THE HISTONE CHAPERONE SPN1 PRESERVES SUBNUCLEOSOMAL STRUCTURES AT PROMOTERS AND NUCLEOSOME POSITIONING IN OPEN READING FRAMES

Spn1 is a multifunctional histone chaperone 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 yeast expressing wild-type or mutants of Spn1. We mapped nucleosome and subnucleosomal protections genome-wide, and surprisingly, we observed a genome-wide loss of subnucleosomal protection over nucleosome-depleted regions (NDRs) in the Spn1-K192N-containing strain, indicating critical functions of Spn1 in maintaining normal chromatin architecture in promoter regions. Additionally, alterations in nucleosome and hexasome positioning were observed in markedly different mutant Spn1 strains, demonstrating that multiple functions of Spn1 are required to maintain proper chromatin structure in open reading frames. Moreover, the impact of these functions correlated with gene expression and length. Taken together, our results reveal a previously unknown role of Spn1 in the maintenance of NDR architecture and deepen our understanding of Spn1-dependent chromatin maintenance over transcribed regions. Overall design: MNase-SSP of following yeast strains: WT, spn1 K192N, aaand spn1 141-305 at 32U and 128U of MNase. Spike-in normalized RNA-seq of following yeast strains: WT, spn1 K192N, aaand spn1 141-305