Figure 5. Glur2 mRNA is associated with PUM2, and PUM2 repressed reporter expression carrying Glur2 3′UTR.

(A) PUM2 proteins are completely removed in Pum2^E67 cortex and hippocampus. Pum2^–/– does not have the full-length wildtype PUM2 proteins but still produced a bigger fusion protein. (B) Western blot of RNA immunoprecipitation experiment revealed that PUM2 proteins could be pulled down in PUM2 immunoprecipitates from the hippocampus (6 animals were used for each genotype). (C) Glur2 mRNA is significantly enriched in PUM2 pull-down, supporting Glur2 mRNA as a potential PUM2 target. Glur2 mRNA levels in the pulldown and input were relative to β-actin, and the enrichment of Glur2 mRNA in the pulldown was based on the Glur2 mRNA enrichment of pull-down over input. (D) Three fragments corresponding to areas of the 3′ UTR containing putative PUM binding motifs were cloned into dual luciferase reporter constructs. (E) PUM2 represses translation via the 3′UTR of Glur2 as shown in dual luciferase assays. Luciferase signals were significantly reduced for all three constructed plasmids carrying different lengths of Glur2 3′ UTRs.