Targeted proteomic study of the cyclin-Cdk module.
The cell division cycle of the yeast S. cerevisiae is driven by one Cdk (cyclin-dependent kinase), which becomes active when bound to one of nine cyclin subunits. Elucidation of Cdk substrates and other Cdk-associated proteins is essential for a full understanding of the cell cycle. Here, we report the results of a targeted proteomics study using affinity purification coupled to mass spectrometry. Our study identified numerous proteins in association with particular cyclin-Cdk complexes. These included phosphorylation substrates, ubiquitination-degradation proteins, adaptors, and inhibitors. Some associations were previously known, and for others, we confirmed their specificity and biological relevance. Using a hypothesis-driven mass spectrometric approach, we also mapped in vivo phosphorylation at Cdk consensus motif-containing peptides within several cyclin-associated candidate Cdk substrates. Our results demonstrate that this approach can be used to detect a host of transient and dynamic protein associations within a biological module.
Archambault
Vincent
V
The Rockefeller University, 1230 York Avenue, New York, NY 10021, USA.
Chang
Emmanuel J
EJ
Drapkin
Benjamin J
BJ
Cross
Frederick R
FR
Chait
Brian T
BT
Rout
Michael P
MP
D000251:Adenosine Triphosphatases; D000595:Amino Acid Sequence; D018797:Cell Cycle Proteins / Q000032:analysis* / Q000378:metabolism; D019926:Cyclin B; D018844:Cyclin-Dependent Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D016213:Cyclins / Q000032:analysis / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000032:analysis; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010766:Phosphorylation; D040901:Proteomics; D029701:Saccharomyces cerevisiae Proteins / Q000032:analysis* / Q000378:metabolism; D014157:Transcription Factors / Q000032:analysis; D014452:Ubiquitins / Q000032:analysis; D000074405:Valosin Containing Protein
C066778:ACE2 protein, S cerevisiae; C491083:CLB3 protein, S cerevisiae; C082721:CLN2 protein, S cerevisiae; D018797:Cell Cycle Proteins; D019926:Cyclin B; D016213:Cyclins; D004268:DNA-Binding Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D014452:Ubiquitins; D018844:Cyclin-Dependent Kinases; D000251:Adenosine Triphosphatases; C000614177:CDC48 protein, S cerevisiae; D000074405:Valosin Containing Protein