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Asymmetrically dividing Drosophila neuroblasts utilize two spatially and temporally independent cytokinesis pathways.

Roth M, Roubinet C, Iffländer N, Ferrand A, Cabernard C - Nat Commun (2015)

Bottom Line: In most metazoan cells, contractile ring placement is regulated by the mitotic spindle through the centralspindlin complex, and potentially also the chromosomal passenger complex (CPC).Drosophila neuroblasts, asymmetrically dividing neural stem cells, but also other cells utilize both spindle-dependent and spindle-independent cleavage furrow positioning pathways.However, the relative contribution of each pathway towards cytokinesis is currently unclear.

View Article: PubMed Central - PubMed

Affiliation: Biozentrum, University of Basel, Klingelbergstrasse 50-70, CH-4056 Basel, Switzerland.

ABSTRACT
Precise cleavage furrow positioning is required for faithful chromosome segregation and cell fate determinant distribution. In most metazoan cells, contractile ring placement is regulated by the mitotic spindle through the centralspindlin complex, and potentially also the chromosomal passenger complex (CPC). Drosophila neuroblasts, asymmetrically dividing neural stem cells, but also other cells utilize both spindle-dependent and spindle-independent cleavage furrow positioning pathways. However, the relative contribution of each pathway towards cytokinesis is currently unclear. Here we report that in Drosophila neuroblasts, the mitotic spindle, but not polarity cues, controls the localization of the CPC component Survivin. We also show that Survivin and the mitotic spindle are required to stabilize the position of the cleavage furrow in late anaphase and to complete furrow constriction. These results support the model that two spatially and temporally separate pathways control different key aspects during asymmetric cell division, ensuring correct cell fate determinant segregation and neuroblast self-renewal.

No MeSH data available.


Related in: MedlinePlus

The spindle-dependent pathway stabilizes furrow positioning.Myosin intensity was measured along the neuroblast cortex (green gradient line) and averaged for wild-type, scpoz2775/Df(3R)5780 and dlgm52;;pinsP89 mutant neuroblasts at (a) metaphase, (b) anaphase and (c) telophase. Vertical dashed lines represent the forming cleavage furrow. Horizontal dashed lines indicate the difference between the lowest and the highest intensity values. These intensity differences are plotted in (d) for the indicated genotypes. Average values were derived from at least five neuroblasts. Error bars indicate s.d. (e) Cleavage furrow positioning was independently measured at the onset of furrowing for wild type (blue ball), dlg;;pins (green ball), scpo (orange ball) and rod and colcemid (brown ball). The A/B ratio (A, distance from the furrow to the apical cortex; B, distance from the furrow to the basal cortex) was plotted as a ratio in (f). Asterisk (*) denotes statistical significance. P=3.4 × 10−9 (two-sample equal variance t-test; wt vs dlg;;pins), P=0.00054 (two-sample equal variance t-test; wt vs scpo), P=0.00094 (two-sample unequal variance t-test; wt vs rod and colcemid). NS, not significant; P>0.01 (based on two-sample equal or unequal variance t-test). Dashed orange line outlines the cell boundaries. Dashed white line highlights the position of the cleavage furrow. Scale bar, 5 μm. wt, wild type.
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f7: The spindle-dependent pathway stabilizes furrow positioning.Myosin intensity was measured along the neuroblast cortex (green gradient line) and averaged for wild-type, scpoz2775/Df(3R)5780 and dlgm52;;pinsP89 mutant neuroblasts at (a) metaphase, (b) anaphase and (c) telophase. Vertical dashed lines represent the forming cleavage furrow. Horizontal dashed lines indicate the difference between the lowest and the highest intensity values. These intensity differences are plotted in (d) for the indicated genotypes. Average values were derived from at least five neuroblasts. Error bars indicate s.d. (e) Cleavage furrow positioning was independently measured at the onset of furrowing for wild type (blue ball), dlg;;pins (green ball), scpo (orange ball) and rod and colcemid (brown ball). The A/B ratio (A, distance from the furrow to the apical cortex; B, distance from the furrow to the basal cortex) was plotted as a ratio in (f). Asterisk (*) denotes statistical significance. P=3.4 × 10−9 (two-sample equal variance t-test; wt vs dlg;;pins), P=0.00054 (two-sample equal variance t-test; wt vs scpo), P=0.00094 (two-sample unequal variance t-test; wt vs rod and colcemid). NS, not significant; P>0.01 (based on two-sample equal or unequal variance t-test). Dashed orange line outlines the cell boundaries. Dashed white line highlights the position of the cleavage furrow. Scale bar, 5 μm. wt, wild type.

Mentions: Finally, we wanted to test whether Survivin affects Myosin dynamics and cleavage furrow positioning. To this end, we measured Myosin intensity along the neuroblast cortex (from apical to basal) and established intensity profiles (see Methods), allowing us to compare Myosin dynamics between wild-type and scpo mutant neuroblasts. Wild-type and scpo mutant neuroblasts showed comparable Myosin intensity and distribution during metaphase (Figs 5a,d and 7a). During anaphase, wild-type neuroblasts cleared Myosin from the apical cortex, enriching it in the furrow region (Figs 5a and 7b). scpo mutant neuroblasts also cleared Myosin apically but showed less precise Myosin accumulation in the furrow region by anaphase. Furthermore, basal Myosin clearing was delayed in scpo mutant neuroblasts (Figs 5d and 7b). Nevertheless, by telophase, scpo mutant neuroblasts also showed Myosin enrichment in the furrow region, comparable to wild type (Figs 5d and 7c). Similar Myosin dynamics were also observed for aurB RNAi-treated neuroblasts (Fig. 6g and data not shown). We quantified the intensity difference between the lowest and highest intensities at metaphase, anaphase and telophase, but failed to detect a significant difference between wild type and scpo mutants (Fig. 7d). Interestingly, scpo intensity profiles revealed that the furrow region is shifted towards the basal cortex (Fig. 7c). We performed the same analysis on dlg;;pins double mutants and found distinct intensity profiles: Myosin cleared at both poles at the same time, accumulating Myosin in the middle of the cell (Fig. 7b,c). However, intensity differences between the lowest and highest intensities at metaphase, anaphase and telophase were not significantly different in dlg;;pins double mutants compared with wild type (Fig. 7d).


Asymmetrically dividing Drosophila neuroblasts utilize two spatially and temporally independent cytokinesis pathways.

Roth M, Roubinet C, Iffländer N, Ferrand A, Cabernard C - Nat Commun (2015)

The spindle-dependent pathway stabilizes furrow positioning.Myosin intensity was measured along the neuroblast cortex (green gradient line) and averaged for wild-type, scpoz2775/Df(3R)5780 and dlgm52;;pinsP89 mutant neuroblasts at (a) metaphase, (b) anaphase and (c) telophase. Vertical dashed lines represent the forming cleavage furrow. Horizontal dashed lines indicate the difference between the lowest and the highest intensity values. These intensity differences are plotted in (d) for the indicated genotypes. Average values were derived from at least five neuroblasts. Error bars indicate s.d. (e) Cleavage furrow positioning was independently measured at the onset of furrowing for wild type (blue ball), dlg;;pins (green ball), scpo (orange ball) and rod and colcemid (brown ball). The A/B ratio (A, distance from the furrow to the apical cortex; B, distance from the furrow to the basal cortex) was plotted as a ratio in (f). Asterisk (*) denotes statistical significance. P=3.4 × 10−9 (two-sample equal variance t-test; wt vs dlg;;pins), P=0.00054 (two-sample equal variance t-test; wt vs scpo), P=0.00094 (two-sample unequal variance t-test; wt vs rod and colcemid). NS, not significant; P>0.01 (based on two-sample equal or unequal variance t-test). Dashed orange line outlines the cell boundaries. Dashed white line highlights the position of the cleavage furrow. Scale bar, 5 μm. wt, wild type.
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Related In: Results  -  Collection

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f7: The spindle-dependent pathway stabilizes furrow positioning.Myosin intensity was measured along the neuroblast cortex (green gradient line) and averaged for wild-type, scpoz2775/Df(3R)5780 and dlgm52;;pinsP89 mutant neuroblasts at (a) metaphase, (b) anaphase and (c) telophase. Vertical dashed lines represent the forming cleavage furrow. Horizontal dashed lines indicate the difference between the lowest and the highest intensity values. These intensity differences are plotted in (d) for the indicated genotypes. Average values were derived from at least five neuroblasts. Error bars indicate s.d. (e) Cleavage furrow positioning was independently measured at the onset of furrowing for wild type (blue ball), dlg;;pins (green ball), scpo (orange ball) and rod and colcemid (brown ball). The A/B ratio (A, distance from the furrow to the apical cortex; B, distance from the furrow to the basal cortex) was plotted as a ratio in (f). Asterisk (*) denotes statistical significance. P=3.4 × 10−9 (two-sample equal variance t-test; wt vs dlg;;pins), P=0.00054 (two-sample equal variance t-test; wt vs scpo), P=0.00094 (two-sample unequal variance t-test; wt vs rod and colcemid). NS, not significant; P>0.01 (based on two-sample equal or unequal variance t-test). Dashed orange line outlines the cell boundaries. Dashed white line highlights the position of the cleavage furrow. Scale bar, 5 μm. wt, wild type.
Mentions: Finally, we wanted to test whether Survivin affects Myosin dynamics and cleavage furrow positioning. To this end, we measured Myosin intensity along the neuroblast cortex (from apical to basal) and established intensity profiles (see Methods), allowing us to compare Myosin dynamics between wild-type and scpo mutant neuroblasts. Wild-type and scpo mutant neuroblasts showed comparable Myosin intensity and distribution during metaphase (Figs 5a,d and 7a). During anaphase, wild-type neuroblasts cleared Myosin from the apical cortex, enriching it in the furrow region (Figs 5a and 7b). scpo mutant neuroblasts also cleared Myosin apically but showed less precise Myosin accumulation in the furrow region by anaphase. Furthermore, basal Myosin clearing was delayed in scpo mutant neuroblasts (Figs 5d and 7b). Nevertheless, by telophase, scpo mutant neuroblasts also showed Myosin enrichment in the furrow region, comparable to wild type (Figs 5d and 7c). Similar Myosin dynamics were also observed for aurB RNAi-treated neuroblasts (Fig. 6g and data not shown). We quantified the intensity difference between the lowest and highest intensities at metaphase, anaphase and telophase, but failed to detect a significant difference between wild type and scpo mutants (Fig. 7d). Interestingly, scpo intensity profiles revealed that the furrow region is shifted towards the basal cortex (Fig. 7c). We performed the same analysis on dlg;;pins double mutants and found distinct intensity profiles: Myosin cleared at both poles at the same time, accumulating Myosin in the middle of the cell (Fig. 7b,c). However, intensity differences between the lowest and highest intensities at metaphase, anaphase and telophase were not significantly different in dlg;;pins double mutants compared with wild type (Fig. 7d).

Bottom Line: In most metazoan cells, contractile ring placement is regulated by the mitotic spindle through the centralspindlin complex, and potentially also the chromosomal passenger complex (CPC).Drosophila neuroblasts, asymmetrically dividing neural stem cells, but also other cells utilize both spindle-dependent and spindle-independent cleavage furrow positioning pathways.However, the relative contribution of each pathway towards cytokinesis is currently unclear.

View Article: PubMed Central - PubMed

Affiliation: Biozentrum, University of Basel, Klingelbergstrasse 50-70, CH-4056 Basel, Switzerland.

ABSTRACT
Precise cleavage furrow positioning is required for faithful chromosome segregation and cell fate determinant distribution. In most metazoan cells, contractile ring placement is regulated by the mitotic spindle through the centralspindlin complex, and potentially also the chromosomal passenger complex (CPC). Drosophila neuroblasts, asymmetrically dividing neural stem cells, but also other cells utilize both spindle-dependent and spindle-independent cleavage furrow positioning pathways. However, the relative contribution of each pathway towards cytokinesis is currently unclear. Here we report that in Drosophila neuroblasts, the mitotic spindle, but not polarity cues, controls the localization of the CPC component Survivin. We also show that Survivin and the mitotic spindle are required to stabilize the position of the cleavage furrow in late anaphase and to complete furrow constriction. These results support the model that two spatially and temporally separate pathways control different key aspects during asymmetric cell division, ensuring correct cell fate determinant segregation and neuroblast self-renewal.

No MeSH data available.


Related in: MedlinePlus