The DNA in chromatin is tightly wound around nucleosomes — but how does it get that way? Although several factors are known to deposit histones on DNA during DNA synthesis, a report in Molecular Cell now uncovers an independent chromatin-assembly pathway that is not coupled to DNA replication.

Geneviève Almouzni and colleagues were studying HIRA, a protein that was previously shown to interact with histones in mammals. They first isolated the homologue of HIRA in Xenopus laevis, and showed that GST–HIRA could bind to all four core histones (H2A, H2B, H3 and H4) with varying affinities.

The authors next used a supercoiling assay to show that HIRA can facilitate the formation of nucleosomes in vitro. They mixed purified histones with a relaxed DNA plasmid in the presence of recombinant HIRA, then monitored the electrophoretic mobility of the plasmid. Nucleosome assembly causes supercoiling of the DNA, and hence faster migration on a gel.

Almouzni and co-workers then studied nucleosome assembly using Xenopus egg extracts, which are very efficient at chromatin assembly and are enriched in HIRA. Although these extracts could promote nucleosome assembly, extracts that had been immunodepleted of HIRA could not. However, depletion of HIRA did not affect the chromatin-assembly activity that is coupled to nucleotide-excision repair or to DNA synthesis.

These results, conclude the authors, “show that HIRA is critical in a specific chromatin-assembly process that is not coupled to DNA synthesis”. What they also emphasize is that there are at least two distinct nucleosome-assembly pathways.