Protein kinase CK2 catalyzes tyrosine phosphorylation in mammalian cells.

Protein kinase CK2 exhibits oncogenic activity in mice and is over-expressed in a number of tumors or leukemic cells. On the basis of its amino acid sequence and a wealth of experimental information, CK2 has traditionally been classified as a protein serine/threonine kinase. In contrast to this traditional view of CK2, recent evidence has shown that CK2 can also phosphorylate tyrosine residues under some circumstances in vitro and in yeast. In this study, we provide definitive evidence demonstrating that CK2 also exhibits tyrosine kinase activity in mammalian cells. Tyrosine phosphorylation of CK2 in cells and in CK2 immunoprecipitates is dependent on CK2 activity and is inhibited by the CK2 selective inhibitor 4,5,6,7-tetrabromobenzotriazole. Examination of phosphotyrosine profiles in cells reveals a number of proteins, including CK2 itself, which exhibit increased tyrosine phosphorylation when CK2 levels are increased. Peptide arrays to evaluate the specificity determinants for tyrosine phosphorylation by CK2 reveal that its specificity for tyrosine phosphorylation is distinct from its specificity for serine/threonine phosphorylation. Of particular note is the requirement for an aspartic acid immediately C-terminal to the phosphorylatable tyrosine residue. Collectively, these data provide conclusive evidence that CK2 catalyzes the phosphorylation of tyrosine residues in mammalian cells, a finding that adds a new level of complexity to the challenge of elucidating its cellular functions. Furthermore, these results raise the possibility that increased CK2 levels that frequently accompany transformation may contribute to the increased tyrosine phosphorylation that occurs in transformed cells

Protein kinase CK2: Systematic relationships with other posttranslational modifications

© Springer International Publishing Switzerland 2015. A wealth of biochemical and genetic evidence has demonstrated that protein kinase CK2 has critical roles in the regulation and execution of numerous biological processes. Large-scale proteomic and phosphoproteomics studies have further reinforced the widespread impact of CK2 on cellular events through interactions with many cellular proteins or protein complexes and through phosphorylation of a vast number of cellular proteins. Given its global participation in many fundamental processes, it is not surprising that CK2 has been implicated in numerous human diseases, a factor that has spurred interest in CK2 as a candidate for molecular- targeted therapy. Despite this growing profile, many questions regarding its precise mechanisms of regulation remain. In fact, several lines of evidence suggest that CK2 is constitutively active, leading to a speculation that CK2 is an unregulated enzyme. Accordingly, there is an apparent paradox that leads to the question of how an unregulated enzyme such as CK2 can be a participant in regulatory processes. In an effort to resolve this paradox, studies in our lab and others have focused on an investigation of the relationships between CK2 and other cellular pathways. Using a combination of computational predictions and database mining together with proteomic strategies and biochemical assays, we have been elucidating systematic relationships between CK2 and regulatory pathways where CK2 phosphorylation sites overlap other posttranslational modifications. Overall, these studies suggest intriguing mechanisms by which CK2 can participate in regulatory events and also how alterations in CK2 levels that accompany disease may promote pathological rewiring of regulatory pathways

A biosensor to monitor dynamic regulation and function of tumour suppressor PTEN in living cells

Tumour suppressor ​PTEN is a phosphatase that negatively regulates the PI3K/AKT pathway. The ability to directly monitor ​PTEN conformation and function in a rapid, sensitive manner is a key step towards developing anti-cancer drugs aimed at enhancing or restoring ​PTEN-dependent pathways. Here we developed an intramolecular bioluminescence resonance energy transfer (BRET)-based biosensor, capable of detecting signal-dependent ​PTEN conformational changes in live cells. The biosensor retains intrinsic properties of ​PTEN, enabling structure–function and kinetic analyses. BRET shifts, indicating conformational change, were detected following mutations that disrupt intramolecular ​PTEN interactions, promoting plasma membrane targeting and also following physiological ​PTEN activation. Using the biosensor as a reporter, we uncovered ​PTEN activation by several G protein-coupled receptors, previously unknown as ​PTEN regulators. Trastuzumab, used to treat ​ERBB2-overexpressing breast cancers also elicited activation-associated ​PTEN conformational rearrangement. We propose the biosensor can be used to identify pathways regulating ​PTEN or molecules that enhance its anti-tumour activity