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. 2021 Oct 1;12:5779. doi: 10.1038/s41467-021-26049-6

Fig. 4. SPOP-mediated poly-ubiquitination of Geminin inhibits MCM proteins binding to Cdt1.

Fig. 4

a PC-3 cells were infected with lentivirus expressing indicated shRNAs or WT or mutated Geminin and harvested for co-IP and WB analysis. b PC-3 cells were infected with lentivirus expressing empty vector or different SPOP mutants and harvested for co-IP and WB. c The structure of human Cdt1-Geminin is overlaid to that of yeast Cdt1-MCM after conformational optimization. A modeled K27-linked poly-ubiquitin chain attached to Geminin Lys127 (light blue) is incompatible with Cdt1-Geminin binding to the MCM complex. d–f PC-3 cells were infected with lentivirus expressing control shRNA or Geminin-specific shRNA in combination with empty vector (EV), Geminin WT, or K100/127R mutant. Cells were pulse-labeled with 30 μM BrdU prior to WB analysis (d), FACS analysis (e), or quantification (f). Data are presented as the mean ± SD of three independent experiments. Two-tailed unpaired Student’s t-test; ***P < 0.001. Source data are provided in this paper or Mendeley database (10.17632/8n7xt5rkhc.1). Similar results for (a), (b), and (d) panels were obtained in three independent experiments.