| Protein phosphatase-1 (PP1), through interactions with substrate targeting subunits, plays critical roles in the regulation of numerous cellular processes. At the onset of this study, we attempted to exploit sophisticated proteomics technologies, including microcystin affinity chromatography and tandem mass spectrometry, in order to identify novel targeting subunits of the PP1 catalytic subunit. Herein, we describe a newly identified regulatory subunit (PITK; P&barbelow;hosphatase I&barbelow;nteractor T&barbelow;argeting K&barbelow; protein) that specifically targets the catalytic subunit of PP1 to nuclear foci to selectively bind and dephosphorylate the transcriptional regulator heterogeneous nuclear ribonucleoprotein K (hnRNP K) at a regulatory S284 site. Additionally, PITK is phosphorylated in vivo at S1013 and S1017, residues that flank or reside within the PP1C-binding motif, and this phosphorylation negatively regulates the binding of the PP1 catalytic subunit to PITK. Incidentally, a mutant variant, S1013,1017A-PITK, when expressed in intact cells, exhibits an increase in native PP1 binding and elicits a more profound dephosphorylation of hnRNPK at S284. A global analysis of transcription by Affymetrix microarray revealed that the expression of PITK resulted in the highly altered expression of 47 genes, including a marked induction of MEK5 (>14 fold, p<0.007). Additionally, the effects of PITK and S1013,1017A-PITK on transcription could be modulated by the co-expression of hnRNP K. Taken together, our findings provide a putative mechanism by which transcriptional activity of hnRNP K can be discretely controlled through the regulation of PP1 activity. |
