E3 ligases confer specificity to ubiquitination by recognizing target substrates and mediating transfer of ubiquitin from an E2 ubiquitinconjugating enzyme to substrate. The activity of most E3s is specified by a RING domain, which binds to an E2∼ubiquitin thioester and activates discharge of its ubiquitin cargo. E2-E3 complexes can either monoubiquitinate a substrate lysine or synthesize polyubiquitin chains assembled via different lysine residues of ubiquitin. These modifications can have diverse effects on the substrate, ranging from proteasome-dependent proteolysis to modulation of protein function, structure, assembly, and/or localization. Not surprisingly, RING E3mediated ubiquitination can be regulated in a number of ways. RING-based E3s are specified by over 600 human genes, surpassing the 518 protein kinase genes. Accordingly, RING E3s have been linked to the control of many cellular processes and to multiple human diseases. Despite their critical importance, our knowledge of the physiological partners, biological functions, substrates, and mechanism of action for most RING E3s remains at a rudimentary stage.

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RING Domain E3 Ubiquitin Ligases

CK2 (formerly termed "casein kinase 2") is a ubiquitous, highly pleiotropic and constitutively active Ser/Thr protein kinase whose implication in neoplasia, cell survival, and virus infection is supported by an increasing number of arguments. Here an updated inventory of 307 CK2 protein substrates is presented. More than one-third of these are implicated in gene expression and protein synthesis as being either transcriptional factors (60) or effectors of DNA/RNA structure (50) or translational elements. Also numerous are signaling proteins and proteins of viral origin or essential to virus life cycle. In comparison, only a minority of CK2 targets (a dozen or so) are classical metabolic enzymes. An analysis of 308 sites phosphorylated by CK2 highlights the paramount relevance of negatively charged side chains that are (by far) predominant over any other residues at positions n+3 (the most crucial one), n+1, and n+2. Based on this signature, it is predictable that proteins phosphorylated by CK2 are much more numerous than those identified to date, and it is possible that CK2 alone contributes to the generation of the eukaryotic phosphoproteome more so than any other individual protein kinase. The possibility that CK2 phosphosites play some global role, e.g., by destabilizing alpha helices, counteracting caspase cleavage, and generating adhesive motifs, will be discussed.

One-thousand-and-one substrates of protein kinase CK2?

Protein kinase CK2 (9casein kinase II9) has traditionally been classified as a messenger-independent protein serine/threonine kinase that is typically found in tetrameric complexes consisting of two catalytic (α and/or α′) subunits and two regulatory β subunits. Accumulated biochemical and genetic evidence indicates that CK2 has a vast array of candidate physiological targets and participates in a complex series of cellular functions, including the maintenance of cell viability. This review summarizes current knowledge of the structural and enzymic features of CK2, and discusses advances that challenge traditional views of this enzyme. For example, the recent demonstrations that individual CK2 subunits exist outside tetrameric complexes and that CK2 displays dual-specificity kinase activity raises new prospects for the precise elucidation of its regulation and cellular functions. This review also discusses a number of the mechanisms that contribute to the regulation of CK2 in cells, and will highlight emerging insights into the role of CK2 in cellular decisions of life and death. In this latter respect, recent evidence suggests that CK2 can exert an anti-apoptotic role by protecting regulatory proteins from caspase-mediated degradation. The mechanistic basis of the observation that CK2 is essential for viability may reside in part in this ability to protect cellular proteins from caspase action. Furthermore, this anti-apoptotic function of CK2 may contribute to its ability to participate in transformation and tumorigenesis.

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Protein kinase CK2: structure, regulation and role in cellular decisions of life and death.

Phosphorylation of β-Catenin by AKT Promotes β-Catenin Transcriptional Activity *

The Tumor Suppressor PTEN Is Phosphorylated by the Protein Kinase CK2 at Its C Terminus IMPLICATIONS FOR PTEN STABILITY TO PROTEASOME-MEDIATED DEGRADATION

http://www3.unisi.it/oliviero_lab/download/Orlandini_et_al._JBC_1998.pdf

Protein Kinase CK2α′ Is Induced by Serum as a Delayed Early Gene and Cooperates with Ha-ras in Fibroblast Transformation

Protein kinase CK2 is a ubiquitous and pleiotropic Ser/Thr protein kinase involved in cell growth and transformation. Here we report the identification by yeast interaction trap of a CK2 interacting protein, UBC3B, which is highly homologous to the E2 ubiquitin conjugating enzyme UBC3/CDC34. UBC3B complements the yeast cdc34-2 cell cycle arrest mutant in S. cerevisiae and transfers ubiquitin to a target substrate in vitro. UBC3B is specifically phosphorylated by CK2 in vitro and in vivo. We mapped by deletions and site directed mutagenesis the phosphorylation site to a serine residue within the C-terminal domain in position 233 of UBC3B and in the corresponding serine residue of UBC3. Following CK2-dependent phosphorylation both UBC3B and UBC3 bind to the F-box protein beta-TrCP, the substrate recognition subunit of an SCF (Skp1, Cul1, F-box) ubiquitin ligase. Furthermore, we observed that co-transfection of CK2alpha' together with UBC3B, but not with UBC3DeltaC, enhances the degradation of beta-catenin. Taken together these data suggest that CK2-dependent phosphorylation of UBC3 and UBC3B functions by regulating beta-TrCP substrate recognition.

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CK2-dependent phosphorylation of the E2 ubiquitin conjugating enzyme UBC3B induces its interaction with beta-TrCP and enhances beta-catenin degradation.

Francesca Semplici

Papers

Protein Kinase CK2α′ Is Induced by Serum as a Delayed Early Gene and Cooperates with Ha-ras in Fibroblast Transformation

CK2-dependent phosphorylation of the E2 ubiquitin conjugating enzyme UBC3B induces its interaction with beta-TrCP and enhances beta-catenin degradation.