Novel asymmetrically localizing components of human centrosomes identified by complementary proteomics methods

Department of Biochemistry and Molecular Biology, University of Southern Denmark, Odense, Denmark.
The EMBO Journal (Impact Factor: 10.43). 03/2011; 30(8):1520-35. DOI: 10.1038/emboj.2011.63
Source: PubMed


Centrosomes in animal cells are dynamic organelles with a proteinaceous matrix of pericentriolar material assembled around a pair of centrioles. They organize the microtubule cytoskeleton and the mitotic spindle apparatus. Mature centrioles are essential for biogenesis of primary cilia that mediate key signalling events. Despite recent advances, the molecular basis for the plethora of processes coordinated by centrosomes is not fully understood. We have combined protein identification and localization, using PCP-SILAC mass spectrometry, BAC transgeneOmics, and antibodies to define the constituents of human centrosomes. From a background of non-specific proteins, we distinguished 126 known and 40 candidate centrosomal proteins, of which 22 were confirmed as novel components. An antibody screen covering 4000 genes revealed an additional 113 candidates. We illustrate the power of our methods by identifying a novel set of five proteins preferentially associated with mother or daughter centrioles, comprising genes implicated in cell polarity. Pulsed labelling demonstrates a remarkable variation in the stability of centrosomal protein complexes. These spatiotemporal proteomics data provide leads to the further functional characterization of centrosomal proteins.

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    • "See also Figureidentify Mid1-specific interactors that would explain its role in cytokinesis; thus, it will be important to determine whether Mid1 has a direct influence on a process that regulates cell division or whether it has substrates that regulate cell division that are as yet unidentified. Second, the Mid2 interactome also identified ASPM (abnormal spindle-like, microcephaly-associated) and CEP128 (centromeric protein 128), which associate with centrosomes, and ASPM has also been linked to the regulation of spindle assembly and cytokinesis (Higgins et al., 2010;Jakobsen et al., 2011;Paramasivam et al., 2007); thus, these proteins should be explored further as potential Mid2 substrates. "
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