G-patch domain and KOW motifs-containing protein, GPKOW; a nuclear RNA-binding protein regulated by protein kinase A

Using yeast two-hybrid screening with the PKA C subunit Cbeta2 as bait, we identified the G-patch domain and KOW motifs-containing protein (GPKOW), also known as the T54 protein or MOS2 homolog, as an interaction partner for Cbeta2. We demonstrate that GPKOW, which contains one G-patch domain and two KOW motifs, is a nuclear RNA-binding protein conserved between species. GPKOW contains two sites that are phosphorylated by PKA in vitro. By RNA immunoprecipitation and site directed mutagenesis of the PKA phosphorylation sites we revealed that GPKOW binds RNA in vivo in a PKA sensitive fashion.GPKOW is a RNA-binding protein that binds RNA in a PKA regulated fashion. Together with our previous results demonstrating that PKA regulates pre-mRNA splicing, our results suggest that PKA phosphorylation is involved in regulating RNA processing at several steps.Post-transcriptional processing of pre-mRNA involves splicing, editing and polyadenylation and starts as soon as pre-mRNAs are produced at their sites of transcription. In addition to small non-coding RNAs (snRNAs) these processes require the activity of RNA-binding proteins (RBPs) [1,2]. The discovery of the heterogeneous nuclear ribonucleoproteins (hnRNP) and other pre-mRNA/mRNA-binding proteins led to the identification of the first amino acid motifs and functional domains that confer binding to RNA [3]. RBPs, which may contain one or multiple RNA-binding domains, are important for RNA metabolism and are involved in RNA stability, splicing, transport, cellular localization, translation and decay [4]. The posttranscriptional RNA regulon model suggests that functionally related mRNAs are co-regulated by RBPs and snRNAs. This allows for a smooth and coordinated regulation of mRNAs and protein synthesis according to specific tasks [5].Little is known about how signal transduction pathways may be involved in regulating post-transcriptional processing of pre-mRNA. For alternative splicing, it is established that reversible