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. 2013 Apr 18;8(4):e61697. doi: 10.1371/journal.pone.0061697

Figure 5. A phosphorylation-independent interaction between Dcp1a and decapping components and RNA.

Figure 5

(A) Flag-tagged Dcp1a and HA-tagged CA or DN MAPKK1 mutants were co-expressed in HEK 293T cells. Cell extracts were immunoprecipitated using M2 beads. Input (5% of cell extracts; left panel) and immunoprecipitated protein complexes (right panel) were analyzed by western blotting using anti-Flag, anti-Ddx6, anti-Edc3, or anti-Edc4. The arrowhead indicates the specific Edc3 band. (B) Co-IP of Dcp1a and other decapping factors in 3T3-L1 cell extracts. Cell extracts from control cells or cells induced to differentiate for 1 h in the presence or absence of 20 µM U0126 were incubated with anti-Dcp1a and protein A–Sepharose. After extensive washing, the precipitated protein complexes were analyzed with western blotting with anti-Dcp1a, anti-Ddx6, anti-Edc3, anti-Edc4, and anti-HuR. The arrowhead indicates the specific Ddx6 band. Similar results were independently reproduced three times.