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. 2021 Apr 12;296:100659. doi: 10.1016/j.jbc.2021.100659

Figure 1.

Figure 1

Several nuclear transport receptors interact directly with FUS in a RanGTP-dependent manner.A, MBP or MBP–FUS was immobilized on amylose beads and incubated with HeLa cell lysate. B, MBP–FUS–EGFP or MBP–EGFP was immobilized on MBP-selector beads and incubated with lysates from HAP1 cells (control) or TNPO1-KO cells. A and B, bound proteins were analyzed by SDS-PAGE followed by Western blotting, detecting TNPO1, importin β, CRM1, and GAPDH with specific antibodies. The immobilized fusion proteins were detected using anti-MBP (A) or anti-GFP (B) antibodies. C, MBP–FUS–EGFP was immobilized on GFP–Trap magnetic agarose beads and incubated with His-tagged nuclear transport receptors (TNPO1, TNPO3, importin β/7, importin β, importin 7, importin 13, exportin 4 (Xpo4), importin α, importin 5, and CRM1) in the absence or presence of RanQ69L(aa1-180)–GTP. Bound proteins were analyzed by SDS-PAGE, followed by Coomassie staining. Input: 40%. FUS binds to TNPO1, TNPO3, importin β/7, importin β, importin 7, importin 13, and exportin 4 but not importin α (in the presence of importin β), importin 5, and CRM1 in a RanGTP-sensitive manner. Similar results were obtained when MBP–FUS–EGFP was immobilized on amylose beads (data not shown).