title: PARC and CUL7 form atypical cullin RING ligase complexes.
abstract: CUL7 and the p53-associated, PARkin-like cytoplasmic protein (PARC) were previously reported to form homodimers and heterodimers, the first demonstration of cullin dimerization. Although a CUL7-based SKP1/CUL1/F-box (SCF)-like complex has been observed, little is known about the existence of a PARC-based SCF-like complex and how PARC interacts with CUL7-based complexes. To further characterize PARC-containing complexes, we examined the ability of PARC to form an SCF-like complex. PARC binds RBX1 and is covalently modified by NEDD8, defining PARC as a true cullin. However, PARC fails to bind SKP1 or F-box proteins, including the CUL7-associated FBXW8. To examine the assembly of PARC- and CUL7-containing complexes, tandem affinity purification followed by multidimensional protein identification technology were used. Multidimensional protein identification technology analysis revealed that the CUL7 interaction with FBXW8 was mutually exclusive of CUL7 binding to PARC or p53. Notably, although heterodimers of CUL7 and PARC bind p53, p53 is not required for the dimerization of CUL7 and PARC. The observed physical separation of FBXW8 and PARC is supported functionally by the generation of Parc-/-, Fbxw8-/- mice, which do not show exacerbation of the Fbxw8-/- phenotype. Finally, all of the PARC and CUL7 subcomplexes examined exhibit E3 ubiquitin ligase activity in vitro. Together, these findings indicate that the intricate assembly of PARC- and CUL7-containing complexes is highly regulated, and multiple subcomplexes may exhibit ubiquitin ligase activity.
authors.affiliation: Department of Medical Oncology, Dana-Farber Cancer Institute, 44 Binney Street, Boston, MA 02115, USA.
authors.forename: Jeffrey R
authors.initials: JR
authors.lastname: Skaar
authors.forename: Laurence
authors.initials: L
authors.lastname: Florens
authors.forename: Takeya
authors.initials: T
authors.lastname: Tsutsumi
authors.forename: Takehiro
authors.initials: T
authors.lastname: Arai
authors.forename: Adriana
authors.initials: A
authors.lastname: Tron
authors.forename: Selene K
authors.initials: SK
authors.lastname: Swanson
authors.forename: Michael P
authors.initials: MP
authors.lastname: Washburn
authors.forename: James A
authors.initials: JA
authors.lastname: DeCaprio
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism*; D002478:Cells, Cultured; D044842:Cullin Proteins / Q000378:metabolism*; D045325:HCT116 Cells; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008969:Molecular Sequence Data; D046912:Multiprotein Complexes / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D017386:Sequence Homology, Amino Acid; D014166:Transferases; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry* / Q000378:metabolism*
chemical_list: C506299:CUL7 protein, human; D002352:Carrier Proteins; D044842:Cullin Proteins; D046912:Multiprotein Complexes; C400192:Cul9 protein, human; D014166:Transferases; D044767:Ubiquitin-Protein Ligases