Asynchronous replication of homologous loci on human active and inactive X chromosomes

The two X chromosomes in mammalian females replicate asynchronously, the inactive later than the active one. Using BrdUrd-sensitive restriction and UV irradiation to identify newly synthesized DNA directly on Southern blots, and restriction fragment length differences to discriminate alleles on active and inactive human X chromosomes, we examined the replication of hypoxanthine phosphoribosyltransferase (HPRT) and clotting factor IX (F9) loci in clonal populations of mouse-human hybrids. We find that HPRT replicates at different times during the period of DNA synthesis (S phase), depending on its activity: It replicates in early S phase, when expressed (on the active X chromosome), and in late S phase when silent (on the inactive X chromosome). Furthermore, when reactivated, the derepressed locus is earlier replicating, supporting a relationship between replication and transcription. Neither F9 allele is expressed in these cells, and both replicate in the second half of S phase, (slightly earlier on active than on inactive X chromosome).