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Cortical Num1p interacts with the dynein intermediate chain Pac11p and cytoplasmic microtubules in budding yeast.

Farkasovsky M, Küntzel H - J. Cell Biol. (2001)

Bottom Line: The bud tip location of Num1p depends on Bni1p but does not require Kar9p, Dyn1p, or cMTs, whereas cMT contacts with polar Num1p dots are reduced in cells lacking Dyn1p.Num1p also forms a complex with Bni1p and Kar9p, although Num1p is not required for Bni1p- and Kar9p-dependent nuclear migration to the bud neck in preanaphase cells.In addition, Num1p may transiently cooperate with an associated Bni1p-Kar9p complex at the bud tip of early anaphase cells.

View Article: PubMed Central - PubMed

Affiliation: Max-Planck Institute for Experimental Medicine, D-37075 Göttingen, Germany.

ABSTRACT
Num1p, a cortical 313-kD protein, controls cytoplasmic microtubule (cMT) functions and nuclear migration through the bud neck in anaphase cells. A green fluorescent protein (GFP)-Num1p fusion protein localizes at the bud tip and the distal mother pole of living cells, apparently forming cMT capture sites at late anaphase. In addition, galactose-induced GFP-Num1p is seen at the bud neck and in lateral regions of the mother cortex. The bud tip location of Num1p depends on Bni1p but does not require Kar9p, Dyn1p, or cMTs, whereas cMT contacts with polar Num1p dots are reduced in cells lacking Dyn1p. Num1p associates with the dynein intermediate chain Pac11p in the presence of Dyn1p, and with the alpha-tubulin Tub3p, as shown by coimmune precipitation of tagged proteins. Num1p also forms a complex with Bni1p and Kar9p, although Num1p is not required for Bni1p- and Kar9p-dependent nuclear migration to the bud neck in preanaphase cells. Our data suggest that Num1p controls nuclear migration during late anaphase by forming dynein-interacting cortical cMT capture sites at both cellular poles. In addition, Num1p may transiently cooperate with an associated Bni1p-Kar9p complex at the bud tip of early anaphase cells.

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Polymyxin B sensitivity of num1Δ and yEGFP3-NUM1 strains. Suspensions containing 10, 100, or 1,000 cells (from left to right) of haploid strains FMY691 (a, NUM1), FMY627 (b, yEGFP3-NUM1) and FMY693 (c, num1Δ) were dotted onto YPGal solid media without (left) or with (right) 0.3 mg/ml polymyxin B.
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Figure 1: Polymyxin B sensitivity of num1Δ and yEGFP3-NUM1 strains. Suspensions containing 10, 100, or 1,000 cells (from left to right) of haploid strains FMY691 (a, NUM1), FMY627 (b, yEGFP3-NUM1) and FMY693 (c, num1Δ) were dotted onto YPGal solid media without (left) or with (right) 0.3 mg/ml polymyxin B.

Mentions: Fig. 1 indicates that the NUM1 wild-type strain FMY691 grows well on YPGal containing 0.3 mg polymyxin B (row a), whereas the num1Δ strain FMY693 does not grow under these conditions (row c). Strain FMY627 expressing a galactose-inducible yEGFP-NUM1 fusion gene grows like wild type (Fig. 1, row b), suggesting that the fusion protein is functional like Num1p in conferring polymyxin B resistance. Furthermore, a centromeric plasmid expressing yEGFP-NUM1 under the control of the NUM1 promoter (pFM313) rescues the nuclear migration defect of num1Δ strains (data not shown).


Cortical Num1p interacts with the dynein intermediate chain Pac11p and cytoplasmic microtubules in budding yeast.

Farkasovsky M, Küntzel H - J. Cell Biol. (2001)

Polymyxin B sensitivity of num1Δ and yEGFP3-NUM1 strains. Suspensions containing 10, 100, or 1,000 cells (from left to right) of haploid strains FMY691 (a, NUM1), FMY627 (b, yEGFP3-NUM1) and FMY693 (c, num1Δ) were dotted onto YPGal solid media without (left) or with (right) 0.3 mg/ml polymyxin B.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2199608&req=5

Figure 1: Polymyxin B sensitivity of num1Δ and yEGFP3-NUM1 strains. Suspensions containing 10, 100, or 1,000 cells (from left to right) of haploid strains FMY691 (a, NUM1), FMY627 (b, yEGFP3-NUM1) and FMY693 (c, num1Δ) were dotted onto YPGal solid media without (left) or with (right) 0.3 mg/ml polymyxin B.
Mentions: Fig. 1 indicates that the NUM1 wild-type strain FMY691 grows well on YPGal containing 0.3 mg polymyxin B (row a), whereas the num1Δ strain FMY693 does not grow under these conditions (row c). Strain FMY627 expressing a galactose-inducible yEGFP-NUM1 fusion gene grows like wild type (Fig. 1, row b), suggesting that the fusion protein is functional like Num1p in conferring polymyxin B resistance. Furthermore, a centromeric plasmid expressing yEGFP-NUM1 under the control of the NUM1 promoter (pFM313) rescues the nuclear migration defect of num1Δ strains (data not shown).

Bottom Line: The bud tip location of Num1p depends on Bni1p but does not require Kar9p, Dyn1p, or cMTs, whereas cMT contacts with polar Num1p dots are reduced in cells lacking Dyn1p.Num1p also forms a complex with Bni1p and Kar9p, although Num1p is not required for Bni1p- and Kar9p-dependent nuclear migration to the bud neck in preanaphase cells.In addition, Num1p may transiently cooperate with an associated Bni1p-Kar9p complex at the bud tip of early anaphase cells.

View Article: PubMed Central - PubMed

Affiliation: Max-Planck Institute for Experimental Medicine, D-37075 Göttingen, Germany.

ABSTRACT
Num1p, a cortical 313-kD protein, controls cytoplasmic microtubule (cMT) functions and nuclear migration through the bud neck in anaphase cells. A green fluorescent protein (GFP)-Num1p fusion protein localizes at the bud tip and the distal mother pole of living cells, apparently forming cMT capture sites at late anaphase. In addition, galactose-induced GFP-Num1p is seen at the bud neck and in lateral regions of the mother cortex. The bud tip location of Num1p depends on Bni1p but does not require Kar9p, Dyn1p, or cMTs, whereas cMT contacts with polar Num1p dots are reduced in cells lacking Dyn1p. Num1p associates with the dynein intermediate chain Pac11p in the presence of Dyn1p, and with the alpha-tubulin Tub3p, as shown by coimmune precipitation of tagged proteins. Num1p also forms a complex with Bni1p and Kar9p, although Num1p is not required for Bni1p- and Kar9p-dependent nuclear migration to the bud neck in preanaphase cells. Our data suggest that Num1p controls nuclear migration during late anaphase by forming dynein-interacting cortical cMT capture sites at both cellular poles. In addition, Num1p may transiently cooperate with an associated Bni1p-Kar9p complex at the bud tip of early anaphase cells.

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