Unusual chromosome architecture and behaviour at an HSR

Amplification of sequences within mammalian chromosomes is often accompanie d by the formation of homogeneously staining regions (HSRs). The arrangemen t of DNA sequences within such amplicons has been investigated, but little is known about the chromosome structure or behaviour of these unusual regio ns. We have analysed the metaphase chromosome structure of the dihydrofolat e reductase (DHFR) amplicon of CHOC400 cells. The chromatin in this region contains hyperacetylated nucleosomes yet, at the same time, appears to be d ensely packed like heterochromatin. The region does not bind heterochromati n proteins. We show that the dense packing of the region is restricted to D NA located close to the chromosome core/scaffold. In contrast, levels of th e chromosome scaffold protein topoisomerase II at HSRs are the same as thos e found at other euchromatic locations. Metaphase chromosome condensation o f the HSR is shown to be sensitive to topoisomerase II inhibitors, and sist er chromatids often appear to remain attached within the HSRs at metaphase. We suggest that these features underlie anaphase bridging and the aberrant interphase structure of the HSR. The DHFR amplicon is widely used as a mod el system to study mammalian DNA replication. We conclude that the higher-o rder chromosome structure of this amplicon is unusual and suggest that caut ion needs to be exercised in extrapolating data from HSRs to normal chromos omal loci.