Figure 4.

MAP1B mRNA associates with QKI. (A) Coimmunoprecipitation of MAP1B mRNA with QKI from the neonatal brain. Cytoplasmic extracts prepared from P2 mouse brainstems was immunoprecipitated with (+) or without anti-QKI antibody (−) as indicated on top of the corresponding lanes. Coprecipitated RNA was subjected to RT-PCR as indicated on the left and Southern hybridization using a ³²P-labled MAP1B probe showed in the bottom panel. Immunoprecipitation was also performed using normal IgG, which does not pull down any mRNA detected by RT-PCR. (B) Top, schematic representation of the MAP1B 3′UTR (base pairs 1752–3813 in the 3′UTR) with the restriction sites (positions in MAP1B 3′UTR marked underneath) used to generate truncated transcripts. Six putative QKI-binding elements recapitulating the consensus sequence identified by SELEX were marked by asterisks (*). Bottom, QKI-binding activity by the MAP1B 5′UTR and various truncated MAP1B 3′UTR. Inset, a representative phosphorimage of captured ³⁵S-QKI by the corresponding truncated MAP1B transcripts. Scintillation counting for RNA-bound QKI is graphically displayed in the bottom panel.