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. 2020 Apr 6;11:1720. doi: 10.1038/s41467-020-15578-1

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© The Author(s) 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 3 — a, b Western blots of indicated proteins in the nuclear (Nuc) and cytoplasmic (Cyto) fractions as well as whole-cell lysates (WCL) of SW620 or SW480 cells transduced by lentiviruses encoding CAS9 and gFBXO22, along with gNS as a non-specific control (a), and retroviruses encoding a shRNA targeting FBXO22 (shFBXO22), along with shNC as a negative control (b). c A doxycycline (DOX)-inducible expression system encoding FBXO22 was introduced by lentiviruses into SW620 cells (SW620-FBXO22^ind). Western blots of indicated proteins in the nuclear and cytoplasmic fractions as well as whole-cell lysates of SW620-FBXO22^ind cells by DOX administration for indicated times (left) or concentrations (right). d Representative images of GFP-tagged FBXO22^WT and FBXO22^mNLS in Hela cells with re-staining of DAPI. Scale bar represents 10 μm. e A DOX-inducible expression system encoding FBXO22^WT or FBXO22^mNLS was introduced by lentiviruses into SW620 cells (SW620-FBXO22^WT-ind or SW620-FBXO22^mNLS-ind). After DOX administration, cells were fractionated and subjected to Western blots of indicated proteins. f 293T cells co-transfected with GFP, GFP-FBXO22^WT or GFP-FBXO22^mNLS and Flag-tagged PTEN^WT or PTEN^4A were subjected to immunoprecipitation with an anti-Flag antibody. Western blots of Flag- and GFP-tagged proteins in the immunoprecipitates are shown. g Western blots of indicated proteins in the nuclear and cytoplasmic fractions as well as whole-cell lysates of two clonally derived 293T cell lines depleted of FBXO22 (FBXO22^KO-1 and FBXO22^KO-2) and a negative control (NS^clone) respectively treated with 50 μg/ml CHX for hours as indicated. The experiments shown in a–g were repeated three times with similar results, and the results of one representative experiment are shown. Source data are provided as a Source Data file.