Identification of novel Drosophila centromere‐associated proteins. Issue 19 (14th July 2014)

• Record Type: Journal Article
• Authors: Barth, Teresa K.; Schade, Georg O. M.; Schmidt, Andreas; Vetter, Irene; Wirth, Marc; Heun, Patrick; Thomae, Andreas W.; Imhof, Axel; Vondriska, Thoms M.; Jensen, Ole N.
• Is Part Of: Proteomics. Volume 14:Issue 19(2014:Oct.)
• Page Start: 2167
• Page End: 2178
• Publication Date: 2014-07-14
• Subjects: Proteins -- Separation -- Periodicals; Bioinformatics -- Periodicals; Proteomics -- Periodicals; Genomes -- Periodicals; Molecular genetics -- Periodicals; 572.605
• DOI: 10.1002/pmic.201400052
• Languages: English
• ISSNs: 1615-9853

Abstract

<abstract abstract-type="main"> <title> <x xml:space="preserve">Abstract</x> </title> <p>Centromeres are chromosomal regions crucial for correct chromosome segregation during mitosis and meiosis. They are epigenetically defined by centromeric proteins such as the centromere‐specific histone H3‐variant centromere protein A (CENP‐A). In humans, 16 additional proteins have been described to be constitutively associated with centromeres throughout the cell cycle, known as the constitutive centromere‐associated network (CCAN). In contrast, only one additional constitutive centromeric protein is known in <italic>Drosophila melanogaster (D.mel)</italic>, the conserved CCAN member CENP‐C. To gain further insights into <italic>D.mel</italic> centromere composition and biology, we analyzed affinity‐purified chromatin prepared from <italic>D.mel</italic> cell lines expressing green fluorescent protein tagged histone three variants by MS. In addition to already‐known centromeric proteins, we identified novel factors that were repeatedly enriched in affinity purification‐MS experiments. We analyzed the cellular localization of selected candidates by immunocytochemistry and confirmed localization to the centromere and other genomic regions for ten factors. Furthermore, RNA interference mediated depletion of CG2051, CG14480, and hyperplastic discs, three of our strongest candidates, leads to elevated mitotic defects. Knockdowns of these candidates neither impair the localization of several known kinetochore proteins nor CENP‐A<sup>CID</sup> loading, suggesting their involvement in alternative pathways that contribute to proper centromere function. In summary, we provide a comprehensive analysis of the proteomic composition of <italic>Drosophila</italic> centromeres. All MS data have been deposited in the ProteomeXchange with identifier PXD000758 (<ext-link ext-link-type="uri" xlink:href="http://proteomecentral.proteomexchange.org/dataset/PXD000758" xlink:type="simple" xmlns:xlink="http://www.w3.org/1999/xlink">http://proteomecentral.proteomexchange.org/dataset/PXD000758</ext-link>).</p> </abstract>