Chromosome replication and transmission are essential for the inheritance of genetic traits, but the mechanisms responsible for these processes remain poorly understood in multicellular eukaryotes. The centromere, which appears as a constriction in metaphase chromosomes, is associated with the kinetochore and serves as the key attachment site to the spindle during mitosis and meiosis. A pressing question in the centromere field today is how centromere identity is propagated from one generation to the next in multicellular eukaryotes. Elucidating the determinants of centromere identity, propagation and function requires identification of the gene products that promote centromere formation and function in trans, and ultimately determining how they interact with the cis-acting elements to perform their essential functions. In essence, we need to understand the organization, replication, and modification of centromeric chromatin. The multifaceted genetic, molecular, cell biological and biochemical approaches required to address these complex questions in higher eukaryotes are likely to succeed in Drosophila. There is over a century of experimental analyses using the fruit fly, which facilitates sophisticated in vivo analyses. Here, we propose genetic, molecular, cell biological and biochemical experiments designed to identify and characterize gene products that promote the assembly and propagation of centromeric chromatin, and to determine their properties and functions. Our entry point into centromeric chromatin will be a conserved histone H3-1ike protein (CID, for Centromere IDentifier) that localizes exclusively to functional centromeres. We will use genetic screens and biochemical isolations to identify genes and proteins that interact with CID, then use a variety of approaches to determine their roles in centromere function. These studies will address specific hypotheses and provide the groundwork for future analysis of inheritance and centromere function in Drosophila and other higher eukaryotes, such as humans.
| Zhang, Weiguo; Mao, Jian-Hua; Zhu, Wei et al. (2016) Centromere and kinetochore gene misexpression predicts cancer patient survival and response to radiotherapy and chemotherapy. Nat Commun 7:12619 |
| Langley, Sasha A; Karpen, Gary H; Langley, Charles H (2014) Nucleosomes shape DNA polymorphism and divergence. PLoS Genet 10:e1004457 |
| Dunleavy, Elaine M; Zhang, Weiguo; Karpen, Gary H (2013) Solo or doppio: how many CENP-As make a centromeric nucleosome? Nat Struct Mol Biol 20:648-50 |
| Dunleavy, Elaine M; Beier, Nicole L; Gorgescu, Walter et al. (2012) The cell cycle timing of centromeric chromatin assembly in Drosophila meiosis is distinct from mitosis yet requires CAL1 and CENP-C. PLoS Biol 10:e1001460 |
| Zhang, Weiguo; Colmenares, Serafin U; Karpen, Gary H (2012) Assembly of Drosophila centromeric nucleosomes requires CID dimerization. Mol Cell 45:263-9 |
| Mellone, Barbara G; Grive, Kathryn J; Shteyn, Vladimir et al. (2011) Assembly of Drosophila centromeric chromatin proteins during mitosis. PLoS Genet 7:e1002068 |
| Dunleavy, Elaine M; Almouzni, Geneviève; Karpen, Gary H (2011) H3.3 is deposited at centromeres in S phase as a placeholder for newly assembled CENP-A in G? phase. Nucleus 2:146-57 |
| Williamson, Adam; Wickliffe, Katherine E; Mellone, Barbara G et al. (2009) Identification of a physiological E2 module for the human anaphase-promoting complex. Proc Natl Acad Sci U S A 106:18213-8 |
| Karpen, Gary H (2009) Preparation of high-molecular-weight DNA from Drosophila embryos. Cold Spring Harb Protoc 2009:pdb.prot5254 |
| Mellone, Barbara G; Zhang, Weiguo; Karpen, Gary H (2009) Frodos found: Behold the CENP-a ""Ring"" bearers. Cell 137:409-12 |
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