Friday, April 19, 2024
4/19/2024
PROJECT SUMMARY Histone proteins package DNA into chromatin and regulate all DNA-templated biological processes in eukaryotes. Consistent with their essential function, mutations or misregulation of histones result in many diseases. While core histones primarily function in genome packaging, histone variants can replace canonical histones at unique genomic locations for specialized roles such as DNA damage response (DDR) or gene expression. Despite their essential functions, histone repertoires have undergone distinct lineage-specific changes. Such evolutionary novelty via gene fusions, duplications or sequence divergence is unexpected in conserved protein families and suggestive of an adaptive advantage for sequence innovation. This proposal will use evolutionary innovations in eukaryotic histone H2A repertoires to investigate the causes and consequences of evolutionary turnover of histone proteins. The most common eukaryotic H2A repertoire is made up of core histone H2A, and histone variants H2A.X and H2A.Z that are involved in DDR and gene regulation, respectively. However, two evolutionary shifts, both likely selectively advantageous, have occurred in yeast and Drosophila H2A repertoires. In yeast, H2A.X, entirely replaced canonical H2A, likely improving DDR and affecting processes like meiosis that depend on DDR. In Drosophila, H2A.X fused with H2A.Z giving rise to a unique H2Av variant. This fusion enriches DDR at heterochromatin which potentially restricts DDR- based transposition events to gene poor regions. To identify functional consequences of these evolutionary innovations, the applicant will re-engineer the ancestral eukaryotic H2A repertoire in yeast and flies. Specifically in S. cerevisiae, the applicant will engineer a core H2A and two variants H2A.X and H2A.Z, preventing H2A.X from being the core histone (Aim 1). In D. melanogaster, the fusion histone H2Av will be separated into H2A.X and H2A.Z uncoupling DDR and gene regulation functions (Aim 2). Changes to organismal chromatin packaging, and relevant biological functions including DNA repair, meiosis, fertility, and transposition will be interrogated. In Aim 3, the applicant will apply tools learnt in Aims 1 and 2 to study divergence of eukaryotic core H2A. Histones have tail sequences which are heavily post-translationally modified and play crucial roles for higher order chromatin packaging and protein interactions. The high sequence divergence across eukaryotic histone tails suggests that tails could facilitate unique lineage-specific functions. By engineering different tail sequences in yeast and flies, Aim 3 will reveal changes to chromatin packaging, and changes to processes such as mating, and quiescence in yeast, and fertility and development in flies. To launch this work, the applicant requires training in genetics, genomics, and phenotypic assays in two model organisms, yeast and flies. By leveraging, her expertise in evolutionary analyses and the power of well-established tools in yeast and flies simultaneously, in the future the applicant will study the biological basis and consequences of histone innovation including her own previous discoveries and co-evolving mechanisms.
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