Tum/RacGAP functions as a switch activating the Pav/kinesin-6 motor.
Centralspindlin is essential for central spindle and cleavage furrow formation. Drosophila centralspindlin consists of a kinesin-6 motor (Pav/kinesin-6) and a GTPase-activating protein (Tum/RacGAP). Centralspindlin localization to the central spindle is mediated by Pav/kinesin-6. While Tum/RacGAP has well-documented scaffolding functions, whether it influences Pav/kinesin-6 function is less well-explored. Here we demonstrate that both Pav/kinesin-6 and the centralspindlin complex (co-expressed Pav/Tum) have strong microtubule bundling activity. Centralspindlin also has robust plus-end-directed motility. In contrast, Pav/kinesin-6 alone cannot move microtubules. However, the addition of Tum/RacGAP or a 65 amino acid Tum/RacGAP fragment to Pav/kinesin-6 restores microtubule motility. Further, ATPase assays reveal that microtubule-stimulated ATPase activity of centralspindlin is seven times higher than that of Pav/kinesin-6. These findings are supported by in vivo studies demonstrating that in Tum/RacGAP-depleted S2 Drosophila cells, Pav/kinesin-6 exhibits severely reduced localization to the central spindle and an abnormal concentration at the centrosomes.
Department of Biology, University of Hawaii at Hilo, 200 West Kawili Street, Hilo, Hawaii 96720, USA.|Department of Molecular, Cellular and Developmental Biology, University of California, Santa Cruz, California 95064, USA.
Li
L
Tao
Department of Molecular, Cellular and Developmental Biology, University of California, Santa Cruz, California 95064, USA.
Barbara
B
Fasulo
Department of Molecular, Cellular and Developmental Biology, University of California, Santa Cruz, California 95064, USA.
Brandt
B
Warecki
Department of Molecular, Cellular and Developmental Biology, University of California, Santa Cruz, California 95064, USA.
William
W
Sullivan
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D018385:Centrosome / Q000378:metabolism; D029721:Drosophila Proteins / Q000235:genetics / Q000378:metabolism*; D004331:Drosophila melanogaster / Q000166:cytology / Q000235:genetics / Q000378:metabolism; D020690:GTPase-Activating Proteins / Q000235:genetics / Q000378:metabolism*; D016547:Kinesins / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D008856:Microscopy, Fluorescence; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism; D020409:Molecular Motor Proteins / Q000235:genetics / Q000378:metabolism*; D046912:Multiprotein Complexes / Q000235:genetics / Q000378:metabolism; D034622:RNA Interference
D029721:Drosophila Proteins; D020690:GTPase-Activating Proteins; D008869:Microtubule-Associated Proteins; D020409:Molecular Motor Proteins; D046912:Multiprotein Complexes; C490516:pav protein, Drosophila; C502951:tum protein, Drosophila; D000251:Adenosine Triphosphatases; D016547:Kinesins