Destabilization of chromosome structure by histone H3 lysine 27 methylation

Abstract

27Chromosome and genome stability are important for normal cell function as instability often 28 correlates with disease and dysfunction of DNA repair mechanisms. Many organisms maintain 29 supernumerary or accessory chromosomes that deviate from standard chromosomes. The 30 pathogenic fungus Zymoseptoria tritici has as many as eight accessory chromosomes, which are 31 highly unstable during meiosis and mitosis, transcriptionally repressed, show enrichment of 32 repetitive elements, and enrichment with heterochromatic histone methylation marks, e.g., 33trimethylation of H3 lysine 9 or lysine 27 (H3K9me3, H3K27me3). To elucidate the role of 34 heterochromatin on genome stability in Z. tritici, we deleted the genes encoding the 35 methyltransferases responsible for H3K9me3 and H3K27me3, kmt1 and kmt6, respectively, and 36 generated a double mutant. We combined experimental evolution and genomic analyses to 37 determine the impact of these deletions on chromosome and genome stability, both in vitro and 38 in planta. We used whole genome sequencing, ChIP-seq, and RNA-seq to compare changes in 39 genome and chromatin structure, and differences in gene expression between mutant and 40 wildtype strains. Analyses of genome and ChIP-seq data in H3K9me3-deficient strains revealed 41 dramatic chromatin reorganization, where H3K27me3 is mostly relocalized into regions that are 42 enriched with H3K9me3 in wild type. Many genome rearrangements and formation of new 43 chromosomes were found in the absence of H3K9me3, accompanied by activation of transposable 44 elements. In stark contrast, loss of H3K27me3 actually increased the stability of accessory 45 chromosomes under normal growth conditions in vitro, even without large scale changes in gene 46 activity. We conclude that H3K9me3 is important for the maintenance of genome stability 47 because it disallows H3K27me3 in these regions. In this system, H3K27me3 reduces the overall 48 stability of accessory chromosomes, generating a "metastable" state for these quasi-essential 49 regions of the genome. 50 3 Author Summary 51 Genome and chromosome stability are essential to maintain normal cell function and viability. 52However, differences in genome and chromosome structure are frequently found in organisms 53 that undergo rapid adaptation to changing environmental conditions, and in humans are often 54 found in cancer cells. We study genome instability in a fungal pathogen that exhibits a high degree 55 of genetic diversity. Regions that show extraordinary diversity in this pathogen are the 56 transposon-rich accessory chromosomes, which contain few genes that are of unknown benefit 57 to the organism but maintained in the population and thus considered "quasi essential". 58Accessory chromosomes in all fungi studied so far are enriched with markers for 59 heterochromatin, namely trimethylation of H3 lysine 9 and 27 (H3K9me3, H3K27me3). We show 60 that loss of these heterochromatin marks has strong but opposing effects on genome stability. 61While loss of the transposon-associate...

Loss of Polycomb Protein EZH2 causes major depletion of H3K27 and H3K9 tri-methylation and developmental defects in the fungusPodospora anserina

“…In N. crassa, disruption of H3K27me3 methyltransferase activity did not lead to developmental defects (Jamieson et al 2013) but to abnormal chromosome conformation (Klocko et al 2016). By contrast, in the pathogenic fungus Zymoseptoria tritici absence of H3K27me3 reduces the loss of its accessory chromosomes, whereas loss of H3K9me3 induced many genomic rearrangements (Möller et al, 2019). Moreover, in N. crassa and Z. tritici, depletion of H3K9me3 resulted in massive redistribution of H3K27me3 in genomic compartments that are embedded in constitutive H3K9me3 heterochromatin in their respective wild-type backgrounds (Jamieson et al, 2016, Möller et al, 2019.…”

“…By contrast, in the pathogenic fungus Zymoseptoria tritici absence of H3K27me3 reduces the loss of its accessory chromosomes, whereas loss of H3K9me3 induced many genomic rearrangements (Möller et al, 2019). Moreover, in N. crassa and Z. tritici, depletion of H3K9me3 resulted in massive redistribution of H3K27me3 in genomic compartments that are embedded in constitutive H3K9me3 heterochromatin in their respective wild-type backgrounds (Jamieson et al, 2016, Möller et al, 2019. In contrast, the absence of H3K27me3 machinery did not impair the H3K9me3 genomic localization in either of them.…”

Genome compartmentalization predates species divergence in the plant pathogen genus Zymoseptoria

“…We used previously published RNA sequencing data to increase the quality of the gene prediction and combined three distinct methodologies [22,30,32]. As a rst approach, we used GeneMark-ES for an ab initio prediction using the option "--fungus" [50].…”

“…To update expression pro les on the new genome assemblies and new gene predictions of the three Z. tritici isolates IPO323, Zt05 and Zt10, we used previously generated RNA-seq data from in planta and in vitro growth [30,32]. The in planta RNA-seq data was obtained from infected leaves at four different stages corresponding to early and late biotrophic and necrotrophic stages of the three Z. tritici isolates [30].…”

“…Interestingly, the accessory chromosomes in Z. tritici show a low transcriptional activity in vitro as well as in planta [30,31]. This suppression of gene expression correlates with an enrichment of heterochromatin associated with the histone modi cation H3K27me3 on the accessory chromosomes [6,32].…”