Fig. 5.

Recombinant Human antigen R (HuR) binds to PCK-5 RNA and PCK-6/7 RNA, but not PCK-4 RNA. A: PCK-4 RNA, PCK-5 RNA, and PCK-6/7 RNA are fragments of the PCK-3 RNA. Electrophoretic mobility shift assays were performed with PCK-4 RNA (B), PCK-5 RNA (C), and PCK-6/7 RNA (D). The [³²P]-labeled RNAs were incubated with increasing concentrations of recombinant HuR. PCK-4 RNA was digested with RNase T₁ before electrophoresis. Samples were run on 8% native polyacrylamide gels. The solid arrows indicate the digested or unbound RNA and the outlined arrows indicate the RNA:protein complexes. C: nucleotide sequence of PCK-5 RNA. E: competition assay was performed by incubating a constant amount of [³²P]-labeled PCK-6/7 RNA and recombinant HuR with increasing quantities of unlabeled PCK-6/7 RNA or pBS RNA. Samples were run on 5% native polyacrylamide gels. The solid arrows indicate the unbound RNA and the outlined arrows indicate the RNA:protein complexes.