The Histone Chaperone Spn1 Preserves Chromatin Protections at Promoters and Nucleosome Positioning in Open Reading Frames.

Spn1 is a multifunctional histone chaperone that associates with RNA polymerase II during elongation and is essential for life in eukaryotes. While previous work has elucidated regions of the protein important for its many interactions, it is unknown how these domains contribute to the maintenance of chromatin structure. Here, we employ digestion by micrococcal nuclease followed by single-stranded library preparation and sequencing (MNase-SSP) to characterize chromatin structure in expressing wild-type or mutants of Spn1.

Spn1 and Its Dynamic Interactions with Spt6, Histones and Nucleosomes.

Histone chaperones facilitate the assembly and disassembly of nucleosomes and regulate DNA accessibility for critical cellular processes. Spn1 is an essential, highly conserved histone chaperone that functions in transcription initiation and elongation in a chromatin context. Here we demonstrate that Spn1 binds H3-H4 with low nanomolar affinity, residues 85-99 within the acidic N-terminal region of Spn1 are required for H3-H4 binding, and Spn1 binding to H3-H4 dimers does not impede (H3-H4) tetramer formation.

Genome Instability Is Promoted by the Chromatin-Binding Protein Spn1 in .

Cells expend a large amount of energy to maintain their DNA sequence. DNA repair pathways, cell cycle checkpoint activation, proofreading polymerases, and chromatin structure are ways in which the cell minimizes changes to the genome. During replication, the DNA-damage tolerance pathway allows the replication forks to bypass damage on the template strand.

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

The process of transcriptional elongation by RNA polymerase II (RNAPII) in a chromatin context involves a large number of crucial factors. Spn1 is a highly conserved protein encoded by an essential gene and is known to interact with RNAPII and the histone chaperone Spt6. Spn1 negatively regulates the ability of Spt6 to interact with nucleosomes, but the chromatin binding properties of Spn1 are largely unknown.

DNA-mediated association of two histone-bound complexes of yeast Chromatin Assembly Factor-1 (CAF-1) drives tetrasome assembly in the wake of DNA replication.

Nucleosome assembly in the wake of DNA replication is a key process that regulates cell identity and survival. Chromatin assembly factor 1 (CAF-1) is a H3-H4 histone chaperone that associates with the replisome and orchestrates chromatin assembly following DNA synthesis. Little is known about the mechanism and structure of this key complex.