A novel chromatin immunoprecipitation and array (CIA) analysis identifies a 460-kb CENP-A-binding neocentromere DNA

Centromere protein A (CENP-A) is an essential histone H3-related protein th at constitutes the specialized chromatin of an active centromere. It has be en suggested that this protein plays a key role in the epigenetic marking a nd transformation of noncentromeric genomic DNA into functional neocentrome res. Neocentromeres have been identified on more than two-thirds of the hum an chromosomes, presumably involving different noncentromeric DNA sequences , but it is unclear whether some generalized sequence properties account fo r these neocentromeric sites. Using a novel method combining chromatin immu noprecipitation and genomic array hybridization, we have identified a 460-k b CENP-A-binding DNA domain of a neocentromere derived from the 20p12 regio n of an invdup (20p) human marker chromosome. Detailed sequence analysis in dicates that this domain contains no centromeric alpha -satellite, classica l satellites, or other known pericentric repetitive sequence motifs. Putati ve gene loci are detected, suggesting that their presence does not preclude neocentromere formation. The sequence is not significantly different from surrounding non-CENP-A-binding DNA in terms of the prevalence of various in terspersed repeats and binding sites for DNA-interacting proteins (Topoisom erase II and High-Mobility-Group protein I). Notable variations include a h igher AT content similar to that seen in human alpha -satellite DNA and a r educed prevalence of long terminal repeats (LTRs), short interspersed repea ts (SINEs), and Alus. The significance of these features in neocentromeriza tion is discussed.