TY - JOUR
T1 - Noncanonical interaction with microtubules via the N-terminal nonmotor domain is critical for the functions of a bidirectional kinesin
AU - Singh, Sudhir K.
AU - Siegler, Nurit
AU - Pandey, Himanshu
AU - Yanir, Neta
AU - Popov, Mary
AU - Goldstein-Levitin, Alina
AU - Sadan, Mayan
AU - Debs, Garrett
AU - Zarivach, Raz
AU - Frank, Gabriel A.
AU - Kass, Itamar
AU - Sindelar, Charles V.
AU - Zalk, Ran
AU - Gheber, Larisa
N1 - Publisher Copyright:
© 2024 American Association for the Advancement of Science. All rights reserved.
PY - 2024/2/1
Y1 - 2024/2/1
N2 - Several kinesin-5 motors (kinesin-5s) exhibit bidirectional motility. The mechanism of such motility remains unknown. Bidirectional kinesin-5s share a long N-terminal nonmotor domain (NTnmd), absent in exclusively plus-end–directed kinesins. Here, we combined in vivo, in vitro, and cryo–electron microscopy (cryo-EM) studies to examine the impact of NTnmd mutations on the motor functions of the bidirectional kinesin-5, Cin8. We found that NTnmd deletion mutants exhibited cell viability and spindle localization defects. Using cryo-EM, we examined the structure of a microtubule (MT)-bound motor domain of Cin8, containing part of its NTnmd. Modeling and molecular dynamic simulations based on the cryo-EM map suggested that the NTnmd of Cin8 interacts with the C-terminal tail of β-tubulin. In vitro experiments on subtilisin-treated MTs confirmed this notion. Last, we showed that NTnmd mutants are defective in plus-end–directed motility in single-molecule and antiparallel MT sliding assays. These findings demonstrate that the NTnmd, common to bidirectional kinesin-5s, is critical for their bidirectional motility and intracellular functions.
AB - Several kinesin-5 motors (kinesin-5s) exhibit bidirectional motility. The mechanism of such motility remains unknown. Bidirectional kinesin-5s share a long N-terminal nonmotor domain (NTnmd), absent in exclusively plus-end–directed kinesins. Here, we combined in vivo, in vitro, and cryo–electron microscopy (cryo-EM) studies to examine the impact of NTnmd mutations on the motor functions of the bidirectional kinesin-5, Cin8. We found that NTnmd deletion mutants exhibited cell viability and spindle localization defects. Using cryo-EM, we examined the structure of a microtubule (MT)-bound motor domain of Cin8, containing part of its NTnmd. Modeling and molecular dynamic simulations based on the cryo-EM map suggested that the NTnmd of Cin8 interacts with the C-terminal tail of β-tubulin. In vitro experiments on subtilisin-treated MTs confirmed this notion. Last, we showed that NTnmd mutants are defective in plus-end–directed motility in single-molecule and antiparallel MT sliding assays. These findings demonstrate that the NTnmd, common to bidirectional kinesin-5s, is critical for their bidirectional motility and intracellular functions.
UR - http://www.scopus.com/inward/record.url?scp=85184683802&partnerID=8YFLogxK
U2 - 10.1126/sciadv.adi1367
DO - 10.1126/sciadv.adi1367
M3 - Article
C2 - 38324691
AN - SCOPUS:85184683802
SN - 2375-2548
VL - 10
JO - Science advances
JF - Science advances
IS - 6
M1 - eadi1367
ER -