MAPPING PROTEIN DNA INTERACTIONS INVIVO WITH FORMALDEHYDE - EVIDENCE THAT HISTONE-H4 IS RETAINED ON A HIGHLY TRANSCRIBED GENE

We have used formaldehyde-mediated protein-DNA crosslinking within intact cells to examine the in vivo chromatin structure of the D. melanogaster heat shock protein 70 (hsp70) genes. In agreement with previous in vitro studies, we find that the heat shock-mediated transcriptional induction of the hsp70 genes perturbs their chromatin structure, resulting in fewer protein-dna contacts crosslinkable in vivo by formaldehyde. However, contrary to earlier in vitro eivdence that histones may be absent form actively trnascribed genes, we show directly by immunoprecopitation of in vivo-crosslinked chromatin fragments, that at least histone H4 remains bound to hsp70 DNA in vivo, irrespecitve of its rate of transcription. The formaldehyde-based in vivo mapping techniques described in this work are generally applicable, and can be used both to probe protein-DNA interactions with in specific genes and to determine the genomic location of specific chromosomal proteins.