A dependent pathway of gene functions leading to chromosome segregation in Saccharomyces cerevisiae.

Wood, John S.; Hartwell, Leland H.

doi:10.1083/jcb.94.3.718

Cited by 92 publications

(62 citation statements)

References 24 publications

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“…We observed three features that are important for optimizing drug treatment conditions. First, diploid strains responded more uniformly to all of the drugs tested than did haploids, also noted by Wood and Hartwell (24) in their studies with a related compound, methyl benzimidazol-2-yl-carbamate. Second, cells treated in liquid culture with nocodazole responded most completely when the drug was added in early logarithmic phase.…”

Section: Resultsmentioning

confidence: 91%

Components of microtubular structures in Saccharomyces cerevisiae.

Pillus¹,

Solomon²

1986

Proc. Natl. Acad. Sci. U.S.A.

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Most studies of cytoskeletal organelles have concentrated on molecular analyses of abundant and biochemically accessible structures. In many of the classical cases, however, the nature of the system chosen has precluded a concurrent genetic analysis. The mitotic spindle of the yeast Saccharomyces cerevisiae is one example of an organelle that can be studied by both classical and molecular genetics. We show here that this microtubule structure also can be examined biochemically. The spindle can be isolated by selective extractions of yeast cells by using adaptations of methods successfully applied to animal cells. In this way, microtubule-associated proteins of the yeast spindle are identified.Microtubule-associated proteins are the focus of much attention, for it seems plausible that these minor components of microtubule structures may be the factors that help to define specific parameters of the repertoire of microtubule organelles: state of assembly, localization, organization, interaction, and regulation offunction. This suggestion arises from the properties of associated proteins in vitro and from unique localizations of some of them in vivo. It has not yet been possible to integrate these sorts of results into a unified picture of precise microtubule functions in vivo. For this reason, increasing interest is focused on those organisms with genetic systems that may serve to bridge the gulf between molecular characterizations and cellular functions.The budding yeast Saccharomyces provides an especially attractive system for such studies. Its mitotic cell cycle has been well characterized by electron microscopy (1-3), light microscopy (4), and immunofluorescence microscopy (5).Mutations and suppressors of mutations in the f-tubulin gene have been defined, as have a variety of conditional mutations which lead to cell-cycle arrests (6-8). Tubulin can be purified from yeast extracts (both Saccharomyces cerevisiae and Saccharomyces uvarum), relying largely upon the in vitro assembly reaction that has been developed for isolating tubulin from several types ofanimal cells and tissues (9). That the yeast tubulin assembles under the same conditions as those applied to other tubulins demonstrates a common property, but there are important differences as well. In particular, Kilmartin (9) noted a different number ofprotofilaments in the assembled microtubules and different sensitivities to cold and to microtubule-depolymerizing drugs. In the filamentous fungus Aspergillus nidulans (10), focus has been primarily on the tubulin components of its microtubule systems; no associated proteins have been identified.In this paper, we have used an alternative approach to identifying cytoplasmic microtubule components. We describe here the successful application to yeast of a technique that has been used to define microtubule-associated proteins in a variety of vertebrate cells (11). This technique has been used to identify species, cell-type, and cell cycle-specific associated proteins (12,13). Antisera to some of these p...

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Section: Resultsmentioning

confidence: 91%

Components of microtubular structures in Saccharomyces cerevisiae.

Pillus¹,

Solomon²

1986

Proc. Natl. Acad. Sci. U.S.A.

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“…Dbf4 is required for the initiation of DNA replication, and mutations affecting the essential function of either CDC7 or DBF4 affect entry into and progression through S-phase ( Johnston and Thomas 1982;Wood and Hartwell 1982). Cdc7-Dbf4 also affects S-phase progression since it is needed throughout S-phase to activate DNA synthesis from individual replication origins (Bousset and Diffley 1998; Donaldson et al 1998).…”

Section: Resultsmentioning

confidence: 99%

Budding Yeast Dbf4 Sequences Required for Cdc7 Kinase Activation and Identification of a Functional Relationship Between the Dbf4 and Rev1 BRCT Domains

Harkins¹,

Gabrielse²,

Haste³

et al. 2009

Genetics

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Cdc7-Dbf4 is a two-subunit kinase required for initiating DNA replication. The Dbf4 regulatory subunit is required for Cdc7 kinase activity. Previous studies have shown that the C termini of Dbf4 orthologs encode a single (putative) C 2 H 2 zinc (Zn) finger, referred to as ''motif C.'' By mutational analysis we show that the Zn finger is not required for the essential function of Dbf4. However, deletion and point mutants altering conserved Zn-finger residues exhibit a substantially slowed S-phase, DNA damage sensitivity, and a hypo-mutagenic phenotype following UV irradiation. Using two-hybrid and biochemical assays, we show that the Dbf4 Zn finger interacts with Cdc7 and stimulates its kinase activity. However, a separable Dbf4 region also mediates an interaction with Cdc7 such that only the loss of both Cdc7-interacting regions results in lethality. In contrast, an N-terminal BRCT-like domain is not required for induced mutagenesis nor does it interact with Cdc7. By making chimeric Dbf4 proteins that contain known BRCT domains in Saccharomyces cerevisiae, we show that the BRCT domain from Rev1, a translesion DNA polymerase, can uniquely substitute for the Dbf4 BRCT domain. Thus, we have mapped regions on budding yeast Dbf4 required for binding and activating Cdc7 kinase. Our data also suggest that the Dbf4 and Rev1 BRCT domains interact with a common protein or structure, although the precise function of both domains and their binding partners remains elusive.

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“…It is therefore possible that S. cerevisiae might have abandoned one function of its polo-like kinase or that another related kinase might have become specialized for this role, as with the divergence of cdc2 + and cdks in mammalian cells. Although there appears not to be an effect on establishment of bipolar spindle in S. cerevisiae in the currently existing alleles of cdc5, we do note that the CDC5 mutation may have an effect on microtubule behavior, because following the release of a temperature-sensitive cdc5 strain from a block at the nonpermissive temperature, cells become insensitive to the microtubule depolymerizing drug MBC (Wood and Hartwell 1982). However, the very fact that CDC5 and CDC15 affect similar events in the budding yeast and that their fission yeast genes plol + and cdc7 + are both required for septation suggest that the sequential action of these two genes might be required in other species to mediate a fundamental mitotic process.…”

Section: Conservation Of Plol + Functionmentioning

confidence: 99%

“…Although the activity of CDC5 kinase has not been determined with respect to cell cycle progression, its transcript is periodically accumulated around G2/M (Kitada et al 1993). Some circumstantial observations suggest that polo/CDC5 may regulate microtubule behavior, cdc5 mutants show an unusual interaction with MBC, a drug that binds tubulin and depolymerizes microtubules (Wood and Hartwell 1982), and there is a strong interaction of polo with mu- …”

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confidence: 99%

The conserved Schizosaccharomyces pombe kinase plo1, required to form a bipolar spindle, the actin ring, and septum, can drive septum formation in G1 and G2 cells.

Ohkura¹,

Hagan²,

Glover³

1995

Genes Dev.

360

395

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We have identified a Schizosaccharomyces pombe gene with homology to the budding yeast gene CDCS, the Drosophila gene polo, and the mammalian family of genes encoding polo-like kinases. Disruption of this gene, plol +, indicates that it is essential. Loss of plol + function leads to a mitotic arrest in which condensed chromsomes are associated with a monopolar spindle or to the failure of septation following the completion of nuclear division. In the latter case, cells show a failure both in the formation of an F-actin ring and in the deposition of septal material, suggesting that plol + function is required high in the regulatory cascade that controls septation. The overexpression of plol + in wild-type cells also results in the formation of monopolar spindles but also induces the formation of multiple septa without nuclear division. Septation can also be induced in the absence of mitotic commitment and concomitant spindle formation by the overexpression of plol + in cdc25-22 or cdc2-33 cells arrested in G2; in GI cells arrested at Start by the cdclO-V50 mutation, or in cells lacking the cyclin B homolog cdcl3 that undergo repeated S phases in the absence of mitosis.[Key Words: S. pombe; polo homolog; bipolar spindle; actin ring; septum; plol kinase] Received January 20, 1995; revised version accepted March 16, 1995.The polo ~ mutation of Drosophila was first identified as a maternal-effect mutant giving rise to embryos that show abnormal networks of microtubules associated with condensed chromosomes, in which the centrosomal antigen CP190 {formerly called Bx63; Frasch et al. 1986;Whitfield et al. 1988) fails to become organized into centrosomes with microtubule-organizing capability (Sunkel and Glover 1988). The mutation results in multipolar spindles and nondisjunction in male meiosis leading to reduced fertility. There is a high mitotic index in larval neuroblasts, with a wide range of defects including monopolar spindles and spindles with broad poles (Sunkel and Glover 1988;Llamazares et al. 1991).polo encodes a 576-amino-acid protein that has an amino-terminal putative kinase domain and a 300-residue carboxy-terminal domain (Llamazares et al. 1991). The polo kinase is highly conserved. The budding yeast and murine genes, CDC5 (Kitada et al. 1993) and mouse Plk (polo-like kinase; Clay et al. 1993;Lake and Jelinek 1993;Hamanaka et al. 1994), for example, show 52% and 65% identity in the kinase domain and 33% and 43% in the carboxy-terminal domain, with the latter homolo3Corresponding author. gies lying in distinct blocks. The human Plk has also been identified (Golsteyn et al. 1994;Hamanaka et al. 1994;Holtrich et al. 1994), and studies in the mouse suggest that there is another closely related kinase, Snk (serum-induced kinase; Simmons et al. 1992), that falls into the same gene family. These mammalian genes are expressed only in proliferating cells or tissues suggesting a cell cycle role.In mutants of the budding yeast gene CDC5, nuclear division is arrested at a late stage and the spindle is elongated (Ha...

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A dependent pathway of gene functions leading to chromosome segregation in Saccharomyces cerevisiae.

Cited by 92 publications

References 24 publications

Components of microtubular structures in Saccharomyces cerevisiae.

Components of microtubular structures in Saccharomyces cerevisiae.

Budding Yeast Dbf4 Sequences Required for Cdc7 Kinase Activation and Identification of a Functional Relationship Between the Dbf4 and Rev1 BRCT Domains

The conserved Schizosaccharomyces pombe kinase plo1, required to form a bipolar spindle, the actin ring, and septum, can drive septum formation in G1 and G2 cells.

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