Formation of spindle poles by dynein/dynactin-dependent transport of NuMA.
NuMA is a large nuclear protein whose relocation to the spindle poles is required for bipolar mitotic spindle assembly. We show here that this process depends on directed NuMA transport toward microtubule minus ends powered by cytoplasmic dynein and its activator dynactin. Upon nuclear envelope breakdown, large cytoplasmic aggregates of green fluorescent protein (GFP)-tagged NuMA stream poleward along spindle fibers in association with the actin-related protein 1 (Arp1) protein of the dynactin complex and cytoplasmic dynein. Immunoprecipitations and gel filtration demonstrate the assembly of a reversible, mitosis-specific complex of NuMA with dynein and dynactin. NuMA transport is required for spindle pole assembly and maintenance, since disruption of the dynactin complex (by increasing the amount of the dynamitin subunit) or dynein function (with an antibody) strongly inhibits NuMA translocation and accumulation and disrupts spindle pole assembly.
Merdes
A
A
ICMB, University of Edinburgh, Scotland. a.merdes@ed.ac.uk
Heald
R
R
Samejima
K
K
Earnshaw
W C
WC
Cleveland
D W
DW
D001692:Biological Transport; D002451:Cell Compartmentation; D000072159:Dynactin Complex; D004398:Dyneins / Q000302:isolation & purification / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D008677:Metaphase; D008869:Microtubule-Associated Proteins / Q000302:isolation & purification / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism; D008954:Models, Biological; D009685:Nuclear Envelope / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism; D008941:Spindle Apparatus / Q000378:metabolism*
D000072159:Dynactin Complex; D008164:Luminescent Proteins; D008869:Microtubule-Associated Proteins; D009687:Nuclear Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D004398:Dyneins