Cyclin G-associated kinase promotes microtubule outgrowth from chromosomes during spindle assembly.
During mitosis, all chromosomes must attach to microtubules of the mitotic spindle to ensure correct chromosome segregation. Microtubule attachment occurs at specialized structures at the centromeric region of chromosomes, called kinetochores. These kinetochores can generate microtubule attachments through capture of centrosome-derived microtubules, but in addition, they can generate microtubules themselves, which are subsequently integrated with centrosome-derived microtubules to form the mitotic spindle. Here, we have performed a large scale RNAi screen and identify cyclin G-associated kinase (GAK) as a novel regulator of microtubule generation at kinetochores/chromatin. This function of GAK requires its C-terminal J-domain, which is essential for clathrin recycling from endocytic vesicles. Consistently, cells lacking GAK show strongly reduced levels of clathrin on the mitotic spindle, and reduction of clathrin levels also inhibits microtubule generation at kinetochores/chromosomes. Finally, we present evidence that association of clathrin with the spindle is promoted by a signal coming from the chromosomes. These results identify a role for GAK and clathrin in microtubule outgrowth from kinetochores/chromosomes and suggest that GAK acts through clathrin to control microtubule outgrowth around chromosomes.
Department of Medical Oncology, University Medical Center Utrecht, The Netherlands.
Marvin E
ME
Tanenbaum
Tea
T
Vallenius
Erica F
EF
Geers
Lois
L
Greene
Tomi P
TP
Mäkelä
Rene H
RH
Medema
D045744:Cell Line, Tumor; D002503:Centromere / Q000378:metabolism; D020090:Chromosome Segregation / Q000502:physiology; D002877:Chromosomes, Human / Q000235:genetics / Q000502:physiology / Q000648:ultrastructure; D002966:Clathrin / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D006367:HeLa Cells; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism*; D018386:Kinetochores / Q000502:physiology; D008870:Microtubules / Q000378:metabolism* / Q000648:ultrastructure; D008938:Mitosis; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D034741:RNA, Small Interfering; D008941:Spindle Apparatus / Q000235:genetics / Q000378:metabolism*; D014404:Tubulin / Q000378:metabolism
D002966:Clathrin; D047908:Intracellular Signaling Peptides and Proteins; D034741:RNA, Small Interfering; D014404:Tubulin; C486325:GAK protein, human; D017346:Protein Serine-Threonine Kinases