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The yeast motor protein, Kar3p, is essential for meiosis I.

Bascom-Slack CA, Dawson DS - J. Cell Biol. (1997)

Bottom Line: The recognition and alignment of homologous chromosomes early in meiosis is essential for their subsequent segregation at anaphase I; however, the mechanism by which this occurs is unknown.We demonstrate here that, in the absence of the molecular motor, Kar3p, meiotic cells are blocked with prophase monopolar microtubule arrays and incomplete synaptonemal complex (SC) formation. kar3 mutants exhibit very low levels of heteroallelic recombination. kar3 mutants do produce double-strand breaks that act as initiation sites for meiotic recombination in yeast, but at levels severalfold reduced from wild-type.These data are consistent with a meiotic role for Kar3p in the events that culminate in synapsis of homologues.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology and Microbiology, Tufts University School of Medicine, Boston, Massachusetts 02111, USA.

ABSTRACT
The recognition and alignment of homologous chromosomes early in meiosis is essential for their subsequent segregation at anaphase I; however, the mechanism by which this occurs is unknown. We demonstrate here that, in the absence of the molecular motor, Kar3p, meiotic cells are blocked with prophase monopolar microtubule arrays and incomplete synaptonemal complex (SC) formation. kar3 mutants exhibit very low levels of heteroallelic recombination. kar3 mutants do produce double-strand breaks that act as initiation sites for meiotic recombination in yeast, but at levels severalfold reduced from wild-type. These data are consistent with a meiotic role for Kar3p in the events that culminate in synapsis of homologues.

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Analysis of tubulin  and DNA staining pattern of  KAR3 and kar3 cells throughout meiosis. Aliquots of  isogenic wild-type (DJ1 +  pD154.12) and kar3 (DJ1)  cells were fixed at the indicated times after transfer to  sporulation medium and  stained with antitubulin antibodies and the DNA-specific dye DAPI. Antitubulin  and DAPI images of representative cells are shown for  both wild-type (top) and kar3  (bottom) time courses. Arrows in the 5-h kar3 samples  indicate pronounced cytoplasmic projections. The arrow in the 24-h kar3 panel  identifies a highly abnormal  spindle. Small numbers of  unusual spindles were observed in all time points for  the kar3 culture. Bar, 10 μm.
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Figure 1: Analysis of tubulin and DNA staining pattern of KAR3 and kar3 cells throughout meiosis. Aliquots of isogenic wild-type (DJ1 + pD154.12) and kar3 (DJ1) cells were fixed at the indicated times after transfer to sporulation medium and stained with antitubulin antibodies and the DNA-specific dye DAPI. Antitubulin and DAPI images of representative cells are shown for both wild-type (top) and kar3 (bottom) time courses. Arrows in the 5-h kar3 samples indicate pronounced cytoplasmic projections. The arrow in the 24-h kar3 panel identifies a highly abnormal spindle. Small numbers of unusual spindles were observed in all time points for the kar3 culture. Bar, 10 μm.

Mentions: It has been reported that kar3 mutants fail to produce spores when assayed using bright field microscopy (Hoyt et al., 1993; Kurihara et al., 1996). To determine the stage of the meiotic block, homozygous kar3 mutant cells were analyzed by observing the spindle morphology and DNA staining patterns throughout meiosis (Fig. 1). At 5 h after the induction of meiosis, wild-type cells exhibit tubulin arrays typical of yeast prophase spindles (microtubules emanating from duplicated, unseparated SPBs). At the same time, most mutant cells possess similar tubulin arrays that appear slightly longer than those in wild-type cells. Additionally, the mutants often have elongated cytoplasmic microtubules that are not seen in wild-type cells (Fig. 1, 5 h), indicating that the kar3 mutation has an effect early in prophase. By 12 h most wild-type cells have spindles characteristic of metaphase I or later stages of meiosis, and at 24 h the wild-type cells have nearly all formed tetrads. In contrast, kar3 mutants continue to exhibit bush-like tubulin staining. At late time points (24 h and beyond, not shown), monopolar arrays with numerous and/or very long projections become more common (an example is shown in the 24-h time point, Fig. 1).


The yeast motor protein, Kar3p, is essential for meiosis I.

Bascom-Slack CA, Dawson DS - J. Cell Biol. (1997)

Analysis of tubulin  and DNA staining pattern of  KAR3 and kar3 cells throughout meiosis. Aliquots of  isogenic wild-type (DJ1 +  pD154.12) and kar3 (DJ1)  cells were fixed at the indicated times after transfer to  sporulation medium and  stained with antitubulin antibodies and the DNA-specific dye DAPI. Antitubulin  and DAPI images of representative cells are shown for  both wild-type (top) and kar3  (bottom) time courses. Arrows in the 5-h kar3 samples  indicate pronounced cytoplasmic projections. The arrow in the 24-h kar3 panel  identifies a highly abnormal  spindle. Small numbers of  unusual spindles were observed in all time points for  the kar3 culture. Bar, 10 μm.
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Related In: Results  -  Collection

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Figure 1: Analysis of tubulin and DNA staining pattern of KAR3 and kar3 cells throughout meiosis. Aliquots of isogenic wild-type (DJ1 + pD154.12) and kar3 (DJ1) cells were fixed at the indicated times after transfer to sporulation medium and stained with antitubulin antibodies and the DNA-specific dye DAPI. Antitubulin and DAPI images of representative cells are shown for both wild-type (top) and kar3 (bottom) time courses. Arrows in the 5-h kar3 samples indicate pronounced cytoplasmic projections. The arrow in the 24-h kar3 panel identifies a highly abnormal spindle. Small numbers of unusual spindles were observed in all time points for the kar3 culture. Bar, 10 μm.
Mentions: It has been reported that kar3 mutants fail to produce spores when assayed using bright field microscopy (Hoyt et al., 1993; Kurihara et al., 1996). To determine the stage of the meiotic block, homozygous kar3 mutant cells were analyzed by observing the spindle morphology and DNA staining patterns throughout meiosis (Fig. 1). At 5 h after the induction of meiosis, wild-type cells exhibit tubulin arrays typical of yeast prophase spindles (microtubules emanating from duplicated, unseparated SPBs). At the same time, most mutant cells possess similar tubulin arrays that appear slightly longer than those in wild-type cells. Additionally, the mutants often have elongated cytoplasmic microtubules that are not seen in wild-type cells (Fig. 1, 5 h), indicating that the kar3 mutation has an effect early in prophase. By 12 h most wild-type cells have spindles characteristic of metaphase I or later stages of meiosis, and at 24 h the wild-type cells have nearly all formed tetrads. In contrast, kar3 mutants continue to exhibit bush-like tubulin staining. At late time points (24 h and beyond, not shown), monopolar arrays with numerous and/or very long projections become more common (an example is shown in the 24-h time point, Fig. 1).

Bottom Line: The recognition and alignment of homologous chromosomes early in meiosis is essential for their subsequent segregation at anaphase I; however, the mechanism by which this occurs is unknown.We demonstrate here that, in the absence of the molecular motor, Kar3p, meiotic cells are blocked with prophase monopolar microtubule arrays and incomplete synaptonemal complex (SC) formation. kar3 mutants exhibit very low levels of heteroallelic recombination. kar3 mutants do produce double-strand breaks that act as initiation sites for meiotic recombination in yeast, but at levels severalfold reduced from wild-type.These data are consistent with a meiotic role for Kar3p in the events that culminate in synapsis of homologues.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology and Microbiology, Tufts University School of Medicine, Boston, Massachusetts 02111, USA.

ABSTRACT
The recognition and alignment of homologous chromosomes early in meiosis is essential for their subsequent segregation at anaphase I; however, the mechanism by which this occurs is unknown. We demonstrate here that, in the absence of the molecular motor, Kar3p, meiotic cells are blocked with prophase monopolar microtubule arrays and incomplete synaptonemal complex (SC) formation. kar3 mutants exhibit very low levels of heteroallelic recombination. kar3 mutants do produce double-strand breaks that act as initiation sites for meiotic recombination in yeast, but at levels severalfold reduced from wild-type. These data are consistent with a meiotic role for Kar3p in the events that culminate in synapsis of homologues.

Show MeSH
Related in: MedlinePlus