Bub1 and Bub3 are required for normal mitotic progression in Drosophila neuroblasts. C.E. Sunkel ¹, C. Lopes ¹, H. Bousbaa ¹, B. Williams ², M. Costa ¹, M. Goldberg ². 1) Instituto de Biologia Molecular e Celular, Universidade Do Porto, Porto, Portugal; 2) Section of Genetics and Development, Cornell University, USA.

   High fidelity of chromosome segregation is essential for normal cellular viability. The spindle checkpoint involves a signalling system that monitors the attachment of spindle microtubules to the kinetochores of mitotic chromosomes and does not allow progression into anaphase until all kinetochores become properly attached. Previously we identified a homologue for BUB1 and showed that null mutations lead to accelerated entry to anaphase leading to chromosome breakage and apoptosis. We also identified a homologue for BUB3 and showed that both BUB1 and BUB3 accumulate to the unattached kinetochores early in mitosis until the metaphase-anaphase transition when staining is significantly reduced. More recently, we have raised new polyclonal antibodies against both proteins and show that BUB1 and BUB3 co-localise to kinetochores before NEB. Furthermore, we identified a mutant allele of Bub3 and show that loss of Bub3 does not lead to chromosome breakage and apoptosis but to premature sister chromatid separation and significant aneuploidy. Immunolocalisation studies also show that BUB1 does not localise properly in Bub3 mutant cells. Therefore, even though BUB1 and BUB3 have been shown to form a complex and are mutually required for correct localisation, the loss of Bub3 is not equivalent to the loss of Bub1. Analysis of strains carrying mutations for both Bub1 and Bub3 confirm these results and suggest that Bub3 is epistatic to Bub1. Moreover, analysis of cyclin B accumulation in the single and double mutant combinations indicates that these proteins are involved in regulating mitotic progression by regulating the activity of the Anaphase Promoting Complex.