Role of astral microtubules and actin in spindle orientation and migration in the budding yeast, Saccharomyces cerevisiae.

Abstract

Abstract. In the yeast Saccharomyces cerevisiae, before the onset of anaphase, the spindle apparatus is always positioned with one spindle pole at, or through, the neck between the mother cell and the growing bud. This spindle orientation enables proper chromosome segregation to occur during anaphase, allowing one replicated genome to be segregated into the bud and the other to remain in the mother cell. In this study, we synchronized a population of cells before the onset of anaphase such that >90% of the cells in the population had spindles with the correct orientation, and then disrupted specific cytoskeletal elements using temperature-sensitive mutations. Disruption of either the astral microtubules or actin function resulted in improper spindle orientation in "~40-50% of the cells. When cells with disrupted astral microtubules or actin function entered into anaphase, there was a 100-200-fold increase in the frequency of binucleated cell bodies. Thus, the maintenance of proper spindle orientation by these cytoskeletal elements was essential for proper chromosome segregation. These data are consistent with the model that proper spindle orientation is maintained by directly or indirectly tethering the astral microtubules to the actin cytoskeleton.After nuclear migration, but before anaphase, bulk chromosome movement occurs within the nucleus apparently because the chromosomes are attached to a mobile spindle. The frequency and magnitude of bulk chromosome movement is greatly diminished by disruption of the astral microtubules but not by disruption of the nonkinetochore spindle microtubules. These results suggest that astral microtubules are not only important for spindle orientation before anaphase, but they also mediate force on the spindle, generating spindle displacement and in turn chromosome movement. Potential roles for this force in spindle assembly and orientation are discussed. CELL division in the yeast, Saccharomyces cerevisiae, is an asymmetric process in which a bud emerges from a discrete region of the periphery of the parental, "mother" cell. Before anaphase the nucleus migrates from a location in the center of the mother cell to a position at the isthmus, or neck, between the mother cell and the growing bud (Pringle and Hartwell, 1981), and the nuclear envelope becomes extended between the mother cell and bud. Embedded in the nuclear envelope are the two separated spindle pole bodies (SPBs) ~ (Byers and Goetsch, 1974;Byers and Goetsch, 1975). Emanating from the lumenal face of each SPB into the nucleus are microtubules that form the bipolar spindle (Byers and Goetsch, 1974). Radiating away from the cytoplasmic face of each SPB towards the cell periphery are astral or cytoplasmic microtubules. Before the onset of anaphase, the bipolar spindle is always positioned with one SPB at, or through, the neck (Byers 1. Abbreviations used in this paper: DIM, digital imaging microscopy; HU, hydroxyurea; SPB, spindle pole body.and Goetsch, 1975;Pringle and Hartwell, 1981). At the onset of anaphase, the spin...

Profilin is required for the normal timing of actin polymerization in response to thermal stress

“…Thus, the fluorescence measurement is an indirect indication of the relative amount of F-actin in the cell at the time of fixation. Using this method, Lillie and Brown [17] have shown that a shift to high temperature causes a rapid but transient depolymerization of actin, which is accompanied by a transient redistribution of actin-containing structures [17,19,20]. Similar effects on actin organization have been noted upon increasing the osmotic strength of the medium [211.…”

Three-dimensional analysis and ultrastructural design of mitotic spindles from the cdc20 mutant of Saccharomyces cerevisiae.

“…Many but not all MT-related events are affected, including the formation of a normal mitotic spindle and thus normal chromosome segregation (Hartwell et al, 1973;Byers and Goetsch 1974;Sethi et al, 1991) and karyogamy (Sethi et al, 1991). Movement of the nucleus to the bud neck, which depends on the presence of astral MTs (Palmer et al, 1992), is not affected in cdc2O cells, but this role of MTs may not rely on dynamic MT behavior. Although it is unlikely that the Cdc2Op is associated directly with MTs, its role may be to promote MT disassembly through a mechanism that also affects the proteolysis of proteins in other pathways.…”

An actin point‐mutation neighboring the ‘hydrophobic plug’ causes defects in the maintenance of cell polaroty and septum organization in the fission yeast Schizosaccharomyces pombe

“…The yeast actin cytoskeleton has been shown to be involved, not only in late mitotic events such as septum formation and cytokinesis, but also in directing cell wall deposition at the growing region of the cell [5,6]. In the budding yeast Schizosaccharomyces cerevisiae, actin function is also required to maintain proper spindle orientation prior to the onset of anaphase [7]. All these events are, thus, essential for proper chromosome distribution, by which daughter cells inherit a complete set of genetic information required for the next cell cycle.…”