Figure 3.

The spacer of SRPK1 lacks detectable NES. Both Myc-tagged full-length SRPK1 (A) and inactive SRPK1 containing a mutation in the ATP binding site (K109M) (D) were mainly localized in the cytoplasm of transfected HeLa cells. Removal of the spacer (K1-ΔS) resulted in exclusive localization of the kinase in the nucleus (G), colocalizing with phosphorylated SC35 in nuclear speckles 8 h posttransfection (H and I). Prolonged expression of the spacer-deleted kinase (24 h posttransfection, J) induced aggregation of splicing factors in the nucleus (K and L). Nuclear entrance seems to be saturable as indicated by a level of cytoplasmic kinase (J). Inactivation of the kinase activity in spacer-deleted SRPK1 (K-ΔS-K109M) prevented nuclear translocation (M). SC35 was detected with a monoclonal anti-SC35 antibody (B, E, H, K, and N). Potential colocalization was determined by merged images (C, F, I, L, and O).