A Role for Dictyostelium RacE in Cortical Tension and Cleavage Furrow Progression

Abstract

The small GTPase racE is essential for cytokinesis in Dictyostelium. We found that this requirement is restricted to cells grown in suspension. When attached to a substrate, racE null cells form an actomyosin contractile ring and complete cytokinesis normally. Nonetheless, racE null cells fail completely in cytokinesis when in suspension. To understand this conditional requirement for racE, we developed a method to observe cytokinesis in suspension. Using this approach, we found that racE null cells attempt cytokinesis in suspension by forming a contractile ring and cleavage furrow. However, the cells form multiple blebs and fail in cytokinesis by regression of the cleavage furrow. We believe this phenotype is caused by the extremely low level of cortical tension found in racE null cells compared to wild-type cells. The reduced cortical tension of racE null cells is not caused by a decrease in their content of F-actin. Instead, mitotic racE null cells contain abnormal F-actin aggregates. These results suggest that racE is essential for the organization of the cortical cytoskeleton to maintain proper cortical integrity. This function of racE is independent of attachment to a substrate, but can be bypassed by other signaling pathways induced by adhesion to a substrate.

Balance of actively generated contractile and resistive forces controls cytokinesis dynamics

“…The parameters for ⌬ and S c (given in the Fig. 4 legend), which were used to obtain the calculated trajectories, are reasonable given available measured values (14,15). The RacE͞dynacortin mutant trajectory was best modeled by allowing the stretch modulus and a positive ⌬ to drive furrow ingression, consistent with the idea that these proteins slow furrow thinning by providing a brake, which counteracts the Laplace pressure.…”

“…Thus, a feedback mechanism may slow down furrow ingression if the plasma membrane ruptures from the cortex. Blebbing has been observed for nonadherent RacE 24E cells (14). Because nonadherent Dictyostelium have less polymeric actin (14), nonadherent RacE 24E and adherent RacE 24E :dynhp cells may have similarly altered cortical mechanics.…”

“…Blebbing has been observed for nonadherent RacE 24E cells (14). Because nonadherent Dictyostelium have less polymeric actin (14), nonadherent RacE 24E and adherent RacE 24E :dynhp cells may have similarly altered cortical mechanics. When we averaged the fastest velocities from the steepest part of the curves of the individual raw data sets, the velocities were 0.31 Ϯ 0.040 m͞s (n ϭ 16) for ⌬RacE:dynhp and 0.19 Ϯ 0.037 m͞s (n ϭ 10) for RacE 24E :dynhp with the fastest observed velocities at 0.74 m͞s and 0.44 m͞s for ⌬RacE:dynhp and RacE 24E :dynhp, respectively.…”

“…1A). Each protein plays an important role in cytokinesis and cortical mechanics (7,(13)(14)(15)(16)(17)(18). Each mutant strain follows a unique time-dependent trajectory of furrow ingression.…”

Deregulation of HEF1 Impairs M-Phase Progression by Disrupting the RhoA Activation Cycle

“…We and others have also previously determined that overexpression of HEF1 impacts cell spreading, causing initial enhancement of spreading, and subsequent retraction van Seventer et al, 2001;Fashena et al, 2002). Defective cellular attachment can independently result in cytokinetic failure (Orly and Sato, 1979;Ben-Ze'ev and Raz, 1981), and it is well established in some models that force generation arising from spreading at the end of telophase assists in cytokinesis (Burton and Taylor, 1997;Gerald et al, 1998;Nagasaki et al, 2002). However, we have determined that plating HEF1-induced cells on fibronectin or laminin to enhance cell spreading had no appreciable effect on the frequency of defective cytokinesis or number of rounded cells with persistent post-mitotic bridges (our unpublished data).…”