title: Quantitative framework for ordered degradation of APC/C substrates.
abstract: BACKGROUND
  During cell-cycle progression, substrates of a single master regulatory enzyme can be modified in a specific order. Here, we used experimental and computational approaches to dissect the quantitative mechanisms underlying the ordered degradation of the substrates of the ubiquitin ligase APC/C(Cdc20), a key regulator of chromosome segregation in mitosis.
  
  
  RESULTS
  We show experimentally that the rate of catalysis varies with different substrates of APC/C(Cdc20). Using a computational model based on multi-step ubiquitination, we then show how changes in the interaction between a single substrate and APC/C(Cdc20) can alter the timing of degradation onset relative to APC/C(Cdc20) activation, while ensuring a fast degradation rate. Degradation timing and dynamics depend on substrate affinity for the enzyme as well as the catalytic rate at which the substrate is modified. When two substrates share the same pool of APC/C(Cdc20), their relative enzyme affinities and rates of catalysis influence the partitioning of APC/C(Cdc20) among substrates, resulting in substrate competition. Depending on how APC/C(Cdc20) is partitioned among its substrates, competition can have minor or major effects on the degradation of certain substrates. We show experimentally that increased expression of the early APC/C(Cdc20) substrate Clb5 does not delay the degradation of the later substrate securin, arguing against a role for competition with Clb5 in establishing securin degradation timing.
  
  
  CONCLUSIONS
  The degradation timing of APC/C(Cdc20) substrates depends on the multi-step nature of ubiquitination, differences in substrate-APC/C(Cdc20) interactions, and competition among substrates. Our studies provide a conceptual framework for understanding how ordered modification can be established among substrates of the same regulatory enzyme, and facilitate our understanding of how precise temporal control is achieved by a small number of master regulators to ensure a successful cell division cycle.
authors.affiliation: Departments of Physiology and Biochemistry & Biophysics, University of California, San Francisco, CA, 94158, USA.
authors.forename: Dan
authors.initials: D
authors.lastname: Lu
authors.affiliation: Departments of Physiology and Biochemistry & Biophysics, University of California, San Francisco, CA, 94158, USA.
authors.forename: Juliet R
authors.initials: JR
authors.lastname: Girard
authors.affiliation: Departments of Physiology and Biochemistry & Biophysics, University of California, San Francisco, CA, 94158, USA.
authors.forename: Weihan
authors.initials: W
authors.lastname: Li
authors.affiliation: Departments of Physiology and Biochemistry & Biophysics, University of California, San Francisco, CA, 94158, USA.
authors.forename: Arda
authors.initials: A
authors.lastname: Mizrak
authors.affiliation: Departments of Physiology and Biochemistry & Biophysics, University of California, San Francisco, CA, 94158, USA. David.Morgan@ucsf.edu.
authors.forename: David O
authors.initials: DO
authors.lastname: Morgan
mesh_terms: D064173:Anaphase-Promoting Complex-Cyclosome / Q000235:genetics / Q000378:metabolism*; D064199:Cdc20 Proteins / Q000235:genetics / Q000378:metabolism*; D002453:Cell Cycle; D008938:Mitosis; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D064249:Securin / Q000235:genetics / Q000378:metabolism*; D054875:Ubiquitination
chemical_list: C577495:CDC20 protein, S cerevisiae; D064199:Cdc20 Proteins; D029701:Saccharomyces cerevisiae Proteins; D064249:Securin; D064173:Anaphase-Promoting Complex-Cyclosome