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A mechanism for nuclear positioning in fission yeast based on microtubule pushing.

Tran PT, Marsh L, Doye V, Inoué S, Chang F - J. Cell Biol. (2001)

Bottom Line: The MT bundles are organized from medial MT-organizing centers that may function as nuclear attachment sites.After an average of 1.5 min of growth at the cell tip, MT plus ends exhibit catastrophe and shrink back to the nuclear region before growing back to the cell tip.Computer modeling suggests that a balance of these pushing MT forces can provide a mechanism to position the nucleus at the middle of the cell.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Columbia University, New York, New York 10032, USA. pt143@columbia.edu

ABSTRACT
The correct positioning of the nucleus is often important in defining the spatial organization of the cell, for example, in determining the cell division plane. In interphase Schizosaccharomyces pombe cells, the nucleus is positioned in the middle of the cylindrical cell in an active microtubule (MT)-dependent process. Here, we used green fluorescent protein markers to examine the dynamics of MTs, spindle pole body, and the nuclear envelope in living cells. We find that interphase MTs are organized in three to four antiparallel MT bundles arranged along the long axis of the cell, with MT plus ends facing both the cell tips and minus ends near the middle of the cell. The MT bundles are organized from medial MT-organizing centers that may function as nuclear attachment sites. When MTs grow to the cell tips, they exert transient forces produced by plus end MT polymerization that push the nucleus. After an average of 1.5 min of growth at the cell tip, MT plus ends exhibit catastrophe and shrink back to the nuclear region before growing back to the cell tip. Computer modeling suggests that a balance of these pushing MT forces can provide a mechanism to position the nucleus at the middle of the cell.

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MT-dependent oscillation of the SPB. Cells expressing cmd1-GFP (PT.1) exhibited labeling of a medial SPB (arrow) and non-SPB patches (possibly actin patches) at the cell tips (Moser et al. 1997). The SPB was clearly distinguished from the patches by its position and dynamics. SPB dynamics in an untreated wild-type cell (A) or in a cell treated with 100 μg ml−1 TBZ for 5 min (B). The top shows an image of the cell with the medial fluorescent SPB; the middle shows tracings of the path of the SPB in a time-lapse sequence of ∼14 min; and the bottom shows plots of SPB displacement from a mean position over time. Bars, 5 μm.
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Figure 5: MT-dependent oscillation of the SPB. Cells expressing cmd1-GFP (PT.1) exhibited labeling of a medial SPB (arrow) and non-SPB patches (possibly actin patches) at the cell tips (Moser et al. 1997). The SPB was clearly distinguished from the patches by its position and dynamics. SPB dynamics in an untreated wild-type cell (A) or in a cell treated with 100 μg ml−1 TBZ for 5 min (B). The top shows an image of the cell with the medial fluorescent SPB; the middle shows tracings of the path of the SPB in a time-lapse sequence of ∼14 min; and the bottom shows plots of SPB displacement from a mean position over time. Bars, 5 μm.

Mentions: To examine the effects of MTs on the nucleus more closely, we measured SPB dynamics. The SPB is associated with the nucleus and one of the MT bundles during interphase (Ding et al. 1997; Hagan and Yanagida 1995). We used GFP-cmd1p as a marker for the SPB (Moser et al. 1997). Time-lapse microscopy revealed that the SPB oscillated largely in a direction parallel to the long axis of the cell (Fig. 5 A), with an average rate of 1.30 ± 0.36 μm min−1 and a maximum excursion length of 2.62 ± 0.57 μm (Table ). To test if this movement was MT dependent, cells were treated for 5 min with 100 μg ml−1 TBZ, an MT-depolymerizing drug. In TBZ-treated cells, SPB movement was greatly diminished with no noticeable directed movement and its maximum excursion length decreased to 0.67 ± 0.15 μm (Fig. 5 B and Table ). These effects were seen in all the TBZ+ cells examined (n = 21). Thus, the SPB oscillates in an MT-dependent manner.


A mechanism for nuclear positioning in fission yeast based on microtubule pushing.

Tran PT, Marsh L, Doye V, Inoué S, Chang F - J. Cell Biol. (2001)

MT-dependent oscillation of the SPB. Cells expressing cmd1-GFP (PT.1) exhibited labeling of a medial SPB (arrow) and non-SPB patches (possibly actin patches) at the cell tips (Moser et al. 1997). The SPB was clearly distinguished from the patches by its position and dynamics. SPB dynamics in an untreated wild-type cell (A) or in a cell treated with 100 μg ml−1 TBZ for 5 min (B). The top shows an image of the cell with the medial fluorescent SPB; the middle shows tracings of the path of the SPB in a time-lapse sequence of ∼14 min; and the bottom shows plots of SPB displacement from a mean position over time. Bars, 5 μm.
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Related In: Results  -  Collection

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Figure 5: MT-dependent oscillation of the SPB. Cells expressing cmd1-GFP (PT.1) exhibited labeling of a medial SPB (arrow) and non-SPB patches (possibly actin patches) at the cell tips (Moser et al. 1997). The SPB was clearly distinguished from the patches by its position and dynamics. SPB dynamics in an untreated wild-type cell (A) or in a cell treated with 100 μg ml−1 TBZ for 5 min (B). The top shows an image of the cell with the medial fluorescent SPB; the middle shows tracings of the path of the SPB in a time-lapse sequence of ∼14 min; and the bottom shows plots of SPB displacement from a mean position over time. Bars, 5 μm.
Mentions: To examine the effects of MTs on the nucleus more closely, we measured SPB dynamics. The SPB is associated with the nucleus and one of the MT bundles during interphase (Ding et al. 1997; Hagan and Yanagida 1995). We used GFP-cmd1p as a marker for the SPB (Moser et al. 1997). Time-lapse microscopy revealed that the SPB oscillated largely in a direction parallel to the long axis of the cell (Fig. 5 A), with an average rate of 1.30 ± 0.36 μm min−1 and a maximum excursion length of 2.62 ± 0.57 μm (Table ). To test if this movement was MT dependent, cells were treated for 5 min with 100 μg ml−1 TBZ, an MT-depolymerizing drug. In TBZ-treated cells, SPB movement was greatly diminished with no noticeable directed movement and its maximum excursion length decreased to 0.67 ± 0.15 μm (Fig. 5 B and Table ). These effects were seen in all the TBZ+ cells examined (n = 21). Thus, the SPB oscillates in an MT-dependent manner.

Bottom Line: The MT bundles are organized from medial MT-organizing centers that may function as nuclear attachment sites.After an average of 1.5 min of growth at the cell tip, MT plus ends exhibit catastrophe and shrink back to the nuclear region before growing back to the cell tip.Computer modeling suggests that a balance of these pushing MT forces can provide a mechanism to position the nucleus at the middle of the cell.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Columbia University, New York, New York 10032, USA. pt143@columbia.edu

ABSTRACT
The correct positioning of the nucleus is often important in defining the spatial organization of the cell, for example, in determining the cell division plane. In interphase Schizosaccharomyces pombe cells, the nucleus is positioned in the middle of the cylindrical cell in an active microtubule (MT)-dependent process. Here, we used green fluorescent protein markers to examine the dynamics of MTs, spindle pole body, and the nuclear envelope in living cells. We find that interphase MTs are organized in three to four antiparallel MT bundles arranged along the long axis of the cell, with MT plus ends facing both the cell tips and minus ends near the middle of the cell. The MT bundles are organized from medial MT-organizing centers that may function as nuclear attachment sites. When MTs grow to the cell tips, they exert transient forces produced by plus end MT polymerization that push the nucleus. After an average of 1.5 min of growth at the cell tip, MT plus ends exhibit catastrophe and shrink back to the nuclear region before growing back to the cell tip. Computer modeling suggests that a balance of these pushing MT forces can provide a mechanism to position the nucleus at the middle of the cell.

Show MeSH