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The subcellular localization of SF2/ ASF is regulated by direct interaction with SR protein kinases (SRPKs).
Serine/arginine-rich (SR) proteins play an important role in constitutive and alternative pre-mRNA splicing. The C-terminal arginine-serine domain of these proteins, such as SF2/ ASF, mediates protein-protein interactions and is phosphorylated in vivo. Using glutathione S-transferase (GST)-SF2/ ASF-affinity chromatography, the SF2/ ASF kinase activity was co-purified from HeLa cells with a 95-kDa protein, which was recognized by an anti-SR protein kinase ( SRPK) 1 monoclonal antibody. Recombinant SRPK1 and SRPK2 bound to and phosphorylated GST- SF2/ ASF in vitro. Phosphopeptide mapping showed that identical sites were phosphorylated in the pull-down kinase reaction with HeLa extracts and by recombinant SRPKs. Epitope-tagged SF2/ ASF transiently expressed in COS7 cells co-immunoprecipitated with SRPKs. Deletion analysis mapped the phosphorylation sites to a region containing an (Arg-Ser)8 repeat beginning at residue 204, and far-Western analysis showed that the region is required for binding of SRPKs to SF2/ ASF. Further binding studies showed that SRPKs bound unphosphorylated SF2/ ASF but did not bind phosphorylated SF2/ ASF. Expression of an SRPK2 kinase-inactive mutant caused accumulation of SF2/ ASF in the cytoplasm. These results suggest that the formation of complexes between SF2/ ASF and SRPKs, which is influenced by the phosphorylation state of SF2/ ASF, may have regulatory roles in the assembly and localization of this splicing factor.[1]References
- The subcellular localization of SF2/ASF is regulated by direct interaction with SR protein kinases (SRPKs). Koizumi, J., Okamoto, Y., Onogi, H., Mayeda, A., Krainer, A.R., Hagiwara, M. J. Biol. Chem. (1999)
