Regulation of microtubule assembly and organization in mitosis by the AAA+ ATPase Pontin.
To identify novel proteins important for microtubule assembly in mitosis, we have used a centrosome-based complementation assay to enrich for proteins with mitotic functions. An RNA interference (RNAi)-based screen of these proteins allowed us to uncover 13 novel mitotic regulators. We carried out in-depth analyses of one of these proteins, Pontin, which is known to have several functions in interphase, including chromatin remodeling, DNA repair, and transcription. We show that reduction of Pontin by RNAi resulted in defects in spindle assembly in Drosophila S2 cells and in several mammalian tissue culture cell lines. Further characterization of Pontin in Xenopus egg extracts demonstrates that Pontin interacts with the gamma tubulin ring complex (gamma-TuRC). Because depletion of Pontin leads to defects in the assembly and organization of microtubule arrays in egg extracts, our studies suggest that Pontin has a mitosis-specific function in regulating microtubule assembly.
Ducat
Daniel
D
Department of Embryology, Carnegie Institution for Science and Howard Hughes Medical Institute, Baltimore, MD 21210, USA.
Kawaguchi
Shin-Ichi
S
Liu
Hongbin
H
Yates
John R
JR
Zheng
Yixian
Y
D000074183:ATPases Associated with Diverse Cellular Activities; D000818:Animals; D001671:Biochemistry / Q000379:methods; D002352:Carrier Proteins / Q000378:metabolism*; D002843:Chromatin / Q000737:chemistry; D004265:DNA Helicases / Q000378:metabolism*; D004260:DNA Repair; D004330:Drosophila; D029721:Drosophila Proteins / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D008870:Microtubules / Q000378:metabolism* / Q000648:ultrastructure; D008938:Mitosis; D008954:Models, Biological; D034622:RNA Interference; D014981:Xenopus; D029867:Xenopus Proteins / Q000378:metabolism*
D002352:Carrier Proteins; D002843:Chromatin; D029721:Drosophila Proteins; D029867:Xenopus Proteins; D000074183:ATPases Associated with Diverse Cellular Activities; D004265:DNA Helicases; C486549:RUVBL1 protein, human; C486548:Ruvbl1 protein, Xenopus; C504804:pont protein, Drosophila