Making CENs of mammalian artificial chromosomes

Mammalian artificial chromosomes (MACs) hold the promise of providing autonomous vectors for gene therapy in dividing cells. They would not require insertion into the genome and could include sufficient genomic sequences that surround the therapeutic gene to ensure proper tissue-specific and temporal regulation. Several groups have reported successful formation of MACs in human cells using transfection strategies that included alpha satellite DNA, the primary DNA found at normal human centromeres. These results, although extremely encouraging, have limitations such as unpredictable chromosome formation and success thus far in only one transformed human cell line. Examination of other cells where alpha satellite DNA has integrated into ectopic chromosomal locations, as well as naturally occurring dicentric and neocentromere-containing cell lines, suggests that alpha satellite DNA may not be necessary or sufficient for centromere formation. Overall, these results suggest that epigenetic modifications of centromeric DNA are required for efficient centromere formation. Models for this centromere-specific epigenetic modification include a specialized chromatin structure and differential replication timing of centromeric DNA. Thus, further investigation of these centromere-specific epigenetic modifications may suggest strategies for increasing the efficiency of generating human artificial chromosomes for use as gene therapy vectors.