MECHANISM OF NUCLEOSOME DISSOCIATION PRODUCED BY TRANSCRIPTION ELONGATION IN A SHORT CHROMATIN TEMPLATE

We have used a linear DNA template (239 bp) containing a nucleosome po sitioning sequence (NX1) downstream of the T7 RNA polymerase promoter to study the mechanism of transcription elongation through a nucleosom e, Under ionic strength approaching physiological conditions we have o bserved that transcription causes nucleosome dissociation and histone redistribution within the template, We have examined the role of the d ifferent elements that, in principle, could induce nucleosome dissocia tion during transcription, The high affinity of histones for single-st randed DNA observed in titration experiments performed using the purif ied (+) and (-) strands of the NX1 fragment suggests that nucleosome d issociation is not due to the formation of segments of single-stranded DNA by RNA polymerase in the elongation process, Furthermore, our res ults show that although RNA can interact with core histones, the synth esized RNA is not bound to the histones dissociated by transcription, Our results indicate that core histones released during transcription can be bound to naked DNA and chromatin (with or without histones H1-H 5). From the dynamic properties of excess histones bound to chromatin, we suggest a nucleosome transcription mechanism in which displaced hi stones are transiently bound to chromatin and finally are reassembled with DNA after the passage of the polymerase.