The activation of a neocentromere in Drosophila requires proximity to an endogenous centromere.

The centromere is essential for proper segregation and inheritance of genetic information. Centromeres are generally regulated to occur exactly once per chromosome; failure to do so leads to chromosome loss or damage and loss of linked genetic material. The mechanism for faithful regulation of centromere activity and number is unknown. The presence of ectopic centromeres (neocentromeres) has allowed us to probe the requirements and characteristics of centromere activation, maintenance, and structure. We utilized chromosome derivatives that placed a 290-kilobase "test segment" in three different contexts within the Drosophila melanogaster genome--immediately adjacent to (1) centromeric chromatin, (2) centric heterochromatin, or (3) euchromatin. Using irradiation mutagenesis, we freed this test segment from the source chromosome and genetically assayed whether the liberated "test fragment" exhibited centromere activity. We observed that this test fragment behaved differently with respect to centromere activity when liberated from different chromosomal contexts, despite an apparent sequence identity. Test segments juxtaposed to an active centromere produced fragments with neocentromere activity, whereas test segments far from centromeres did not. Once established, neocentromere activity was stable. The imposition of neocentromere activity on juxtaposed DNA supports the hypothesis that centromere activity and identity is capable of spreading and is regulated epigenetically.