Formation of spindle poles by dynein/dynactin-dependent transport of NuMA

NuMA is a large nuclear protein whose relocation to the spindle poles is re quired for bipolar mitotic spindle assembly. We show here that this process depends on directed NuMA transport toward microtubule minus ends powered b y cytoplasmic dynein and its activator dynactin. Upon nuclear envelope brea kdown, large cytoplasmic aggregates of green fluorescent protein (GFP)-tagg ed NuMA stream poleward along spindle fibers in association with the actin- related protein 1 (Arp1) protein of the dynactin complex and cytoplasmic dy nein. Immunoprecipitations and gel filtration demonstrate the assembly of a reversible, mitosis-specific complex of NuMA with dynein and dynactin. NuM A transport is required for spindle pole assembly and maintenance, since di sruption of the dynactin complex (by increasing the amount of the dynamitin subunit) or dynein function (with an antibody) strongly inhibits NuMA tran slocation and accumulation and disrupts spindle pole assembly.