Jcb_201203166 1..14

Abstract

Correct spindle position is an essential feature of cell division from yeast to man. In animal cells, the cleavage furrow is specified so as to bisect the mitotic spindle. Therefore, correct orientation of the spindle along a given axis, as well as its accurate placement on this axis (hereafter collectively referred to as “spindle positioning” for simplicity), is critical for dictating the relative size of daughter cells and for ensuring the proper distribution of cytoplasmic constituents at cell division. Spindle positioning plays an important role during development of metazoan organisms, as well as in stem cell lineages. Despite important progress in recent years, the mechanisms governing spindle positioning remain incompletely understood. Molecules important for spindle positioning have been identified notably in invertebrate systems, including Drosophila melanogaster neuroblasts and Caenorhabditis elegans embryos (reviewed by Gönczy, 2008, Knoblich, 2008; Siller and Doe, 2009). In C. elegans one-cell stage embryos, for instance, forward genetic and RNAi-based functional genomic screens have uncovered that a ternary complex, as well as the minus end– directed motor dynein, is essential for generating pulling forces that act on astral microtubules to position the spindle. This ternary complex comprises the partially redundant heterotrimeric G proteins GOA-1 and GPA-16 and the essentially identical GoLoco proteins GPR-1 and GPR-2 (hereafter collectively referred as GPR-1/2) as well as the coiled-coil domain protein LIN-5 (Gotta and Ahringer, 2001; Colombo et al., 2003; Gotta et al., 2003; Srinivasan et al., 2003; Nguyen-Ngoc et al., 2007). Dynein is a multisubunit AAA ATPase motor protein complex of >1.5 MD that is fundamental for several cellular processes across eukaryotic evolution, including proper organelle distribution, spindle assembly, and kinetochore function (reviewed by Kardon and Vale, 2009). This complex comprises the dynein heavy chain motor protein and dynein intermediate and light chains, as well as several additional factors, including the dynactin complex that is needed for dynein activity. In C. elegans embryos, coimmunoprecipitation experiments indicate that the ternary complex associates with dynein (Couwenbergs et al., 2007; Nguyen-Ngoc et al., 2007; Park and Rose, 2008). Moreover, the presence of dynein at the cell cortex is compromised upon depletion of ternary complex components in early worm embryos (Nguyen-Ngoc et al., 2007). Together, these observations have led to a working model in which the ternary complex would bring dynein to the plasma membrane owing to Correct spindle positioning is fundamental for proper cell division during development and in stem cell lineages. Dynein and an evolutionarily conserved ternary complex (nuclear mitotic apparatus protein [NuMA]–LGN–G in human cells and LIN-5–GPR-1/2–G in Caenorhabditis elegans) are required for correct spindle positioning, but their relationship remains incompletely understood. By analyzing fixed specimens and conducting live-imaging experiments, we uncovered that appropriate levels of ternary complex components are critical for dynein-dependent spindle positioning in HeLa cells and C. elegans embryos. Moreover, using mutant versions of G in both systems, we established that dynein acts at the membrane to direct spindle positioning. Importantly, we identified a region within NuMA that mediates association with dynein. By using this region to target dynein to the plasma membrane, we demonstrated that the mere presence of dynein at that location is sufficient to direct spindle positioning in HeLa cells. Overall, we propose a model in which the ternary complex serves to anchor dynein at the plasma membrane to ensure correct spindle positioning. Cortical dynein is critical for proper spindle positioning in human cells

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Cite this paper

@inproceedings{Kotak2012Jcb_2012031661, title={Jcb_201203166 1..14}, author={Sachin Kotak and Coralie Busso}, year={2012} }