PULSE TREATMENT OF INTERPHASIC HELA-CELLS WITH NANOMOLAR DOSES OF DOCETAXEL AFFECTS CENTROSOME ORGANIZATION AND LEADS TO CATASTROPHIC EXIT OF MITOSIS

In order to investigate the role of centrosome duplication in mitotic spindle morphogenesis, we designed a 1 hour pulse treatment protocol o n synchronized HeLa cells with nanomolar doses of taxoids that might i mpair centrosome biogenesis but would allow the recovery of normal mic rotubule (Mt) dynamics before mitosis, We were prompted to use this ap proach as docetaxel (DOC; taxotere(TM))), a taxoid known to promote Mt polymerization, was shown to be more cytotoxic when applied during S phase, We show that pulse drug exposure is most efficient in late S an d in G(2) and results in a marked disorganization of the centrosome in G(2), the pericentriolar material (PCM) being dissociated from centri oles, Separation of centrosomes at the G(2)-M transition is also impai red and mitotic spindle morphogenesis is grossly abnormal: although in most spindles chromosomes align in a metaphase plate, the two centros omes stay most often unseparated at one pole and most of the NuMA prot ein accumulates at the other, Interestingly, we find that the centroso mes' ability to duplicate is not abolished as they are still able to t rigger parthenogenetic development of frog eggs, Despite spindle asymm etry, the progression through mitosis is not blocked, This results in a catastrophic exit from mitosis, each mitotic cell generating several micronucleated cells linked together by multiple midbodies, Lack of m itotic block appears therefore as the prime cause of cell lethality, T hese experiments suggest that NuMA redistribution at the onset of mito sis depends upon the correct redistribution of PCM between centriole p airs, They also indicate that the presence of aberrant spindle poles d oes not alert the surveillance mechanism controlling the exit of mitos is.