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. 2024 Jan 24;10(4):eadk9394. doi: 10.1126/sciadv.adk9394

Fig. 9. MERVL-gag blocks URI binding to OCT4 and SOX2.

Fig. 9.

(A) Plasmid construct for constitutive and stable EGFP or MERVL-gag overexpression in mESCs. (B) WB of pool of clones from EGFP or MERVL-gag overexpression in mESCs. (C) IF of MERVL-gag, SOX2, and URI in the pool of clones from MERVL-gag overexpressing mESCs. Scale bar, 100 μm. (D) WB of single clones from EGFP or MERVL-gag overexpression in mESCs. (E) IF of MERVL-gag, NANOG, SOX2, and URI in single clones from EGFP or MERVL-gag overexpression in mESCs. Scale bars, 10 μm. (F) IF of MERVL-gag and URI in totipotent early 2C, late 2C, and 4C embryo. Projection, single Z-slice and magnifications for each channel are depicted. Scale bar, 20 μm. (G) Colocalization levels of URI with MERVL-gag (left) and vice versa (right) from (F). t test; ****P < 0.0001. (H to J) Coimmunoprecipitation assays of OCT4 (H), SOX2 (I), and URI (J) in mESCs cultured in presence of 2i/LIF. Short (s.e.) and long (l.e.) exposure are shown. (K to M) Coimmunoprecipitation assays of URI (K) and MERVL-gag in DUX-induced 2C-like cells (L) or MERVL-gag overexpressing cells (M). s.e. and l.e. are shown. (N) Competition pulldowns of IVT SOX2 or OCT4 in combination with increasing IVT MERVL-gag using GST-URI. IVT MERVL-gag quantity is annotated as percentage of loaded IVT inputs. s.e. and l.e. exposures are shown. (O) Pulldown levels of SOX2 and OCT4 from (N). (P) Working model. A 4% of the input was loaded as WCE in coimmunoprecipitation experiments.