CK2-dependent phosphorylation of the E2 ubiquitin conjugating enzyme UBC3B induces its interaction with beta-TrCP and enhances beta-catenin degradation.
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.
Semplici
Francesca
F
Dipartimento di Biologia Molecolare, Università degli Studi di Siena, Italy.
Meggio
Flavio
F
Pinna
Lorenzo A
LA
Oliviero
Salvatore
S
D000595:Amino Acid Sequence; D064173:Anaphase-Promoting Complex-Cyclosome; D000818:Animals; D047390:Casein Kinase II; D002478:Cells, Cultured; D015500:Chloramphenicol O-Acetyltransferase / Q000378:metabolism; D003001:Cloning, Molecular; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D017931:DNA Primers / Q000737:chemistry; D019204:GTP-Binding Proteins / Q000378:metabolism*; D015723:Gene Library; D005816:Genetic Complementation Test; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D008025:Ligases / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D010766:Phosphorylation; D010957:Plasmids; D016133:Polymerase Chain Reaction; D011233:Precipitin Tests; D011401:Promoter Regions, Genetic; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D012441:Saccharomyces cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D017386:Sequence Homology, Amino Acid; D015534:Trans-Activators; D025801:Ubiquitin / Q000378:metabolism; D044763:Ubiquitin-Conjugating Enzymes; D043743:Ubiquitin-Protein Ligase Complexes; D051176:beta Catenin; D044844:beta-Transducin Repeat-Containing Proteins
C471644:BTRC protein, human; C471643:Btrc protein, mouse; C495270:CTNNB1 protein, human; C495267:CTNNB1 protein, mouse; D003598:Cytoskeletal Proteins; D017931:DNA Primers; D029701:Saccharomyces cerevisiae Proteins; D015534:Trans-Activators; D025801:Ubiquitin; D051176:beta Catenin; D044844:beta-Transducin Repeat-Containing Proteins; D015500:Chloramphenicol O-Acetyltransferase; C577449:CDC34 protein, S cerevisiae; C577447:CDC34 protein, human; C577453:Ube2r2 protein, mouse; D044763:Ubiquitin-Conjugating Enzymes; D043743:Ubiquitin-Protein Ligase Complexes; D064173:Anaphase-Promoting Complex-Cyclosome; D005982:Glutathione Transferase; D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases; D019204:GTP-Binding Proteins; D008025:Ligases