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Dictyostelium IQGAP-related protein specifically involved in the completion of cytokinesis.

Adachi H, Takahashi Y, Hasebe T, Shirouzu M, Yokoyama S, Sutoh K - J. Cell Biol. (1997)

Bottom Line: These results indicate that the GAPA protein is specifically involved in the completion of cytokinesis.Recently, it was reported that IQGAPs are putative effectors for Rac and CDC42, members of the Rho family of GTPases, and participate in reorganization of the actin cytoskeleton.Thus, it is possible that Dictyostelium GAPA participates in the severing of the midbody by regulating the actin cytoskeleton through an interaction with a member of small GTPases.

View Article: PubMed Central - PubMed

Affiliation: Department of Life Sciences, Graduate School of Arts and Sciences, University of Tokyo, 3-8-1 Komaba, Meguro-ku, Tokyo 153. f37771@m-unix.cc.u-tokyo.ac.jp

ABSTRACT
The gapA gene encoding a novel RasGTPase-activating protein (RasGAP)-related protein was found to be disrupted in a cytokinesis mutant of Dictyostelium that grows as giant and multinucleate cells in a dish culture. The predicted sequence of the GAPA protein showed considerable homology to those of Gap1/Sar1 from fission yeast and the COOH-terminal half of mammalian IQGAPs, the similarity extending beyond the RasGAP-related domain. In suspension culture, gapA- cells showed normal growth in terms of the increase in cell mass, but cytokinesis inefficiently occurred to produce spherical giant cells. Time-lapse recording of the dynamics of cell division in a dish culture revealed that, in the case of gapA- cells, cytokinesis was very frequently reversed at the step in which the midbody connecting the daughter cells should be severed. Earlier steps of cytokinesis in the gapA- cells seemed to be normal, since myosin II was accumulated at the cleavage furrow. Upon starvation, gapA- cells developed and formed fruiting bodies with viable spores, like the wild-type cells. These results indicate that the GAPA protein is specifically involved in the completion of cytokinesis. Recently, it was reported that IQGAPs are putative effectors for Rac and CDC42, members of the Rho family of GTPases, and participate in reorganization of the actin cytoskeleton. Thus, it is possible that Dictyostelium GAPA participates in the severing of the midbody by regulating the actin cytoskeleton through an interaction with a member of small GTPases.

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Reversion of cytokinesis  observed in gapA− cells. Wildtype AX2 (upper frames) and mutant D42-2 (lower frames) cells  were axenically grown on coverslips. Cell division was observed  under a microscope connected to  a time-lapse video recorder. The  number in each frame indicates  the time in min. In the lower  frames (D42-2), the mutant cells  (open and filled triangles) attempted to divide into two daughter cells. For the cell with the  filled triangle, cytokinesis was reversed at the step where the two  daughter cells were connected by  a thin cytoplasmic bridge (midbody; arrowhead). In contrast, the  cell with the open triangle completed cytokinesis like wild-type  cells. Bar, 10 μm.
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Figure 8: Reversion of cytokinesis observed in gapA− cells. Wildtype AX2 (upper frames) and mutant D42-2 (lower frames) cells were axenically grown on coverslips. Cell division was observed under a microscope connected to a time-lapse video recorder. The number in each frame indicates the time in min. In the lower frames (D42-2), the mutant cells (open and filled triangles) attempted to divide into two daughter cells. For the cell with the filled triangle, cytokinesis was reversed at the step where the two daughter cells were connected by a thin cytoplasmic bridge (midbody; arrowhead). In contrast, the cell with the open triangle completed cytokinesis like wild-type cells. Bar, 10 μm.

Mentions: Second, the dynamics of cell division on a glass surface were monitored by means of time-lapse video recording. gapA− cells (Fig. 8, lower frames) stopped protruding pseudopodia, became rounded, and then became constricted to produce daughter cells linked by a thin cytoplasmic bridge (midbody; Fig. 8, arrowheads). By this stage, cytokinesis of the gapA− cells was similar to that of the wild-type cells (Fig. 8, upper frames), although it sometimes took longer for constriction in the mutant. This bridge, however, frequently remained intact for a long time, and finally cytokinesis was reversed to yield a single cell (Fig. 8, filled triangle). This reversion occurred for ∼50% (n = 44) of the smallest mutant cells that attempted to divide into two. The failed cytokinesis accumulated to produce giant and multinucleate cells. These results suggest that, in gapA− cells, earlier stages of cytokinesis are rarely affected, whereas the fidelity of the latest severing of the midbody is greatly reduced. Consistent with this notion, myosin II was localized at the cleavage furrow in the dividing mutant cells as in the dividing wild-type cells (Fig. 9). In interphase mutant cells, myosin II was also predominantly localized at the periphery of the cells, as in the wild-type cells. The localization of actin filaments in interphase mutant cells was also indistinguishable from that in the wildtype cells (data not shown).


Dictyostelium IQGAP-related protein specifically involved in the completion of cytokinesis.

Adachi H, Takahashi Y, Hasebe T, Shirouzu M, Yokoyama S, Sutoh K - J. Cell Biol. (1997)

Reversion of cytokinesis  observed in gapA− cells. Wildtype AX2 (upper frames) and mutant D42-2 (lower frames) cells  were axenically grown on coverslips. Cell division was observed  under a microscope connected to  a time-lapse video recorder. The  number in each frame indicates  the time in min. In the lower  frames (D42-2), the mutant cells  (open and filled triangles) attempted to divide into two daughter cells. For the cell with the  filled triangle, cytokinesis was reversed at the step where the two  daughter cells were connected by  a thin cytoplasmic bridge (midbody; arrowhead). In contrast, the  cell with the open triangle completed cytokinesis like wild-type  cells. Bar, 10 μm.
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Related In: Results  -  Collection

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Figure 8: Reversion of cytokinesis observed in gapA− cells. Wildtype AX2 (upper frames) and mutant D42-2 (lower frames) cells were axenically grown on coverslips. Cell division was observed under a microscope connected to a time-lapse video recorder. The number in each frame indicates the time in min. In the lower frames (D42-2), the mutant cells (open and filled triangles) attempted to divide into two daughter cells. For the cell with the filled triangle, cytokinesis was reversed at the step where the two daughter cells were connected by a thin cytoplasmic bridge (midbody; arrowhead). In contrast, the cell with the open triangle completed cytokinesis like wild-type cells. Bar, 10 μm.
Mentions: Second, the dynamics of cell division on a glass surface were monitored by means of time-lapse video recording. gapA− cells (Fig. 8, lower frames) stopped protruding pseudopodia, became rounded, and then became constricted to produce daughter cells linked by a thin cytoplasmic bridge (midbody; Fig. 8, arrowheads). By this stage, cytokinesis of the gapA− cells was similar to that of the wild-type cells (Fig. 8, upper frames), although it sometimes took longer for constriction in the mutant. This bridge, however, frequently remained intact for a long time, and finally cytokinesis was reversed to yield a single cell (Fig. 8, filled triangle). This reversion occurred for ∼50% (n = 44) of the smallest mutant cells that attempted to divide into two. The failed cytokinesis accumulated to produce giant and multinucleate cells. These results suggest that, in gapA− cells, earlier stages of cytokinesis are rarely affected, whereas the fidelity of the latest severing of the midbody is greatly reduced. Consistent with this notion, myosin II was localized at the cleavage furrow in the dividing mutant cells as in the dividing wild-type cells (Fig. 9). In interphase mutant cells, myosin II was also predominantly localized at the periphery of the cells, as in the wild-type cells. The localization of actin filaments in interphase mutant cells was also indistinguishable from that in the wildtype cells (data not shown).

Bottom Line: These results indicate that the GAPA protein is specifically involved in the completion of cytokinesis.Recently, it was reported that IQGAPs are putative effectors for Rac and CDC42, members of the Rho family of GTPases, and participate in reorganization of the actin cytoskeleton.Thus, it is possible that Dictyostelium GAPA participates in the severing of the midbody by regulating the actin cytoskeleton through an interaction with a member of small GTPases.

View Article: PubMed Central - PubMed

Affiliation: Department of Life Sciences, Graduate School of Arts and Sciences, University of Tokyo, 3-8-1 Komaba, Meguro-ku, Tokyo 153. f37771@m-unix.cc.u-tokyo.ac.jp

ABSTRACT
The gapA gene encoding a novel RasGTPase-activating protein (RasGAP)-related protein was found to be disrupted in a cytokinesis mutant of Dictyostelium that grows as giant and multinucleate cells in a dish culture. The predicted sequence of the GAPA protein showed considerable homology to those of Gap1/Sar1 from fission yeast and the COOH-terminal half of mammalian IQGAPs, the similarity extending beyond the RasGAP-related domain. In suspension culture, gapA- cells showed normal growth in terms of the increase in cell mass, but cytokinesis inefficiently occurred to produce spherical giant cells. Time-lapse recording of the dynamics of cell division in a dish culture revealed that, in the case of gapA- cells, cytokinesis was very frequently reversed at the step in which the midbody connecting the daughter cells should be severed. Earlier steps of cytokinesis in the gapA- cells seemed to be normal, since myosin II was accumulated at the cleavage furrow. Upon starvation, gapA- cells developed and formed fruiting bodies with viable spores, like the wild-type cells. These results indicate that the GAPA protein is specifically involved in the completion of cytokinesis. Recently, it was reported that IQGAPs are putative effectors for Rac and CDC42, members of the Rho family of GTPases, and participate in reorganization of the actin cytoskeleton. Thus, it is possible that Dictyostelium GAPA participates in the severing of the midbody by regulating the actin cytoskeleton through an interaction with a member of small GTPases.

Show MeSH
Related in: MedlinePlus