Dual kinase-mediated regulation of PITK by CaMKII and GSK3.
Phosphatase Interactor Targeting K protein (PITK) was previously identified as a novel PP1 targeting subunit implicated in modulating the phosphorylation of the transcriptional regulator heterogeneous nuclear ribonucleoprotein K (hnRNP K) [Kwiek NC, Thacker DF, Datto MB, Megosh HB, Haystead TA. Cell Signal 18 (10) (2006) 1769.]. Through the phosphorylation of PITK at S1013 and S1017 (residues that flank or reside within a PP1C-binding motif), the binding of the PP1 catalytic subunit to PITK, and subsequently the activity of the holoenzyme, are discretely controlled. Herein, we demonstrate that PITK phosphorylation at S1013 and S1017 also dictates the subcellular localization of the holoenzyme. Whereas both wildtype-and an S1013,1017D-PITK mutant displayed a speckled nuclear localization, a constitutively dephosphorylated form of PITK (S1013,1017A-PITK) resulted in a diffuse localization throughout the cell including the cytoplasm. Additionally, through the use of unbiased proteomics techniques, we provide evidence for a dual kinase-mediated regulation of the PITK holoenzyme whereby PITK phosphorylation at S1017 is catalyzed by calcium/calmodulin-dependent kinase II-delta (CaMKIIdelta), promoting the subsequent phosphorylation of S1013 by glycogen synthase kinase-3 (GSK3) in vitro. Taken together, our findings provide further insight into the regulation of PITK, PP1, and hnRNP K by reversible phosphorylation.
Kwiek
Nicole C
NC
Department of Pharmacology and Cancer Biology, Duke University Medical Center, Box 3813, Durham NC 27710, United States.
Thacker
Drew F
DF
Haystead
Timothy A J
TA
D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D002460:Cell Line; D002522:Chlorocebus aethiops; D038362:Glycogen Synthase Kinase 3 / Q000032:analysis / Q000378:metabolism*; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000032:analysis / Q000378:metabolism*; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
C514564:ANKRD28 protein, human; D047908:Intracellular Signaling Peptides and Proteins; D011994:Recombinant Proteins; C517660:CAMK2D protein, human; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D038362:Glycogen Synthase Kinase 3