Long transposon-rich centromeres in an oomycete reveal divergence of centromere features in Stramenopila-Alveolata-Rhizaria lineages

Abstract

26Centromeres are chromosomal regions that serve as platforms for kinetochore assembly 27 and spindle attachments, ensuring accurate chromosome segregation during cell division. 28Despite functional conservation, centromere DNA sequences are diverse and often repetitive, 29 making them challenging to assemble and identify. Here, we describe centromeres in an 30 oomycete Phytophthora sojae by combining long-read sequencing-based genome assembly and 31 chromatin immunoprecipitation for the centromeric histone CENP-A followed by high-throughput 32 sequencing (ChIP-seq). P. sojae centromeres cluster at a single focus at different life stages and 33 during nuclear division. We report an improved genome assembly of the P. sojae reference strain, 34 which enabled identification of 15 enriched CENP-A binding regions as putative centromeres. 35By focusing on a subset of these regions, we demonstrate that centromeres in P. sojae are 36 regional, spanning 211 to 356 kb. Most of these regions are transposon-rich, poorly transcribed, 37 and lack the histone modification H3K4me2 but are embedded within regions with the 38 heterochromatin marks H3K9me3 and H3K27me3. Strikingly, we discovered a Copia-like 39 transposon (CoLT) that is highly enriched in the CENP-A chromatin. Similar clustered elements 40 are also found in oomycete relatives of P. sojae, and may be applied as a criterion for prediction 41 of oomycete centromeres. This work reveals a divergence of centromere features in oomycetes 42 as compared to other organisms in the Stramenopila-Alveolata-Rhizaria (SAR) supergroup 43 including diatoms and Plasmodium falciparum that have relatively short and simple regional 44 centromeres. Identification of P. sojae centromeres in turn also augments the genome assembly. 45 46 Significance Statement 49Oomycetes are fungal-like microorganisms that belong to the stramenopiles within the 50 Stramenopila-Alveolata-Rhizaria (SAR) supergroup. The Phytophthora oomycetes are infamous 51 as plant killers, threatening crop production worldwide. Because of the highly repetitive nature of 52 their genomes, assembly of oomycete genomes presents challenges that impede identification of 53 centromeres, which are chromosomal sites mediating faithful chromosome segregation. We 54 report long-read sequencing-based genome assembly of the Phytophthora sojae reference strain, 55 which facilitated the discovery of centromeres. P. sojae harbors large regional centromeres 56 enriched for a Copia-like transposon that is also found in discrete clusters in other oomycetes. 57This study provides insight into the oomycete genome organization, and broadens our knowledge 58 of the centromere structure, function and evolution in eukaryotes. 59 60 Accurate segregation of chromosomes during mitosis and meiosis is critical for the 61 development and reproduction of all eukaryotic organisms. Centromeres are specialized regions 62 of chromosomes that mediate kinetochore formation, spindle attachment, and sister chromatid 63 segregation during cell division (1, 2). T...

Epigenetic dynamics of centromeres and neocentromeres inCryptococcus deuterogattii

“…This feature is also observed in several other fungi, including Neurospora crassa and Fusarium fujikuroi, in which centromeres are enriched for H3K9me2/3 [44,45]. Similar observations have been made in the oomycete Phytophthora sojae [46]. This is in contrast to S. pombe, in which the CENP-A-enriched regions lack heterochromatin, but pericentric regions are enriched for heterochromatin [19,38,39].…”

Centromere evolution in the fungal genusVerticillium

“…Centromeres are often enriched for a variety of different retrotransposons and other repetitive elements (15,16,25,29,30). We similarly observed that centromeres in all Verticillium species are repeat-rich.…”

“…H3K4me2 borders most centromeres in Z. tritici (27), and is associated with centromeres in S. pombe and some animals and plants (64-67). H3K4me2 has not been observed at centromeres in most fungi, including V. dahliae, and in the oomycete P. sojae (30). Changes in heterochromatin in N. crassa leads to altered CenH3 positioning (25), suggesting that heterochromatin is similarly required for centromere maintenance and function in V. dahliae.…”

“…Knowledge on centromeres has been impaired by their repetitive nature, which hampers their assembly and subsequent analyses (15,28). However, recent advances in long-read sequencing technologies enables to study the constitution and evolution of centromeres (11,16,(29)(30)(31). By using long-read sequencing technologies in combination with optical mapping, we previously generated gapless genome assemblies of two strains of the fungal plant pathogen Verticillium dahliae (32), whose genomes are characterized by genome rearrangements and the occurrence of lineage-specific (LS) regions (7,8,(33)(34)(35)) that are hypervariable between V. dahliae strains and contain genes with roles in adaptive evolution to plant hosts (7,8,33,35).…”

“…where centromeres contain six retrotransposons (Tcn1-6) that nearly exclusively occur at centromeres(16). Similarly, centromeres of the oomycete plant pathogen Phytophthora sojae contain multiple types of repeats, but they are enriched for a single element called CoLT (Copia-Like Transposon)(30). The strong associations of specific repeats to centromeres could directly or indirectly link these elements to centromere function.…”