The elongation factor Spn1 is a multi-functional chromatin binding protein

Area of Science:

• Molecular Biology
• Epigenetics
• Chromatin Biology

Background:

• Transcriptional elongation by RNA polymerase II (RNAPII) requires numerous factors within chromatin.
• Spn1 is a conserved protein interacting with RNAPII and the histone chaperone Spt6, known to negatively regulate Spt6-nucleosome interactions.

Purpose of the Study:

• To investigate the chromatin binding properties and functional roles of Spn1.
• To elucidate the contribution of Spn1's core domain to its interactions and functions in vivo.

Main Methods:

• Biochemical assays to test binding of Spn1 to DNA, histones, and nucleosomes.
• In vitro nucleosome assembly assays.
• Analysis of Spn1 core domain interactions with Spt6 and RNAPII.
• Chromatin immunoprecipitation to assess promoter recruitment.
• Synthetic genetic interaction analysis with histone chaperone mutants.

Main Results:

• Full-length Spn1 binds DNA, histones H3-H4, mononucleosomes, and nucleosomal arrays, exhibiting weak nucleosome assembly activity.
• Spn1's core domain shows suboptimal interactions with histones, nucleosomes, and DNA, and fails in vitro nucleosome assembly.
• The Spn1 core domain binds Spt6 and RNAPII but is not stably recruited to the CYC1 promoter in vivo.
• Spn1 mutants display synthetic genetic interactions with deletions of histone chaperone genes.

Conclusions:

• Spn1 functions as a histone binding factor with significant histone chaperone activities.
• Chromatin interactions are crucial for Spn1's in vivo functions during transcription.
• Spn1 plays a vital role in regulating chromatin structure and gene expression.