Dynamic release of nuclear RanGTP triggers TPX2-dependent microtubule assembly during the apoptotic execution phase.
During apoptosis, the interphase microtubule network is dismantled then later replaced by a novel, non-centrosomal microtubule array. These microtubules assist in the peripheral redistribution of nuclear fragments in the apoptotic cell; however, the regulation of apoptotic microtubule assembly is not understood. Here, we demonstrate that microtubule assembly depends upon the release of nuclear RanGTP into the apoptotic cytoplasm because this process is blocked in apoptotic cells overexpressing dominant-negative GDP-locked Ran (T24N). Actin-myosin-II contractility provides the impetus for Ran release and, consequently, microtubule assembly is blocked in blebbistatin- and Y27632-treated apoptotic cells. Importantly, the spindle-assembly factor TPX2 (targeting protein for Xklp2), colocalises with apoptotic microtubules, and siRNA silencing of TPX2, but not of the microtubule motors Mklp1 and Kid, abrogates apoptotic microtubule assembly. These data provide a molecular explanation for the assembly of the apoptotic microtubule network, and suggest important similarities with the process of RanGTP- and TPX2-mediated mitotic spindle formation.
Moss
David K
DK
Cell Biology Laboratories, Department of Biochemistry, University of Bristol, School of Medical Sciences, University Walk, Bristol BS8 1TD, UK.
Wilde
Andrew
A
Lane
Jon D
JD
D000199:Actins / Q000378:metabolism; D000903:Antibiotics, Antineoplastic / Q000378:metabolism; D017209:Apoptosis / Q000502:physiology*; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism*; D005231:Fatty Acids, Unsaturated / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism*; D024462:Myosin Type II / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D008941:Spindle Apparatus / Q000378:metabolism; D020931:ran GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*
D000199:Actins; D000903:Antibiotics, Antineoplastic; D018797:Cell Cycle Proteins; D005231:Fatty Acids, Unsaturated; D008869:Microtubule-Associated Proteins; D009687:Nuclear Proteins; D034741:RNA, Small Interfering; D011993:Recombinant Fusion Proteins; C122167:TPX2 protein, human; D024462:Myosin Type II; D020931:ran GTP-Binding Protein; C038753:leptomycin B