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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 CIPC.(A) Ase1 tagged with GFP was expressed from the native promoter in wild-type cultured in EMM medium containing DMSO or 260 μM CIPC for 5 hours at 30°C. Sad1-mCherry was used as an SPB maker. The bars indicate 5 μm. (B) The Δase1 strain expressing GFP-Atb2 from the native nda3 promoter integrated at the lys1 locus and Sad1-mCherry from the native locus was cultured in EMM medium containing DMSO or 260 μM CIPC for 5 hours at 30°C. The bars indicate 5 μm. (C) Each strain was streaked on YES medium with or without CIPC (260 μM) and incubated at 30°C for 5 days.
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pone.0120109.g008: Relationship between Ase1 and CIPC.(A) Ase1 tagged with GFP was expressed from the native promoter in wild-type cultured in EMM medium containing DMSO or 260 μM CIPC for 5 hours at 30°C. Sad1-mCherry was used as an SPB maker. The bars indicate 5 μm. (B) The Δase1 strain expressing GFP-Atb2 from the native nda3 promoter integrated at the lys1 locus and Sad1-mCherry from the native locus was cultured in EMM medium containing DMSO or 260 μM CIPC for 5 hours at 30°C. The bars indicate 5 μm. (C) Each strain was streaked on YES medium with or without CIPC (260 μM) and incubated at 30°C for 5 days.

Mentions: Because the misorientation of MTs could be caused by a defect in the microtubule organizing center (MTOC), we visually examined certain MTOC components in the Δscp3 strain. As shown in Figs. 7 and 8A, the localization and intensity of the fluorescent signal of the three proteins, Alp4, Mto1 and Ase1, were visibly normal in the Δscp3 strain, further suggesting that MT misorientation in this strain is due to the loss of bundling.


Fission yeast Scp3 potentially maintains microtubule orientation through bundling.

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

Relationship between Ase1 and CIPC.(A) Ase1 tagged with GFP was expressed from the native promoter in wild-type cultured in EMM medium containing DMSO or 260 μM CIPC for 5 hours at 30°C. Sad1-mCherry was used as an SPB maker. The bars indicate 5 μm. (B) The Δase1 strain expressing GFP-Atb2 from the native nda3 promoter integrated at the lys1 locus and Sad1-mCherry from the native locus was cultured in EMM medium containing DMSO or 260 μM CIPC for 5 hours at 30°C. The bars indicate 5 μm. (C) Each strain was streaked on YES medium with or without CIPC (260 μM) and incubated at 30°C for 5 days.
© Copyright Policy
Related In: Results  -  Collection

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

pone.0120109.g008: Relationship between Ase1 and CIPC.(A) Ase1 tagged with GFP was expressed from the native promoter in wild-type cultured in EMM medium containing DMSO or 260 μM CIPC for 5 hours at 30°C. Sad1-mCherry was used as an SPB maker. The bars indicate 5 μm. (B) The Δase1 strain expressing GFP-Atb2 from the native nda3 promoter integrated at the lys1 locus and Sad1-mCherry from the native locus was cultured in EMM medium containing DMSO or 260 μM CIPC for 5 hours at 30°C. The bars indicate 5 μm. (C) Each strain was streaked on YES medium with or without CIPC (260 μM) and incubated at 30°C for 5 days.
Mentions: Because the misorientation of MTs could be caused by a defect in the microtubule organizing center (MTOC), we visually examined certain MTOC components in the Δscp3 strain. As shown in Figs. 7 and 8A, the localization and intensity of the fluorescent signal of the three proteins, Alp4, Mto1 and Ase1, were visibly normal in the Δscp3 strain, further suggesting that MT misorientation in this strain is due to the loss of bundling.

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