Structure of the RZZ complex and molecular basis of its interaction with Spindly.
Kinetochores are macromolecular assemblies that connect chromosomes to spindle microtubules (MTs) during mitosis. The metazoan-specific ≈800-kD ROD-Zwilch-ZW10 (RZZ) complex builds a fibrous corona that assembles on mitotic kinetochores before MT attachment to promote chromosome alignment and robust spindle assembly checkpoint signaling. In this study, we combine biochemical reconstitutions, single-particle electron cryomicroscopy, cross-linking mass spectrometry, and structural modeling to build a complete model of human RZZ. We find that RZZ is structurally related to self-assembling cytosolic coat scaffolds that mediate membrane cargo trafficking, including Clathrin, Sec13-Sec31, and αβ'ε-COP. We show that Spindly, a dynein adaptor, is related to BicD2 and binds RZZ directly in a farnesylation-dependent but membrane-independent manner. Through a targeted chemical biology approach, we identify ROD as the Spindly farnesyl receptor. Our results suggest that RZZ is dynein's cargo at human kinetochores.
Department of Structural Biochemistry, Max Planck Institute of Molecular Physiology, 44227 Dortmund, Germany.
Shyamal
S
Mosalaganti
Department of Mechanistic Cell Biology, Max Planck Institute of Molecular Physiology, 44227 Dortmund, Germany.
Jenny
J
Keller
Department of Mechanistic Cell Biology, Max Planck Institute of Molecular Physiology, 44227 Dortmund, Germany.
Anika
A
Altenfeld
Department of Chemical Biology, Max Planck Institute of Molecular Physiology, 44227 Dortmund, Germany.
Michael
0000-0003-4868-9276
M
Winzker
Gene Center, Ludwig-Maximilians-Universität München, 81377 Munich, Germany.
Pascaline
P
Rombaut
Department of Structural Biochemistry, Max Planck Institute of Molecular Physiology, 44227 Dortmund, Germany.
Michael
0000-0002-6612-6656
M
Saur
Department of Mechanistic Cell Biology, Max Planck Institute of Molecular Physiology, 44227 Dortmund, Germany.
Arsen
A
Petrovic
Department of Mechanistic Cell Biology, Max Planck Institute of Molecular Physiology, 44227 Dortmund, Germany.
Annemarie
0000-0003-2327-6512
A
Wehenkel
Department of Mechanistic Cell Biology, Max Planck Institute of Molecular Physiology, 44227 Dortmund, Germany.
Sabine
S
Wohlgemuth
Department of Mechanistic Cell Biology, Max Planck Institute of Molecular Physiology, 44227 Dortmund, Germany.
Franziska
F
Müller
Department of Mechanistic Cell Biology, Max Planck Institute of Molecular Physiology, 44227 Dortmund, Germany.
Stefano
0000-0001-6380-6560
S
Maffini
Department of Mechanistic Cell Biology, Max Planck Institute of Molecular Physiology, 44227 Dortmund, Germany.
Tanja
0000-0002-9680-8586
T
Bange
Gene Center, Ludwig-Maximilians-Universität München, 81377 Munich, Germany.
Franz
F
Herzog
Department of Chemical Biology, Max Planck Institute of Molecular Physiology, 44227 Dortmund, Germany.|Department of Chemistry and Chemical Biology, Technical University Dortmund, 44227 Dortmund, Germany.
Herbert
H
Waldmann
Department of Structural Biochemistry, Max Planck Institute of Molecular Physiology, 44227 Dortmund, Germany stefan.raunser@mpi-dortmund.mpg.de andrea.musacchio@mpi-dortmund.mpg.de.
Stefan
0000-0001-9373-3016
S
Raunser
Department of Mechanistic Cell Biology, Max Planck Institute of Molecular Physiology, 44227 Dortmund, Germany stefan.raunser@mpi-dortmund.mpg.de andrea.musacchio@mpi-dortmund.mpg.de.|Centre for Medical Biotechnology, Faculty of Biology, University Duisburg-Essen, 45141 Essen, Germany.
Andrea
0000-0003-2362-8784
A
Musacchio
D002352:Carrier Proteins / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000378:metabolism; D045744:Cell Line, Tumor; D004398:Dyneins / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D018386:Kinetochores / Q000378:metabolism* / Q000502:physiology; D059566:M Phase Cell Cycle Checkpoints / Q000502:physiology; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism; D008938:Mitosis / Q000502:physiology; D021381:Protein Transport / Q000502:physiology; D008941:Spindle Apparatus / Q000378:metabolism* / Q000502:physiology*
D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D008869:Microtubule-Associated Proteins; D004398:Dyneins