[Distribution of transcription-active DNA sequences in rat liver chromatin during fragmentation with endogenous...

Distribution of the transcriptionally active sequences in rat liver chromatin DNA fragments released from chromatin sites with different sensitivity to endogenous Ca2+, Mg2+-DNases was studied by the dot hybridization method with cloned DNA-probes. The internal fragment of the rat chromosomal ceruloplasmin gene containing coding sequences and the promoter region of the metallothionein-I gene were specific probes for the transcriptionally active liver chromatin. cDNA of beta-globin gene was the control for the transcriptionally inert DNA sequences. Mononucleosomal length, DNA (175-215 bp) formed in the course of mile nuclease digestion (less than 1% mononucleosomes and acid-soluble material) was 3-6-fold enriched in transcribed sequences of the ceruloplasmin gene as compared to oligonucleosomal DNA fragments produced in the same digestion conditions (fractions containing fragments 750-850 and greater than 2000 bp). The relative amount of the ceruloplasmin gene in mononucleosomal length DNA formed during the mild digestion was 12-25-fold greater than in total rat liver DNA and 25-50-fold greater than in the mononucleosomal DNA (150-175 bp) produced in the course of extensive digestion (80-85% mononucleosomes and 15-20% acid-soluble material) by endogenous Ca2+, Mg2+-DNases. The promoter metallothionein-I gene sequences exhibited the same distribution among chromatin fragments as the coding ceruloplasmin gene sequence. Distribution of the nontranscribed beta-globin sequences in liver chromatin DNA fragments produced at different stages of nuclease digestion was not related to chromatin sensitivity to endogenous DNases. These sequences were distributed similarly in different DNA size classes and total DNA.