Résumé
The method described here allows the mapping of protein-DNA interactions through the ability of formaldehyde to cross-link proteins and nucleic acids in living cells. Formaldehyde is a very reactive dipolar compound which reacts with the amino groups of proteins and amino acids (1, 2). It shows no reactivity, however, towards free double-stranded DNA, and thus does not cause the extensive DNA damage seen after prolonged exposure to other crosslinking reagents such as UV. Each formaldehyde molecule has the capacity to interact with two amino groups. Therefore DNA-protein, protein-protein, and RNA-protein cross-links are rapidly formed after formaldehyde treatment, creating a stable structure which prevents the redistribution of cellular components. Furthermore, a simple heat treatment is sufficient to reverse the reaction equilibrium, and to allow the isolation of pure DNA for further analysis (3). Formaldehyde crosslinking combined with chromatin immuno precipitation (IP) is a way of mapping the in vivo distribution of chromatin• associated proteins. As such, this technique is of great value in the analysis of protein-DNA interactions, even more so when studying proteins which do not show specific DNA binding activities in vitro. This has recently been demonstrated for several chromatin associated proteins, such as Polycomb (PC) in Drosophila (4) and the Silent Information Regulators (SIR) proteins in budding yeast (5). Additionally, analysis of the crosslinking pattern can not only allow mapping of sites of protein-DNA interaction, but can also give an estimation of the relative binding affinity to different sequences across a large genomic region.