In most eukaryotes, centromeres are epigenetically defined by nucleosomes that contain the histone H3 variant centromere protein A (CENP-A). Specific targeting of the CENP-A-loading chaperone to the centromere is vital for stable centromere propagation; however, the existence of ectopic centromeres (neocentromeres) indicates that this chaperone can function in different chromatin environments. The mechanism responsible for accommodating the CENP-A chaperone at noncentromeric regions is poorly understood. Here, we report the identification of transient, immature neocentromeres in Schizosaccharomyces pombe that show reduced association with the CENP-A chaperone Scm3, owing to persistence of the histone H2A variant H2A.Z. After the acquisition of adjacent heterochromatin or relocation of the immature neocentromeres to subtelomeric regions, H2A.Z was depleted and Scm3 was replenished, thus leading to subsequent stabilization of the neocentromeres. These findings provide new insights into histone variant-mediated epigenetic control of neocentromere establishment.