Teleost fishes, thanks to their rapid evolution of sex determination mechanisms, provide remarkable opportunities to study the formation of sex chromosomes and the mechanisms driving the birth of new ...master sex determining (MSD) genes. However, the evolutionary interplay between the sex chromosomes and the MSD genes they harbor is rather unexplored. We characterized a male-specific duplicate of the anti-Müllerian hormone (amh) as the MSD gene in Northern Pike (Esox lucius), using genomic and expression evidence as well as by loss-of-function and gain-of-function experiments. Using RAD-Sequencing from a family panel, we identified Linkage Group (LG) 24 as the sex chromosome and positioned the sex locus in its sub-telomeric region. Furthermore, we demonstrated that this MSD originated from an ancient duplication of the autosomal amh gene, which was subsequently translocated to LG24. Using sex-specific pooled genome sequencing and a new male genome sequence assembled using Nanopore long reads, we also characterized the differentiation of the X and Y chromosomes, revealing a small male-specific insertion containing the MSD gene and a limited region with reduced recombination. Our study reveals an unexpectedly low level of differentiation between a pair of sex chromosomes harboring an old MSD gene in a wild teleost fish population, and highlights both the pivotal role of genes from the amh pathway in sex determination, as well as the importance of gene duplication as a mechanism driving the turnover of sex chromosomes in this clade.
The NuA4/TIP60 acetyltransferase complex is a key regulator of genome expression and stability. Here we identified MBTD1 as a stable subunit of the complex, and we reveal that, via a histone reader ...domain for H4K20me1/2, MBTD1 allows TIP60 to associate with specific gene promoters and to promote the repair of DNA double-strand breaks by homologous recombination. It was previously suggested that TIP60-dependent acetylation of H4 regulates binding of the non-homologous end joining factor 53BP1, which engages chromatin through simultaneous binding of H4K20me2 and H2AK15ub. We find that the TIP60 complex regulates association of 53BP1 partly by competing for H4K20me2 and by regulating H2AK15ub. Ubiquitylation of H2AK15 by RNF168 inhibits chromatin acetylation by TIP60, while this residue can be acetylated by TIP60 in vivo, blocking its ubiquitylation. Altogether, these results uncover an intricate mechanism orchestrated by the TIP60 complex to regulate 53BP1-dependent repair through competitive bivalent binding and modification of chromatin.
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•We identify MBTD1 as a stable subunit of the TIP60/NuA4 complex•MBTD1 as part of TIP60 regulates transcription and repair of DNA breaks•MBTD1 competes with 53BP1 for binding to the H4K20me mark•TIP60 acetylates H2AK15 and blocks its ubiquitylation by RNF168
Jacquet et al. identify H4K20me-binding MBTD1 as a stable subunit of the TIP60/NuA4 acetyltransferase complex. MBTD1 assists TIP60 in gene-specific transcription and repair of DNA double-strand breaks by homologous recombination. Moreover, TIP60 regulates the bivalent association of 53BP1 to chromatin to influence repair pathway choice.
Roses have high cultural and economic importance as ornamental plants and in the perfume industry. We report the rose whole-genome sequencing and assembly and resequencing of major genotypes that ...contributed to rose domestication. We generated a homozygous genotype from a heterozygous diploid modern rose progenitor, Rosa chinensis 'Old Blush'. Using single-molecule real-time sequencing and a meta-assembly approach, we obtained one of the most comprehensive plant genomes to date. Diversity analyses highlighted the mosaic origin of 'La France', one of the first hybrids combining the growth vigor of European species and the recurrent blooming of Chinese species. Genomic segments of Chinese ancestry identified new candidate genes for recurrent blooming. Reconstructing regulatory and secondary metabolism pathways allowed us to propose a model of interconnected regulation of scent and flower color. This genome provides a foundation for understanding the mechanisms governing rose traits and should accelerate improvement in roses, Rosaceae and ornamentals.
Although originally primarily a system for functional biology, Arabidopsis thaliana has, owing to its broad geographical distribution and adaptation to diverse environments, developed into a powerful ...model in population genomics. Here we present chromosome-level genome assemblies of 69 accessions from a global species range. We found that genomic colinearity is very conserved, even among geographically and genetically distant accessions. Along chromosome arms, megabase-scale rearrangements are rare and typically present only in a single accession. This indicates that the karyotype is quasi-fixed and that rearrangements in chromosome arms are counter-selected. Centromeric regions display higher structural dynamics, and divergences in core centromeres account for most of the genome size variations. Pan-genome analyses uncovered 32,986 distinct gene families, 60% being present in all accessions and 40% appearing to be dispensable, including 18% private to a single accession, indicating unexplored genic diversity. These 69 new Arabidopsis thaliana genome assemblies will empower future genetic research.
Conventional affinity purification followed by mass spectrometry (AP-MS) analysis is a broadly applicable method used to decipher molecular interaction networks and infer protein function. However, ...it is sensitive to perturbations induced by ectopically overexpressed target proteins and does not reflect multilevel physiological regulation in response to diverse stimuli. Here, we developed an interface between genome editing and proteomics to isolate native protein complexes produced from their natural genomic contexts. We used CRISPR/Cas9 and TAL effector nucleases (TALENs) to tag endogenous genes and purified several DNA repair and chromatin-modifying holoenzymes to near homogeneity. We uncovered subunits and interactions among well-characterized complexes and report the isolation of MCM8/9, highlighting the efficiency and robustness of the approach. These methods improve and simplify both small- and large-scale explorations of protein interactions as well as the study of biochemical activities and structure-function relationships.
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•Streamlined affinity purification of protein complexes using CRISPR/Cas9 and TALENs•Purification of native holoenzymes regulated under physiological conditions•Blueprint for the systematic and unbiased mapping of the human protein interactome
Characterization of multiprotein complexes via affinity purification is optimal when expression of the bait protein is specified by its natural genomic context. Dalvai et al. implement a robust genome-editing-based approach to tag endogenous genes and purify native complexes from human cells to near homogeneity.
Advances in deciphering the functional architecture of eukaryotic genomes have been facilitated by recent breakthroughs in sequencing technologies, enabling a more comprehensive representation of ...genes and repeat elements in genome sequence assemblies, as well as more sensitive and tissue-specific analyses of gene expression. Here we show that PacBio sequencing has led to a substantially improved genome assembly of Medicago truncatula A17, a legume model species notable for endosymbiosis studies
, and has enabled the identification of genome rearrangements between genotypes at a near-base-pair resolution. Annotation of the new M. truncatula genome sequence has allowed for a thorough analysis of transposable elements and their dynamics, as well as the identification of new players involved in symbiotic nodule development, in particular 1,037 upregulated long non-coding RNAs (lncRNAs). We have also discovered that a substantial proportion (~35% and 38%, respectively) of the genes upregulated in nodules or expressed in the nodule differentiation zone colocalize in genomic clusters (270 and 211, respectively), here termed symbiotic islands. These islands contain numerous expressed lncRNA genes and display differentially both DNA methylation and histone marks. Epigenetic regulations and lncRNAs are therefore attractive candidate elements for the orchestration of symbiotic gene expression in the M. truncatula genome.
Abstract
Motivation
Long-read sequencing technologies can be employed to detect and map DNA modifications at the nucleotide resolution on a genome-wide scale. However, published software packages ...neglect the integration of genomic annotation and comprehensive filtering when analyzing patterns of modified bases detected using Pacific Biosciences (PacBio) or Oxford Nanopore Technologies (ONT) data. Here, we present DNA Modification Annotation (DNAModAnnot), a R package designed for the global analysis of DNA modification patterns using adapted filtering and visualization tools.
Results
We tested our package using PacBio sequencing data to analyze patterns of the 6-methyladenine (6mA) in the ciliate Paramecium tetraurelia, in which high 6mA amounts were previously reported. We found P. tetraurelia 6mA genome-wide distribution to be similar to other ciliates. We also performed 5-methylcytosine (5mC) analysis in human lymphoblastoid cells using ONT data and confirmed previously known patterns of 5mC. DNAModAnnot provides a toolbox for the genome-wide analysis of different DNA modifications using PacBio and ONT long-read sequencing data.
Availability and implementation
DNAModAnnot is distributed as a R package available via GitHub (https://github.com/AlexisHardy/DNAModAnnot).
Supplementary information
Supplementary data are available at Bioinformatics online.
During DNA replication, thousands of replication origins are activated across the genome. Chromatin architecture contributes to origin specification and usage, yet it remains unclear which chromatin ...features impact on DNA replication. Here, we perform a RNAi screen for chromatin regulators implicated in replication control by measuring RPA accumulation upon replication stress. We identify six factors required for normal rates of DNA replication and characterize a function of the bromodomain and PHD finger‐containing protein 3 (BRPF3) in replication initiation. BRPF3 forms a complex with HBO1 that specifically acetylates histone H3K14, and genomewide analysis shows high enrichment of BRPF3, HBO1 and H3K14ac at ORC1‐binding sites and replication origins found in the vicinity of TSSs. Consistent with this, BRPF3 is necessary for H3K14ac at selected origins and efficient origin activation. CDC45 recruitment, but not MCM2‐7 loading, is impaired in BRPF3‐depleted cells, identifying a BRPF3‐dependent function of HBO1 in origin activation that is complementary to its role in licencing. We thus propose that BRPF3‐HBO1 acetylation of histone H3K14 around TSS facilitates efficient activation of nearby replication origins.
Synopsis
A distinct origin activation role of HBO1 acetyltransferase provides new insight into how demarcation of chromatin surrounding transcription start sites affects the regulation of nearby replication origins.
siRNA screen identifies chromatin regulators important for DNA replication, including the scaffold protein BRPF3.
BRPF3 regulates origin firing by directing the acetyltransferase HBO1 to target histone H3K14 in chromatin surrounding replication origins.
HBO1‐BRPF3 complex function in origin activation is separate from and complementary to HBO1‐JADE1 function in origin licensing.
Reduced origin activation upon BRPF3 depletion protects cells against replication stress‐induced DNA damage.
An RNAi screen for chromatin regulators of replication control reveals an origin activation role of HBO1 acetyltransferase role that is separate from its function in origin licensing.
Grapevine is one of the most important fruit species in the world. In order to better understand genetic basis of traits variation and facilitate the breeding of new genotypes, we sequenced, ...assembled, and annotated the genome of the American native Vitis riparia, one of the main species used worldwide for rootstock and scion breeding. A total of 164 Gb raw DNA reads were obtained from Vitis riparia resulting in a 225X depth of coverage. We generated a genome assembly of the V. riparia grape de novo using the PacBio long-reads that was phased with the 10x Genomics Chromium linked-reads. At the chromosome level, a 500 Mb genome was generated with a scaffold N50 size of 1 Mb. More than 34% of the whole genome were identified as repeat sequences, and 37,207 protein-coding genes were predicted. This genome assembly sets the stage for comparative genomic analysis of the diversification and adaptation of grapevine and will provide a solid resource for further genetic analysis and breeding of this economically important species.
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