[Show abstract][Hide abstract] ABSTRACT: Ubiquitin ligases, together with their cognate ubiquitin-conjugating enzymes, are responsible for the ubiquitylation of proteins,
a process that regulates a myriad of eukaryotic cellular functions. The first cullin-RING ligase discovered, yeast SCFCdc4, functions with the conjugating enzyme Cdc34 to regulate the cell cycle. Cdc34 orthologs are notable for their highly acidic
C-terminal extension. Here we confirm that the Cdc34 acidic C-terminal tail has a role in Cdc34 binding to SCFCdc4 and makes a major contribution to the submicromolar Km of Cdc34 for SCFCdc4. Moreover, we demonstrate that a key functional property of the tail is its acidity. Our analysis also uncovers an unexpected
new function for the acidic tail in promoting catalysis. We demonstrate that SCF is functional when Cdc34 is fused to the
C terminus of Cul1 and that this fusion retains partial function even when the acidic tail has been deleted. The Cdc34-SCF
fusion proteins that lack the acidic tail must interact in a fundamentally different manner than unfused SCF and wild type
Cdc34, demonstrating that distinct mechanisms of E2 recruitment to E3, as is seen in nature, can sustain substrate ubiquitylation.
Finally, a search of the yeast proteome uncovered scores of proteins containing highly acidic stretches of amino acids, hinting
that electrostatic interactions may be a common mechanism for facilitating protein assembly.
Preview · Article · Oct 2009 · Journal of Biological Chemistry
[Show abstract][Hide abstract] ABSTRACT: Exit from mitosis is characterized by a precipitous decline in cyclin-dependent kinase (Cdk) activity, dissolution of mitotic structures, and cytokinesis. In Saccharomyces cerevisiae, mitotic exit is driven by a protein phosphatase, Cdc14, which is in part responsible for counteracting Cdk activity. Throughout interphase, Cdc14 is sequestered in the nucleolus, but successful anaphase activates the mitotic exit network (MEN), which triggers dispersal of Cdc14 throughout the cell by a mechanism that has remained unknown. In this study, we show that a MEN component, protein kinase Dbf2-Mob1, promotes transfer of Cdc14 to the cytoplasm and consequent exit from mitosis by direct phosphorylation of Cdc14 on serine and threonine residues adjacent to a nuclear localization signal (NLS), thereby abrogating its NLS activity. Our results define a mechanism by which the MEN promotes exit from mitosis.
Preview · Article · Mar 2009 · The Journal of Cell Biology