Facilitated Transcription through the Nucleosome at High Ionic Strength Occurs via a Histone Octamer Transfer Mechanism

Wendy Walter,Vasily M Studitsky

doi:10.1074/jbc.m103704200

Abstract

The rate of transcription through the nucleosome, the fine structure of the nucleosomal barrier, and the fate of the nucleosome during transcription at different salt concentrations were analyzed using linear 227-base pair mononucleosomal templates containing a uniquely positioned nucleosome core. At lower ionic strength (30 mm NaCl), the nucleosome constitutes a strong barrier for SP6 RNA polymerase. At higher ionic strength (330 mm NaCl), the rates of transcription on nucleosomal and histone-free DNA templates are very similar. At both higher and lower ionic strengths, the complete histone octamer is transferred over the same distance by fundamentally similar mechanisms. The data indicate that even at the rate of transcription characteristic of histone-free DNA, the transfer intermediates can be formed quite efficiently. This suggests possible mechanisms that could facilitate transcription through the nucleosome at physiological ionic strength.

Highlights

The most basic unit of DNA packaging in eukaryotes is the nucleosome
Increasing the salt concentration to 0.3– 0.5 M NaCl (0.15 M NaCl is considered to be physiological ionic strength) during transcription allows the rate of transcription of chromatin templates to reach those of naked DNA [11, 12], recapitulating the highly efficient transcription of chromatin templates in vivo
We have shown that an increase in ionic strength progressively weakens the nucleosomal barrier to transcription (Fig. 2)

Summary

Introduction

The most basic unit of DNA packaging in eukaryotes is the nucleosome. A nucleosome core particle consists of 146 bp1 of DNA wrapped around a histone octamer. At higher ionic strength (330 mM NaCl), the rates of transcription on nucleosomal and histone-free DNA templates are very similar. The relative rates of transcription were analyzed in reactions containing increasing concentrations of salt for both DNA and nucleosomal templates, and transcribed nucleosomal templates were analyzed for nucleosome displacement at both higher and lower ionic strength.

Results

Conclusion