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Dynamics of multiple nuclei in Ashbya gossypii hyphae depend on the control of cytoplasmic microtubules length by Bik1, Kip2, Kip3, and not on a capture/shrinkage mechanism.

Grava S, Philippsen P - Mol. Biol. Cell (2010)

Bottom Line: Growing cMTs slide along the hyphal cortex and exert pulling forces on nuclei.Surprisingly, a capture/shrinkage mechanism seems to be absent in A. gossypii. cMTs reaching a hyphal tip do not shrink, and cMT +ends accumulate in hyphal tips.Thus, differences in cMT dynamics and length control between budding yeast and A. gossypii are key elements in the adaptation of the cMT cytoskeleton to much longer cells and much higher degrees of nuclear mobilities.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Microbiology, Biozentrum, University of Basel, CH-4056 Basel, Switzerland.

ABSTRACT
Ashbya gossypii has a budding yeast-like genome but grows exclusively as multinucleated hyphae. In contrast to budding yeast where positioning of nuclei at the bud neck is a major function of cytoplasmic microtubules (cMTs), A. gossypii nuclei are constantly in motion and positioning is not an issue. To investigate the role of cMTs in nuclear oscillation and bypassing, we constructed mutants potentially affecting cMT lengths. Hyphae lacking the plus (+)end marker Bik1 or the kinesin Kip2 cannot polymerize long cMTs and lose wild-type nuclear movements. Interestingly, hyphae lacking the kinesin Kip3 display longer cMTs concomitant with increased nuclear oscillation and bypassing. Polymerization and depolymerization rates of cMTs are 3 times higher in A. gossypii than in budding yeast and cMT catastrophes are rare. Growing cMTs slide along the hyphal cortex and exert pulling forces on nuclei. Surprisingly, a capture/shrinkage mechanism seems to be absent in A. gossypii. cMTs reaching a hyphal tip do not shrink, and cMT +ends accumulate in hyphal tips. Thus, differences in cMT dynamics and length control between budding yeast and A. gossypii are key elements in the adaptation of the cMT cytoskeleton to much longer cells and much higher degrees of nuclear mobilities.

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cMT interactions with the cell cortex generate SPB movements in A. gossypii cells. (A) Examples of 1Z plane time-lapse movies with GFP-Tub1 cells. a, cMT grows and sweeps the cortex. The SPB does not significantly move laterally but is pushed longitudinally. Yellow arrows represent the position of the SPB at t0. b, cMT grows and pushes the SPB against the cortex. c, cMT grows and slides along the cortex resulting in a longitudinal SPB movement. a–c, dashed lines represent the uppermost positions of SPBs in the image sequences. The cMT +ends are marked with white arrows. (B) A cMT grows and then slides along the cortex in a GFP-Tub1 Bik1-Cherry hypha (sliding occurs at t36–t54). Asterisk (*) shows a Bik1 dot moving along the cMT. Numbers indicate time in seconds. Bars, 5 μm.
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Figure 3: cMT interactions with the cell cortex generate SPB movements in A. gossypii cells. (A) Examples of 1Z plane time-lapse movies with GFP-Tub1 cells. a, cMT grows and sweeps the cortex. The SPB does not significantly move laterally but is pushed longitudinally. Yellow arrows represent the position of the SPB at t0. b, cMT grows and pushes the SPB against the cortex. c, cMT grows and slides along the cortex resulting in a longitudinal SPB movement. a–c, dashed lines represent the uppermost positions of SPBs in the image sequences. The cMT +ends are marked with white arrows. (B) A cMT grows and then slides along the cortex in a GFP-Tub1 Bik1-Cherry hypha (sliding occurs at t36–t54). Asterisk (*) shows a Bik1 dot moving along the cMT. Numbers indicate time in seconds. Bars, 5 μm.

Mentions: Interestingly, in addition to the bright spot visible at the cMT +tip, lower intensity Bik1 spots were visible along the cMT (Figures 1B and 3A). We hypothesized that those additional Bik1 spots can either be brought to the cMT +tip by a +end directed kinesin motor or moved toward the −end via a −end-directed motor, or alternatively be located at the +tip of a short cMT bundled together with a longer cMT. Figure 2C shows that two Bik1 spots located at the +tip of two cMTs emanating from different SPBs can eventually fuse (fluorescence intensity at t30: spot S1 = 10.8, spot S2 = 7.8; fluorescence intensity at t36: spot S = 17.6). This suggests that a +tip dot of Bik1 could attach to another cMT and move along it to finally form parallel cMT bundles. Bundling of cMT from one SPB or different SPBs may be a way to increase cMT stability. Altogether, the very low frequency of cMT catastrophes, the higher cMT polymerization rate, and the possibility that cMTs form parallel bundles can at least partially explain why cMT grow longer in A. gossypii than in budding yeast.


Dynamics of multiple nuclei in Ashbya gossypii hyphae depend on the control of cytoplasmic microtubules length by Bik1, Kip2, Kip3, and not on a capture/shrinkage mechanism.

Grava S, Philippsen P - Mol. Biol. Cell (2010)

cMT interactions with the cell cortex generate SPB movements in A. gossypii cells. (A) Examples of 1Z plane time-lapse movies with GFP-Tub1 cells. a, cMT grows and sweeps the cortex. The SPB does not significantly move laterally but is pushed longitudinally. Yellow arrows represent the position of the SPB at t0. b, cMT grows and pushes the SPB against the cortex. c, cMT grows and slides along the cortex resulting in a longitudinal SPB movement. a–c, dashed lines represent the uppermost positions of SPBs in the image sequences. The cMT +ends are marked with white arrows. (B) A cMT grows and then slides along the cortex in a GFP-Tub1 Bik1-Cherry hypha (sliding occurs at t36–t54). Asterisk (*) shows a Bik1 dot moving along the cMT. Numbers indicate time in seconds. Bars, 5 μm.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2965685&req=5

Figure 3: cMT interactions with the cell cortex generate SPB movements in A. gossypii cells. (A) Examples of 1Z plane time-lapse movies with GFP-Tub1 cells. a, cMT grows and sweeps the cortex. The SPB does not significantly move laterally but is pushed longitudinally. Yellow arrows represent the position of the SPB at t0. b, cMT grows and pushes the SPB against the cortex. c, cMT grows and slides along the cortex resulting in a longitudinal SPB movement. a–c, dashed lines represent the uppermost positions of SPBs in the image sequences. The cMT +ends are marked with white arrows. (B) A cMT grows and then slides along the cortex in a GFP-Tub1 Bik1-Cherry hypha (sliding occurs at t36–t54). Asterisk (*) shows a Bik1 dot moving along the cMT. Numbers indicate time in seconds. Bars, 5 μm.
Mentions: Interestingly, in addition to the bright spot visible at the cMT +tip, lower intensity Bik1 spots were visible along the cMT (Figures 1B and 3A). We hypothesized that those additional Bik1 spots can either be brought to the cMT +tip by a +end directed kinesin motor or moved toward the −end via a −end-directed motor, or alternatively be located at the +tip of a short cMT bundled together with a longer cMT. Figure 2C shows that two Bik1 spots located at the +tip of two cMTs emanating from different SPBs can eventually fuse (fluorescence intensity at t30: spot S1 = 10.8, spot S2 = 7.8; fluorescence intensity at t36: spot S = 17.6). This suggests that a +tip dot of Bik1 could attach to another cMT and move along it to finally form parallel cMT bundles. Bundling of cMT from one SPB or different SPBs may be a way to increase cMT stability. Altogether, the very low frequency of cMT catastrophes, the higher cMT polymerization rate, and the possibility that cMTs form parallel bundles can at least partially explain why cMT grow longer in A. gossypii than in budding yeast.

Bottom Line: Growing cMTs slide along the hyphal cortex and exert pulling forces on nuclei.Surprisingly, a capture/shrinkage mechanism seems to be absent in A. gossypii. cMTs reaching a hyphal tip do not shrink, and cMT +ends accumulate in hyphal tips.Thus, differences in cMT dynamics and length control between budding yeast and A. gossypii are key elements in the adaptation of the cMT cytoskeleton to much longer cells and much higher degrees of nuclear mobilities.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Microbiology, Biozentrum, University of Basel, CH-4056 Basel, Switzerland.

ABSTRACT
Ashbya gossypii has a budding yeast-like genome but grows exclusively as multinucleated hyphae. In contrast to budding yeast where positioning of nuclei at the bud neck is a major function of cytoplasmic microtubules (cMTs), A. gossypii nuclei are constantly in motion and positioning is not an issue. To investigate the role of cMTs in nuclear oscillation and bypassing, we constructed mutants potentially affecting cMT lengths. Hyphae lacking the plus (+)end marker Bik1 or the kinesin Kip2 cannot polymerize long cMTs and lose wild-type nuclear movements. Interestingly, hyphae lacking the kinesin Kip3 display longer cMTs concomitant with increased nuclear oscillation and bypassing. Polymerization and depolymerization rates of cMTs are 3 times higher in A. gossypii than in budding yeast and cMT catastrophes are rare. Growing cMTs slide along the hyphal cortex and exert pulling forces on nuclei. Surprisingly, a capture/shrinkage mechanism seems to be absent in A. gossypii. cMTs reaching a hyphal tip do not shrink, and cMT +ends accumulate in hyphal tips. Thus, differences in cMT dynamics and length control between budding yeast and A. gossypii are key elements in the adaptation of the cMT cytoskeleton to much longer cells and much higher degrees of nuclear mobilities.

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