The ATP,Mg-dependent protein phosphatase: Regulation by casein kinase-1

AbstractThe free modulator subunit of the ATP,Mg-dependent phosphatase is phosphorylated up to 1 mol per mol by casein kinase-1, up to 1.85 mol per mol after dephosphorylation by the PCSH1 phosphatase, but 10-fold less when purified in the presence of NaF, suggesting an in vivo phosphorylation of the casein kinase-1 sites. Peptide mapping of 32P-modulator labeled by casein kinase-1 or -2 shows a different phosphorylation pattern. Phosphorylation of the inactive phosphatase by casein kinase-1 prevents the subsequent kinase FA-mediated activation, while it does not impair the activated phosphatase

Phosphorylation of the phosphatase modulator subunit (inhibitor-2) by casein kinase-1 Identification of the phosphorylation sites

AbstractThe isolated modulator subunit of the inactive protein phosphatase-1 is phosphorylated in vitro by casein kinase-1 at two different site Ser-86 and Ser-174. The Ser-86 site is a common target for casein kinase-1 and casein kinase-2, but is preferentially phosphorylated by the former enzyme. The Ser-174 site seems to be specific for casein kinase-1, and is phosphorylated at a slower rate. These results give a new insight into the in vitro phosphorylation pattern of the modulator subunit of the phosphatase and provides additional data on the specificity of casein kinase-1

Phosphorylation of histone deacetylase 7 by protein kinase D mediates T cell receptor–induced Nur77 expression and apoptosis

The molecular basis of thymocyte negative selection, a crucial mechanism in establishing central tolerance, is not yet resolved. Histone deacetylases (HDACs) have emerged as key transcriptional regulators in several major developmental programs. Recently, we showed that the class IIa member, HDAC7, regulates negative selection by repressing expression of Nur77, an orphan nuclear receptor involved in antigen-induced apoptosis of thymocytes. Engagement of the T cell receptor (TCR) alleviates this repression through phosphorylation-dependent nuclear exclusion of HDAC7. However, the identity of the TCR-activated kinase that phosphorylates and inactivates HDAC7 was still unknown. Here, we demonstrate that TCR-induced nuclear export of HDAC7 and Nur77 expression is mediated by activation of protein kinase D (PKD). Indeed, active PKD stimulates HDAC7 nuclear export and Nur77 expression. In contrast, inhibition of PKD prevents TCR-mediated nuclear exclusion of HDAC7 and associated Nur77 activation. Furthermore, we show that HDAC7 is an interaction partner and a substrate for PKD. We identify four serine residues in the NH2 terminus of HDAC7 as targets for PKD. More importantly, a mutant of HDAC7 specifically deficient in phosphorylation by PKD, inhibits TCR-mediated apoptosis of T cell hybridomas. These findings indicate that PKD is likely to play a key role in the signaling pathways controlling negative selection