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. 2020 Mar 22;9(3):773. doi: 10.3390/cells9030773

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© 2020 by the authors.

Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).

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(A) Venn diagrams showing overlaps for Gis1-bound proteins under various conditions. The numbers of proteins bound to Gis1 identified from extracts prepared from cells grown under various conditions are shown here. (B) TAP pull-down confirms that Mot3, Oye2, and Pbp4 interact with Gis1. Extracts prepared from cells expressing TAP-tagged Mot3, Pbp4, or Oye2 and FLAG-tagged Gis1 were incubated with IgG-Sepharose 6 beads. Input proteins (In) and pulled-down (Pull) proteins were electrophoresed and detected by anti-TAP or anti-FLAG antibodies. Blots were probed with anti-TAP antibodies and then stripped and probed with anti-Flag antibodies. (C) The Gis1 ZnF domain, but not the JmjN/C domain, is required for interaction with Mot3. Extracts prepared from cells expressing TAP-tagged Mot3 and FLAG-tagged Gis1ΔZnF or Myc-tagged Gis1ΔNC were incubated with gG-Sepharose 6 beads. Input proteins (In) and pulled-down (Pull) proteins were electrophoresed and detected by anti-TAP, anti-Myc, or anti-FLAG antibodies. Blots were probed with anti-TAP antibodies and then stripped and probed with anti-Flag or anti-Myc antibodies.