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Fission yeast Scp3 potentially maintains microtubule orientation through bundling.

Ozaki K, Chikashige Y, Hiraoka Y, Matsumoto T - PLoS ONE (2015)

Bottom Line: In this study, we investigated the function of Scp3 along with the effect of CIPC in the fission yeast Schizosaccharomyces pombe.Functional analysis suggested that Scp3 functions independently from Ase1, a protein largely required for the bundling of the mitotic spindle.These results suggested that multiple systems are independently involved to ensure microtubule orientation by bundling in fission yeast.

View Article: PubMed Central - PubMed

Affiliation: Graduate School of Biostudies, Kyoto University, Kyoto, Kyoto, Japan.

ABSTRACT
Microtubules play important roles in organelle transport, the maintenance of cell polarity and chromosome segregation and generally form bundles during these processes. The fission yeast gene scp3+ was identified as a multicopy suppressor of the cps3-81 mutant, which is hypersensitive to isopropyl N-3-chlorophenylcarbamate (CIPC), a poison that induces abnormal multipolar spindle formation in higher eukaryotes. In this study, we investigated the function of Scp3 along with the effect of CIPC in the fission yeast Schizosaccharomyces pombe. Microscopic observation revealed that treatment with CIPC, cps3-81 mutation and scp3+ gene deletion disturbed the orientation of microtubules in interphase cells. Overexpression of scp3+ suppressed the abnormal orientation of microtubules by promoting bundling. Functional analysis suggested that Scp3 functions independently from Ase1, a protein largely required for the bundling of the mitotic spindle. A strain lacking the ase1+ gene was more sensitive to CIPC, with the drug affecting the integrity of the mitotic spindle, indicating that CIPC has a mitotic target that has a role redundant with Ase1. These results suggested that multiple systems are independently involved to ensure microtubule orientation by bundling in fission yeast.

No MeSH data available.


Related in: MedlinePlus

Relationship between Ase1 and Scp3.(A) Interphase MTs were observed in the Δase1 strain overexpressing scp3+ or the Δscp3 strain overexpressing ase1+. Each gene was expressed from the nmt promoter of pREP81 in EMM medium at 30°C. (B) Nonparametric Mann-Whitney U test of (A). The red lines are the median. More than 150 microtubules were observed for each condition. (C) Mitotic cells overexpressing ase1+ were observed. ase1+ expression was induced for 18 hours. GFP-Atb2 expressed from the nda3 promoter integrated at the lys1 locus was used as a maker of microtubules and Sad1-mCherry for SPB. (D) Cells with normal spindle (upper) and with multiple SPBs (bottom) were counted for each strain. (E) scp3+ was overexpressed in the Δase1 strain. Aberrant mitotic spindles are marked with arrowheads. The bars indicate 5 μm.
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pone.0120109.g006: Relationship between Ase1 and Scp3.(A) Interphase MTs were observed in the Δase1 strain overexpressing scp3+ or the Δscp3 strain overexpressing ase1+. Each gene was expressed from the nmt promoter of pREP81 in EMM medium at 30°C. (B) Nonparametric Mann-Whitney U test of (A). The red lines are the median. More than 150 microtubules were observed for each condition. (C) Mitotic cells overexpressing ase1+ were observed. ase1+ expression was induced for 18 hours. GFP-Atb2 expressed from the nda3 promoter integrated at the lys1 locus was used as a maker of microtubules and Sad1-mCherry for SPB. (D) Cells with normal spindle (upper) and with multiple SPBs (bottom) were counted for each strain. (E) scp3+ was overexpressed in the Δase1 strain. Aberrant mitotic spindles are marked with arrowheads. The bars indicate 5 μm.

Mentions: Because the phenotype of the Δscp3 strain most resembled that caused by deletion of the ase1+ gene [26,27], we attempted to investigate the functional relationship between Scp3 and Ase1. As shown in Fig. 6A and 6B, overexpression of scp3+ or ase1+ promoted the bundling of MTs in a strain lacking the other gene, suggesting that these two genes promote bundling independently. We also found that ase1+ overexpression was toxic. The mitotic spindle often elongated beyond one of the SPBs in cells overexpression of ase1+, likely due to the abnormal bundling of MTs composing the mitotic spindle (Fig. 6C and 6D). We also noticed that (1) mitotic cells overexpressing ase1+ contained more than 2 SPBs, which could be caused by fragmentation of SPB and (2) this effect was more prominent in cells lacking scp3+ (Fig. 6D), suggesting that Scp3 might be required for maintenance of the integrity of SPB when the spindle dynamics was perturbed.


Fission yeast Scp3 potentially maintains microtubule orientation through bundling.

Ozaki K, Chikashige Y, Hiraoka Y, Matsumoto T - PLoS ONE (2015)

Relationship between Ase1 and Scp3.(A) Interphase MTs were observed in the Δase1 strain overexpressing scp3+ or the Δscp3 strain overexpressing ase1+. Each gene was expressed from the nmt promoter of pREP81 in EMM medium at 30°C. (B) Nonparametric Mann-Whitney U test of (A). The red lines are the median. More than 150 microtubules were observed for each condition. (C) Mitotic cells overexpressing ase1+ were observed. ase1+ expression was induced for 18 hours. GFP-Atb2 expressed from the nda3 promoter integrated at the lys1 locus was used as a maker of microtubules and Sad1-mCherry for SPB. (D) Cells with normal spindle (upper) and with multiple SPBs (bottom) were counted for each strain. (E) scp3+ was overexpressed in the Δase1 strain. Aberrant mitotic spindles are marked with arrowheads. The bars indicate 5 μm.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4359140&req=5

pone.0120109.g006: Relationship between Ase1 and Scp3.(A) Interphase MTs were observed in the Δase1 strain overexpressing scp3+ or the Δscp3 strain overexpressing ase1+. Each gene was expressed from the nmt promoter of pREP81 in EMM medium at 30°C. (B) Nonparametric Mann-Whitney U test of (A). The red lines are the median. More than 150 microtubules were observed for each condition. (C) Mitotic cells overexpressing ase1+ were observed. ase1+ expression was induced for 18 hours. GFP-Atb2 expressed from the nda3 promoter integrated at the lys1 locus was used as a maker of microtubules and Sad1-mCherry for SPB. (D) Cells with normal spindle (upper) and with multiple SPBs (bottom) were counted for each strain. (E) scp3+ was overexpressed in the Δase1 strain. Aberrant mitotic spindles are marked with arrowheads. The bars indicate 5 μm.
Mentions: Because the phenotype of the Δscp3 strain most resembled that caused by deletion of the ase1+ gene [26,27], we attempted to investigate the functional relationship between Scp3 and Ase1. As shown in Fig. 6A and 6B, overexpression of scp3+ or ase1+ promoted the bundling of MTs in a strain lacking the other gene, suggesting that these two genes promote bundling independently. We also found that ase1+ overexpression was toxic. The mitotic spindle often elongated beyond one of the SPBs in cells overexpression of ase1+, likely due to the abnormal bundling of MTs composing the mitotic spindle (Fig. 6C and 6D). We also noticed that (1) mitotic cells overexpressing ase1+ contained more than 2 SPBs, which could be caused by fragmentation of SPB and (2) this effect was more prominent in cells lacking scp3+ (Fig. 6D), suggesting that Scp3 might be required for maintenance of the integrity of SPB when the spindle dynamics was perturbed.

Bottom Line: In this study, we investigated the function of Scp3 along with the effect of CIPC in the fission yeast Schizosaccharomyces pombe.Functional analysis suggested that Scp3 functions independently from Ase1, a protein largely required for the bundling of the mitotic spindle.These results suggested that multiple systems are independently involved to ensure microtubule orientation by bundling in fission yeast.

View Article: PubMed Central - PubMed

Affiliation: Graduate School of Biostudies, Kyoto University, Kyoto, Kyoto, Japan.

ABSTRACT
Microtubules play important roles in organelle transport, the maintenance of cell polarity and chromosome segregation and generally form bundles during these processes. The fission yeast gene scp3+ was identified as a multicopy suppressor of the cps3-81 mutant, which is hypersensitive to isopropyl N-3-chlorophenylcarbamate (CIPC), a poison that induces abnormal multipolar spindle formation in higher eukaryotes. In this study, we investigated the function of Scp3 along with the effect of CIPC in the fission yeast Schizosaccharomyces pombe. Microscopic observation revealed that treatment with CIPC, cps3-81 mutation and scp3+ gene deletion disturbed the orientation of microtubules in interphase cells. Overexpression of scp3+ suppressed the abnormal orientation of microtubules by promoting bundling. Functional analysis suggested that Scp3 functions independently from Ase1, a protein largely required for the bundling of the mitotic spindle. A strain lacking the ase1+ gene was more sensitive to CIPC, with the drug affecting the integrity of the mitotic spindle, indicating that CIPC has a mitotic target that has a role redundant with Ase1. These results suggested that multiple systems are independently involved to ensure microtubule orientation by bundling in fission yeast.

No MeSH data available.


Related in: MedlinePlus