• W2034774438 creator A5090728150 @default.
• W2034774438 date "2013-08-01" @default.
• W2034774438 modified "2023-10-11" @default.
• W2034774438 title "Identification of Cargo Proteins Specific for Importin-β with Importin-α Applying a Stable Isotope Labeling by Amino Acids in Cell Culture (SILAC)-based in Vitro Transport System*" @default.
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• W2034774438 doi "https://doi.org/10.1074/jbc.m113.489286" @default.
• W2034774438 hasPubMedCentralId "https://www.ncbi.nlm.nih.gov/pmc/articles/3750152" @default.
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• W2034774438 hasPublicationYear "2013" @default.
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