Fig. 4

Validation of ERβ–AGO2 interaction in the nucleus. a Co-immunoprecipitation of ERβ and AGO2 from nuclear extracts of Ct-ERβ or wild-type (wt; ERβ−, control) cells transiently transfected with a Myc-tagged AGO2 expression vector. b Co-immunoprecipitation of ERβ with AGO2, PRPF8, FXR1, and EIF6 from nuclear extracts of a Tet-inducible MCF-7 cell clone expressing Myc-Flag-ERβ; doxy doxycycline. c Top left: western blots of AGO2 and AGO1 co-immunoprecipitation with ERβ from nuclear extracts of Ct-ERβ cells before and after AGO2 silencing. Heatmap: amount (expressed as LFQ value normalized with respect to ERβ LFQ value) of proteins co-immunoprecipitated with ERβ in control (NT) and AGO2 ‘knock-down’ (shAGO2) cells measured in three biological replicates. FC average fold-change in shAGO2 vs control samples (NT). Only statistically significant changes in protein content are reported. ESR2 (bait) and AGO2 are highlighted in red and blue, respectively. Red arrowheads mark known AGO2 interactors from protein–protein interaction databases [46, 47] and the literature [12]