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. Author manuscript; available in PMC: 2014 Apr 30.
Published in final edited form as: Nat Commun. 2013;4:2388. doi: 10.1038/ncomms3388

Figure 7. HuR binding to the HuRBS prevents the recruitment of Ago2 to the HMGB1 mRNA.

Figure 7

(a) Western blot analysis was performed using antibodies against HuR and α-tubulin as a loading control on total cell extracts obtained from exponentially growing C2C12 cells treated with a control (Ctr) or HuR specific (siHuR) siRNA. (b) Immunoprecipitation experiments were performed using a monoclonal Ago2 antibody, or anti-IgG antibody as a control, on the total cell lysates described in (a). The immunoprecipitation of Ago2 was then assessed by western blotting using an anti-Ago2 antibody. (c–e) RNA was isolated from the immunoprecipitate described above, and quantitative RT-PCR was performed using primers specific to (c) HMGB1 (d) miR-1192 (e) miR-16. The levels of HMGB1 mRNA, miR-1192 and miR-16 in each IP, relative to those in the IgG IP, were respectively normalized against the GAPDH mRNA and U6 levels. Error bars represent S.E.M of three independent experiments.