A molecular mechanism of mitotic centrosome assembly in Drosophila

Abstract

Centrosomes comprise a pair of centrioles surrounded by pericentriolar material (PCM). The PCM expands dramatically as cells enter mitosis, but it is unclear how this occurs. In this study, we show that the centriole protein Asl initiates the recruitment of DSpd-2 and Cnn to mother centrioles; both proteins then assemble into co-dependent scaffold-like structures that spread outwards from the mother centriole and recruit most, if not all, other PCM components. In the absence of either DSpd-2 or Cnn, mitotic PCM assembly is diminished; in the absence of both proteins, it appears to be abolished. We show that DSpd-2 helps incorporate Cnn into the PCM and that Cnn then helps maintain DSpd-2 within the PCM, creating a positive feedback loop that promotes robust PCM expansion around the mother centriole during mitosis. These observations suggest a surprisingly simple mechanism of mitotic PCM assembly in flies.DOI: http://dx.doi.org/10.7554/eLife.03399.001

SPD-2/CEP192 and CDK Are Limiting for Microtubule-Organizing Center Function at the Centrosome

“…A similar function had previously been proposed for PLP and Cnn in Drosophila; however, neither of these proteins have clear homologs in C. elegans [29,31]. Our findings are consistent with a recent report showing that, in Drosophila, SPD-2 works together with Cnn to recruit mitotic PCM to the mother centriole [32]. The total amount of SPD-2 in the cell had previously been suggested to limit PCM size in mitotic cells [33].…”

Centrosome Biology: The Ins and Outs of Centrosome Assembly

“…However, the situation in flies seems to be a little more complex. Previously, in embryos, both Cnn and Spd-2 were shown to be initially recruited into a central region from where they subsequently moved outwards [5,6,16] ( Figure 1C). To address whether this behavior was specific to embryos or whether it also occurred in other cell types, Conduit and Raff [15] performed FRAP of Spd-2-GFP and GFP-Cnn at centrosomes in mitotic larval brain cells.…”

“…These centrosomes contained intact centrioles surrounded by an extensive salt-resistant matrix that was capable of recruiting PCM components [3,4]. Early insights into the molecular composition of a PCM scaffold came from the identification of two key centrosomal proteins, Spd-2 and Centrosomin (Cnn) in flies and SPD-2 and SPD-5 in worms, which together form a scaffold around the mother centriole that recruits other PCM components [5][6][7][8][9]. Indeed, utilizing an in vitro PCM assembly system, Woodruff et al [10] recently reported that recombinant SPD-5 polymerizes in a concentration-and time-dependent manner to form micrometer-sized porous networks.…”

“…Based on its requirement for centrosome lattice assembly and the fact that it is also phosphorylated by PLK-1, SPD-5 is assumed to be the functional homologue of Cnn in worms, yet Laos et al [14] did not report centrosomal flaring or its equivalent in C. elegans embryos. However, the findings in flies do provide a model for mitotic PCM expansion that occurs in both embryos and brain cells: Spd-2 assembles around the mother centriole, then moves outwards, where it helps to recruit other PCM components, including Cnn and Polo; Cnn then becomes phosphorylated by Polo, leading to the formation of a multimeric Cnn scaffold [5,6,15]. This scaffold allows more Spd-2 and Polo to accumulate, creating a positive feedback loop that allows more Cnn to be incorporated, thereby expanding the PCM scaffold away from the mother centriole.…”

Superresolution live imaging of plant cells using structured illumination microscopy

“…Concerning live applications, SIM may involve 2D and 3D subdiffraction resolution of widely spread intracellular structures such as plasmodesmata 34 or cell walls 35 and simultaneous dual-color time-lapse imaging of actin and clathrin-coated vesicles 36 . Of particular importance was the recent implementation of fluorescence recovery after photobleaching analysis during SIM time-lapse imaging 37 .…”