The spindle midzone microtubule-associated proteins Ase1p and Cin8p affect the number and orientation of astral microtubules in Saccharomyces cerevisiae

Abstract

The nucleus of the budding yeast S. cerevisiae has to move to the bud neck during mitosis in order for proper DNA segregation to take place. This movement is mediated by spindle and astral microtubules, and it relies on forces generated by microtubule-associated motor proteins. When budding yeast cells express the non-cleavable cohesin subunit, Scc1-RRDD, sister chromatid separation is blocked, preventing the spindle from elongating. Thus, in the presence of Scc1-RRDD nuclear positioning is mediated solely by forces acting through astral microtubules. We have previously shown that under these conditions cells exit mitosis with the nucleus in the mother cells, and that the position of the nucleus is determined, at least in part, by the FEAR pathway, which regulates various aspects of mitotic exit. When the FEAR pathway is inactivated, cells expressing Scc1-RRDD exit mitosis with the nucleus in the daughter cells (referred to as a "daughterly phenotype"). In order to find additional proteins that participate in nuclear positioning, we screened a series of mutant strains for those that displayed a daughterly phenotype when Scc1-RRDD was expressed. The most prominent defects were seen in ase1delta and cin8delta mutant cells. Both Ase1p and Cin8p were previously shown to be nuclear and to be involved in spindle function. We show here that deletion of ASE1 or CIN8 causes a defect in SPB separation and leads to an abnormal number of astral microtubules and a change in their orientation within the cell. Taken together, these results suggest that in budding yeast Ase1p and Cin8p affect nuclear positioning through astral micro-tubule-dependent mechanisms.