title: Human CUL1 forms an evolutionarily conserved ubiquitin ligase complex (SCF) with SKP1 and an F-box protein.
abstract: The SCF ubiquitin ligase complex of budding yeast triggers DNA replication by catalyzing ubiquitination of the S phase cyclin-dependent kinase inhibitor SIC1. SCF is composed of three proteins-ySKP1, CDC53 (Cullin), and the F-box protein CDC4-that are conserved from yeast to humans. As part of an effort to identify components and substrates of a putative human SCF complex, we isolated hSKP1 in a two-hybrid screen with hCUL1, the closest human homologue of CDC53. Here, we show that hCUL1 associates with hSKP1 in vivo and directly interacts with both hSKP1 and the human F-box protein SKP2 in vitro, forming an SCF-like particle. Moreover, hCUL1 complements the growth defect of yeast cdc53(ts) mutants, associates with ubiquitination-promoting activity in human cell extracts, and can assemble into functional, chimeric ubiquitin ligase complexes with yeast SCF components. Taken together, these data suggest that hCUL1 functions as part of an SCF ubiquitin ligase complex in human cells. Further application of biochemical assays similar to those described here can now be used to identify regulators/components of hCUL1-based SCF complexes, to determine whether the hCUL2-hCUL5 proteins also are components of ubiquitin ligase complexes in human cells, and to screen for chemical compounds that modulate the activities of the hSKP1 and hCUL1 proteins.
authors.affiliation: Division of Biology, California Institute of Technology, Pasadena, CA 91125, USA.
authors.forename: S A
authors.initials: SA
authors.lastname: Lyapina
authors.forename: C C
authors.initials: CC
authors.lastname: Correll
authors.forename: E T
authors.initials: ET
authors.lastname: Kipreos
authors.forename: R J
authors.initials: RJ
authors.lastname: Deshaies
mesh_terms: D042846:CDC2-CDC28 Kinases; D018797:Cell Cycle Proteins / Q000378:metabolism*; D017124:Conserved Sequence; D044842:Cullin Proteins; D051357:Cyclin-Dependent Kinase 2; D050756:Cyclin-Dependent Kinase Inhibitor Proteins; D018844:Cyclin-Dependent Kinases / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000378:metabolism; D019143:Evolution, Molecular; D044783:F-Box Proteins; D000073979:F-Box-WD Repeat-Containing Protein 7; D005656:Fungal Proteins / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008025:Ligases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D044786:S-Phase Kinase-Associated Proteins; D029701:Saccharomyces cerevisiae Proteins; D044767:Ubiquitin-Protein Ligases; D014452:Ubiquitins / Q000378:metabolism
chemical_list: C100988:Cdc53 protein, S cerevisiae; D018797:Cell Cycle Proteins; C100318:Cullin 1; D044842:Cullin Proteins; D050756:Cyclin-Dependent Kinase Inhibitor Proteins; D004791:Enzyme Inhibitors; D044783:F-Box Proteins; D000073979:F-Box-WD Repeat-Containing Protein 7; C513273:FBXW7 protein, human; D005656:Fungal Proteins; D044786:S-Phase Kinase-Associated Proteins; C086926:SIC1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D014452:Ubiquitins; D044767:Ubiquitin-Protein Ligases; D017346:Protein Serine-Threonine Kinases; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases; D008025:Ligases