Microinjected F-actin into dividing newt eggs moves toward the next cleavage furrow together with Ca2+ stores with inositol 1,4,5-trisphosphate receptor in a microtubule- and microtubule motor-dependent manner

Abstract

It has been reported that F-actin is transported to the presumptive cleavage furrow along the cortex during anaphase-cytokinesis, an event termed cortical actin flow in animal cultured cells. The motor source has remained unknown. We reported that Ca2+ stores with IP3 receptor (IP3R) was re-distributed from the polar cortex during metaphase to the presumptive cleavage furrow just before the onset of furrowing, and that Ca2+ stores with IP3R microinjected into dividing newt eggs moved toward the presumptive cleavage furrow during anaphase-cytokinesis in a microtubule-dependent manner, and that Ca2+ store-enriched microsome fractions induced the cleavage furrow as the putative cleavage stimulus. Because the distribution of F-actin and Ca2+ stores with IP3R during metaphase to cytokinesis is similar, we considered that this cortical actin flow may be powered by transportation of Ca2+ stores with IP3R. Purified F-actin labeled with phalloidin-rhodamine was microinjected into the dividing newt eggs and the eggs observed under a confocal microscope. We found that the microinjected F-actin moved linearly toward the next cleavage furrow and that this movement was blocked by nocodazole, microtubule-depolarizing agent and AMP-PNP, a blocking agent of microtubule motors. Co-microinjected rhodamine-labeled F-actin and sacro/endoplasmic reticulum Ca2+-ATPase (SERCA)-GFP-labeled Ca2+ stores with IP3R co-moved and co-accumulated to the next cleavage furrow. These results strongly suggest that Ca2+ stores with IP3R, which is transferred by microtubule-based motility as cleavage stimulus, act as an F-actin translocator.