OVERCOMING A NUCLEOSOMAL BARRIER TO TRANSCRIPTION

We have studied the kinetics of transcription through a nucleosome core. RNA polymerase transcribes the first similar to 25 bp of nucleosomal DNA rapidly, but then hits a barrier and continues slowly to the nucleosomal dyad region. Here, the barrier disappears and the transcript is completed at a rapid rate, as if on free DNA, indicating that histone octamer transfer is completed as polymerase reaches the dyad, If DNA behind the polymerase is removed during transcription, the barrier does not appear until the polymerase has penetrated up to 15 bp farther into the nucleosome. On a longer template, the barrier is almost eliminated. We have shown previously that the octamer is transferred around the transcribing polymerase via an intermediate containing an intranucleosomal DNA loop, Our results exclude the possibility that polymerase has difficulty breaking histone-DNA contacts and suggest instead that polymerase pauses because it has difficulty transcribing DNA in the loop.