Depicted is a cross section through the centrosome: CP91 resides in the inner core layer as two distinguishable layers (darker yellow) and in close proximity to Cep192 (blue), which is located in the outer core layers. BioID2 for expression in and applying a knock-in strategy for the expression of BioID2-tagged centrosomal fusion proteins. Thus, we were able to identify various centrosomal Cep192 interaction partners, including CDK5RAP2, which was previously allocated to the inner corona structure, and several core components. Studies employing overexpression of GFP-Cep192 as well as depletion of endogenous Cep192 revealed that Cep192 is a key protein for the recruitment of corona components during centrosome biogenesis and is required to maintain a stable corona structure. centrosome provides the best-established model for an acentriolar centrosome outside the Opisthokonta. The centrosome consists of a layered core structure surrounded by a corona, in which -tubulin containing nodules are embedded. According to ultrastructural analyses, the core structure consists VEZF1 of three major layers: two outer layers with slightly lower electron density and one central layer with very high electron density [7,8]. A closer look reveals that these major layers can be subdivided into several sublayers. Although earlier work suggested that the layers make up a box-shaped core structure [9], now there is no doubt that they form a cylindrical stack of layers [8,10] as was also found in the related amoebozoan [11]. Layered structures occur also in yeasts; however, they are most likely analogous to the core layers, not homologous [1]. Unlike in centriolar centrosomes, duplication of the centrosome does not take place during S-phase but starts only at the G2/M transition [12]. First, the whole centrosome increases in size and the corona dissociates, along with the microtubule-nucleation complexes. This is accompanied by the disassembly of all pre-existing microtubules. Next, the CCT251545 remaining core structure enters a fenestra in the nuclear envelope, and the central layer disappears. In prometaphase the remaining outer layers start to CCT251545 separate, each residing in its own fenestra in the nuclear envelope. The former outer layers act as mitotic centrosomes, and upon separation they nucleate spindle microtubules, CCT251545 forming a central spindle. In metaphase, astral microtubules appear. Starting with anaphase, the plaque-shaped mitotic centrosomes undergo a folding process, in which the inner, microtubule-nucleating surface becomes more and more exposed to the cytoplasm. In telophase, the folding process of each mitotic centrosome completes with a scission at the kink of the fold, and the re-appearance of the central layer. This process implicates an inside-to-outside reversal of the outer layers in each cell cycle [12] and implies that the two CCT251545 outer layers have the same protein composition. The new centrosomes then exit their fenestrae in the nuclear envelope, but remain attached to the cytosolic surface of the nucleus. At this time, the microtubule nucleating surface of the new core structure differentiates into the new corona. Our group is still making progress in characterizing the complete set of structural and regulatory proteins involved in this process. Meanwhile, forty-two proteins have been identified as centrosomal or centrosome-associated (reviewed by Gr?f et al. in this special issue of Cells). However, despite recent advances in the assignment of novel centrosomal core proteins to the corona and the individual core layers [13,14,15,16,17], the composition of the two outer layers and their interaction with the corona components have remained elusive. CCT251545 In this paper we focus on Cep192. In mammalian cells, Cep192 (SPD-2 in orthologue of Cep192 is the major component of the outer core layers, and that it interacts with CDK5RAP2, a major recruiting factor of -tubulin complexes. In analogy to animal.
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