Redistribution of Actin during Assembly and Reassembly of the Contractile Ring in Grasshopper Spermatocytes

Abstract

Cytokinesis in animal cells requires the assembly of an actomyosin contractile ring to cleave the cell. The ring is highly dynamic; it assembles and disassembles during each cell cleavage, resulting in the recurrent redistribution of actin. To investigate this process in grasshopper spermatocytes, we mechanically manipulated the spindle to induce actin redistribution into ectopic contractile rings, around reassembled lateral spindles. To enhance visualization of actin, we folded the spindle at its equator to convert the remnants of the partially assembled ring into a concentrated source of actin. Filaments from the disintegrating ring aligned along reorganizing spindle microtubules, suggesting that their incorporation into the new ring was mediated by microtubules. We tracked incorporation by speckling actin filaments with Qdots and/or labeling them with Alexa 488-phalloidin. The pattern of movement implied that actin was transported along spindle microtubules, before entering the ring. By double-labeling dividing cells, we imaged actin filaments moving along microtubules near the contractile ring. Together, our findings indicate that in one mechanism of actin redistribution, actin filaments are transported along spindle microtubule tracks in a plus-end–directed fashion. After reaching the spindle midzone, the filaments could be transported laterally to the ring. Notably, actin filaments undergo a dramatic trajectory change as they enter the ring, implying the existence of a pulling force. Two other mechanisms of actin redistribution, cortical flow and de novo assembly, are also present in grasshopper, suggesting that actin converges at the nascent contractile ring from diffuse sources within the cytoplasm and cortex, mediated by spindle microtubules.

Dissecting Contractile Ring Assembly Using a Multimode Microtechnique

“…2). In light of our recent findings in spermatocytes of silkworms [3] and grasshoppers [4], these results demonstrate a mechanism by which peripheral spindle microtubules could mechanically sweep cortical actin filaments into the incipient contractile ring. …”

F-actin localization dynamics during appressorium formation inColletotrichum graminicola

“…In animal cells there are two sources of actin filaments involved in the assembly of the contractile ring: de novo assembly around the cell equator and recruitment of pre-existing actin filaments (Cao andWang 1990a, Chen et al 2008). The pre-existing actin filaments are redistributed to the cleavage furrow by cortical flow or via spindle microtubules (Cao and Wang 1990b, Zhou and Wang 2008, Alsop et al 2009). In fission yeast the de novo-assembled F-actin is essential to the assembly of the contractile ring (Pelham and Chang 2002).…”

Astral microtubules physically redistribute cortical actin filaments to the incipient contractile ring

“…This has also been demonstrated in Drosophila spermatocytes . Similar flow of cortical actin has also been seen in grasshopper spermatocytes [Alsop et al, 2009] and cultured mammalian cells [Zhou and Wang, 2008]. In silkworm spermatocytes, manipulation of dynamic microtubules can drive the flow of cortical actin to the site of cleavage [Chen et al, 2008].…”

“…In other words, microtubules from the newly created monopolar spindle elongated toward and interacted with the cortex, resulting in the directionally biased movement of actin away from the spindle pole [Alsop et al, 2009]. In other words, microtubules from the newly created monopolar spindle elongated toward and interacted with the cortex, resulting in the directionally biased movement of actin away from the spindle pole [Alsop et al, 2009].…”