Extended Data Fig. 10: Further information miRNA pulldown and direct transfection of AML12 cells.

a) Analyses for Molecular Function (left) and Cellular Component Gene Ontology (right) of the 67 proteins identified in the proteomic study. b) Table showing average values for relative binding enrichment of the proteins listed in the first column to the miRNA constructs shown in the top row. The columns 2-5 refer to miR-34c and its CGGGAG-containing version, while columns 6-8 refer to miR-26a and its CGGGAG-containing version. In both cases, binding to wild-type miRNAs was set as 1, and the binding of the other miRNA constructs (scramble and CGGGAG-containing version) was normalized respect to that. Only those proteins showing a log2 Fold Enrichment of CGGGAG-containing miRNA version versus wild-type miRNA >3 (>8 fold) were included. c) Brown adipocytes overexpressing wild type miR-34c (OE-miR-34c-5p-WT, grey bars) or CGGGAG-containing miR-34c (OE-miR-34c-5p-CGGGAG, red bars) were transfected with either control siRNA, Alyref siRNA or Fus siRNA (as indicated in x-axes) and analyzed by qPCR for knockdown efficiency for Alyref (left graph) or Fus (right graph), or by immunoblotting (right). *P≤0.05 (Kruskal-Wallis followed by Mann-Whitney U test), n=3. d) EXOmotif-containing miR-34c versions have the same efficiency in reducing target gene expression than wild-type miR-34c. AML12 hepatocytes were directly transfected with mimic miR-34c wild-type or its mutant versions miR-34-UGUGU, miR-34-CAUG and miR-34-CGGGAG or non-targeting miRNA (control) for 24 h and expression of predicted and experimentally-validated miR-34c target genes were analyzed. TATA-box binding protein (Tbp) was used as housekeeping gene. n=6. *P≤0.05, ***P≤0.001 (ANOVA followed by Bonferroni post-hoc test). Data are expressed as mean ± SEM.