The histone code of the fungal genus Aspergillus uncovered by evolutionary and proteomic analyses

Abstract

Chemical modifications of DNA and histone proteins impact the organization of chromatin within the nucleus. Changes in these modifications, catalyzed by different chromatin-modifying enzymes, influence chromatin organization, which in turn is thought to impact the spatial and temporal regulation of gene expression. While combinations of different histone modifications, the histone code, have been studied in several model species, we know very little about histone modifications in the fungal genus Aspergillus, whose members are generally well-studied due to their importance as models in cell and molecular biology as well as their medical and biotechnological relevance. Here, we used phylogenetic analyses in 94 Aspergilli as well as other fungi to uncover the occurrence and evolutionary trajectories of enzymes and protein complexes with roles in chromatin modifications or regulation. We found that these enzymes and complexes are highly conserved in Aspergilli, pointing towards a complex repertoire of chromatin modifications. Nevertheless, we also observed few recent gene duplications or losses, highlighting Aspergillus species to further study the roles of specific chromatin modifications. SET7 (KMT6) and other components of PRC2 (Polycomb Repressive Complex 2), which is responsible for methylation on histone H3 at lysine 27 in many eukaryotes including fungi, are absent in Aspergilli as well as in closely related Penicillium species, suggesting that these lost the capacity for this histone modification. We corroborated our computational predictions by performing untargeted mass spectrometry analysis of histone post-translational modifications in Aspergillus nidulans. This systematic analysis will pave the way for future research into the complexity of the histone code and its functional implications on genome architecture and gene regulation in fungi. Data SummaryThe predicted proteomes used in this study are publicly available at the JGI (Joint Genome Institute) MycoCosm repository (1); the species names and abbreviations are listed in Supplementary Table 1. To evaluate the completeness of the predicted proteomes and to obtain a species phylogeny, 758 fungal BUSCO (Benchmarking Universal Single-Copy Ortholog) genes were used, and their names are listed in Supplementary Table 2. The fasta, trimmed alignment, and maximum-likelihood phylogenetic tree files can be found in Supplementary Data 1 and 2 deposited at Zenodo (10.5281/zenodo.6586562). The mass spectrometry results mentioned in Supplementary Table 6 are deposited in the PRIDE database with the dataset identifier PXD033478. Impact StatementUncovering how fungi regulate gene expression and genome organization is an important step to understand how they control biological processes such as growth or biosynthesis of bioactive molecules. Despite the known importance of chromatin modifications in controlling a plethora of biological processes across eukaryotes, they remain particularly poorly understood in filamentous fungi, even in model organisms like Aspergilli. Our systematic evolutionary analysis provides a robust framework for the functional analysis of chromatin modifications in Aspergillus species and other fungi. Our results do not only implicate candidate enzymes to play a role in new chromatin modifications, but they also point at species that have experienced duplications or losses of genes encoding enzymes for specific chromatin modifications. We therefore expect that this work will set the stage for future research into the complexity of the histone code and its functional implications on gene regulation and genome organization in fungi.