K11-linked polyubiquitination in cell cycle control revealed by a K11 linkage-specific antibody.
Polyubiquitination is a posttranslational modification where ubiquitin chains containing isopeptide bonds linking one of seven ubiquitin lysines with the C terminus of an adjoining ubiquitin are covalently attached to proteins. While functions of K48- and K63-linked polyubiquitin are understood, the role(s) of noncanonical K11-linked chains is less clear. A crystal structure of K11-linked diubiquitin demonstrates a distinct conformation from K48- or K63-linked diubiquitin. We engineered a K11 linkage-specific antibody and use it to demonstrate that K11 chains are highly upregulated in mitotic human cells precisely when substrates of the ubiquitin ligase anaphase-promoting complex (APC/C) are degraded. These chains increased with proteasomal inhibition, suggesting they act as degradation signals in vivo. Inhibition of the APC/C strongly impeded the formation of K11-linked chains, suggesting that a single ubiquitin ligase is the major source of mitotic K11-linked chains. Our results underscore the importance of K11-linked ubiquitin chains as critical regulators of mitotic protein degradation.
Matsumoto
Marissa L
ML
Department of Antibody Engineering, Genentech, Inc., South San Francisco, CA 94080, USA.
Wickliffe
Katherine E
KE
Dong
Ken C
KC
Yu
Christine
C
Bosanac
Ivan
I
Bustos
Daisy
D
Phu
Lilian
L
Kirkpatrick
Donald S
DS
Hymowitz
Sarah G
SG
Rape
Michael
M
Kelley
Robert F
RF
Dixit
Vishva M
VM
D064173:Anaphase-Promoting Complex-Cyclosome; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000276:immunology / Q000494:pharmacology*; D002453:Cell Cycle / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D011499:Protein Processing, Post-Translational / Q000502:physiology*; D025801:Ubiquitin / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D043743:Ubiquitin-Protein Ligase Complexes / Q000378:metabolism*; D054875:Ubiquitination / Q000502:physiology*
D000911:Antibodies, Monoclonal; D025801:Ubiquitin; D043743:Ubiquitin-Protein Ligase Complexes; D064173:Anaphase-Promoting Complex-Cyclosome