Specific histone-histone contacts are ruptured when nucleosomes unfold at low ionic strength

Area of Science:

• Molecular Biology
• Chromatin Structure
• Biochemistry

Background:

• Chromatin, the complex of DNA and proteins that forms chromosomes, is essential for genome organization and regulation.
• Nucleosomes, the basic repeating units of chromatin, consist of DNA wrapped around histone proteins.
• Understanding nucleosome stability and unfolding is crucial for deciphering gene regulation and DNA accessibility.

Purpose of the Study:

• To investigate the ordered unfolding of the nucleosome core at low ionic strengths.
• To identify specific histone-H2B and histone-H4 (H2B-H4) binding sites involved in nucleosome stability.
• To characterize the conformational transitions of nucleosomes under varying ionic conditions.

Main Methods:

• Gentle lysis of nuclei in low ionic strength solutions.
• Contact-site cross-linking in intact nuclei to identify protein-protein interactions.
• Ultracentrifugation of nucleosomes at various salt concentrations.
• Analysis of purified nucleosome monomers as controls.

Main Results:

• Nucleosomes rupture at a major H2B-H4 binding site during gentle lysis in very low ionic strength solutions.
• H2A-H2B interactions remain unperturbed during this unfolding process.
• At least four distinct H2B-H4 contacts are broken, confirmed by cross-linking studies.
• Sedimentation analysis reveals a significant conformational transition in the ionic strength range where H2B-H4 sites rupture.

Conclusions:

• The H2B-H4 binding site is a critical point of nucleosome unfolding at low ionic strength.
• Nucleosome unfolding is a stepwise process, with specific histone-histone interactions being more sensitive to ionic conditions.
• These findings provide insights into the dynamic nature of chromatin structure and its response to environmental changes.