Nuclear Envelope Integrity in Health and Disease: Consequences on Genome Instability and Inflammation

The dynamic nature of the nuclear envelope (NE) is often underestimated. The NE protects, regulates, and organizes the eukaryote genome and adapts to epigenetic changes and to its environment. The NE morphology is characterized by a wide range of diversity and abnormality such as invagination and blebbing, and it is a diagnostic factor for pathologies such as cancer. Recently, the micronuclei, a small nucleus that contains a full chromosome or a fragment thereof, has gained much attention. The NE of micronuclei is prone to collapse, leading to DNA release into the cytoplasm with consequences ranging from the activation of the cGAS/STING pathway, an innate immune response, to the creation of chromosomal instability. The discovery of those mechanisms has revolutionized the understanding of some inflammation-related diseases and the origin of complex chromosomal rearrangements, as observed during the initiation of tumorigenesis. Herein, we will highlight the complexity of the NE biology and discuss the clinical symptoms observed in NE-related diseases. The interplay between innate immunity, genomic instability, and nuclear envelope leakage could be a major focus in future years to explain a wide range of diseases and could lead to new classes of therapeutics.

Overcoming uncollapsed haplotypes in long-read assemblies of non-model organisms

Background Long-read sequencing is revolutionizing genome assembly: as PacBio and Nanopore technologies become more accessible in technicity and in cost, long-read assemblers flourish and are starting to deliver chromosome-level assemblies. However, these long reads are usually error-prone, making the generation of a haploid reference out of a diploid genome a difficult enterprise. Failure to properly collapse haplotypes results in fragmented and structurally incorrect assemblies and wreaks havoc on orthology inference pipelines, yet this serious issue is rarely acknowledged and dealt with in genomic projects, and an independent, comparative benchmark of the capacity of assemblers and post-processing tools to properly collapse or purge haplotypes is still lacking. Results We tested different assembly strategies on the genome of the rotifer Adineta vaga, a non-model organism for which high coverages of both PacBio and Nanopore reads were available. The assemblers we tested (Canu, Flye, NextDenovo, Ra, Raven, Shasta and wtdbg2) exhibited strikingly different behaviors when dealing with highly heterozygous regions, resulting in variable amounts of uncollapsed haplotypes. Filtering reads generally improved haploid assemblies, and we also benchmarked three post-processing tools aimed at detecting and purging uncollapsed haplotypes in long-read assemblies: HaploMerger2, purge_haplotigs and purge_dups. Conclusions We provide a thorough evaluation of popular assemblers on a non-model eukaryote genome with variable levels of heterozygosity. Our study highlights several strategies using pre and post-processing approaches to generate haploid assemblies with high continuity and completeness. This benchmark will help users to improve haploid assemblies of non-model organisms, and evaluate the quality of their own assemblies.

Humanizing the yeast origin recognition complex

The Origin Recognition Complex (ORC) is an evolutionarily conserved six-subunit protein complex that binds specific sites at many locations to coordinately replicate the entire eukaryote genome. Though highly conserved in structure, ORC’s selectivity for replication origins has diverged tremendously between yeasts and humans to adapt to vastly different life cycles. In this work, we demonstrate that the selectivity determinant of ORC for DNA binding lies in a 19-amino acid insertion helix in the Orc4 subunit, which is present in yeast but absent in human. Removal of this motif from Orc4 transforms the yeast ORC, which selects origins based on base-specific binding at defined locations, into one whose selectivity is dictated by chromatin landscape and afforded with plasticity, as reported for human. Notably, the altered yeast ORC has acquired an affinity for regions near transcriptional start sites (TSSs), which the human ORC also favors.

Bacterial Genes Outnumber Archaeal Genes in Eukaryotic Genomes

Eukaryotes are typically depicted as descendants of archaea, but their genomes are evolutionary chimeras with genes stemming from archaea and bacteria. Which prokaryotic heritage predominates? Here, we have clustered 19,050,992 protein sequences from 5,443 bacteria and 212 archaea with 3,420,731 protein sequences from 150 eukaryotes spanning six eukaryotic supergroups. By downsampling, we obtain estimates for the bacterial and archaeal proportions. Eukaryotic genomes possess a bacterial majority of genes. On average, the majority of bacterial genes is 56% overall, 53% in eukaryotes that never possessed plastids, and 61% in photosynthetic eukaryotic lineages, where the cyanobacterial ancestor of plastids contributed additional genes to the eukaryotic lineage. Intracellular parasites, which undergo reductive evolution in adaptation to the nutrient rich environment of the cells that they infect, relinquish bacterial genes for metabolic processes. Such adaptive gene loss is most pronounced in the human parasite Encephalitozoon intestinalis with 86% archaeal and 14% bacterial derived genes. The most bacterial eukaryote genome sampled is rice, with 67% bacterial and 33% archaeal genes. The functional dichotomy, initially described for yeast, of archaeal genes being involved in genetic information processing and bacterial genes being involved in metabolic processes is conserved across all eukaryotic supergroups.

Estimating the quality of eukaryotic genomes recovered from metagenomic analysis

Eukaryotes make up a large fraction of microbial biodiversity. However, the field of metagenomics has been heavily biased towards the study of just the prokaryotic fraction. This focus has driven the necessary methodological developments to enable the recovery of prokaryotic genomes from metagenomes, which has reliably yielded genomes from thousands of novel species. More recently, microbial eukaryotes have gained more attention, but there is yet to be a parallel explosion in the number of eukaryotic genomes recovered from metagenomic samples. One of the current deficiencies is the lack of a universally applicable and reliable tool for the estimation of eukaryote genome quality. To address this need, we have developed EukCC, a tool for estimating the quality of eukaryotic genomes based on the dynamic selection of single copy marker gene sets, with the aim of applying it to metagenomics datasets. We demonstrate that our method outperforms current genome quality estimators and have applied EukCC to datasets from two different biomes to enable the identification of novel genomes, including a eukaryote found on the human skin and a Bathycoccus species obtained from a marine sample.

Structure of SWI/SNF chromatin remodeller RSC bound to a nucleosome

Chromatin remodelling complexes of the SWI/SNF family function in the formation of nucleosome-depleted regions and transcriptionally active promoters in the eukaryote genome. The structure of the Saccharomyces cerevisiae SWI/SNF family member RSC in complex with a nucleosome substrate reveals five protein modules and suggests key features of the remodelling mechanism. A DNA-interacting module grasps extra-nucleosomal DNA and helps to recruit RSC to promoters. The ATPase and arm modules sandwich the nucleosome disc with their ‘SnAC’ and ‘finger’ elements, respectively. The translocase motor engages with the edge of the nucleosome at superhelical location +2 to pump DNA along the nucleosome, resulting in a sliding of the histone octamer along DNA. The results elucidate how nucleosome-depleted regions are formed and provide a basis for understanding human chromatin remodelling complexes of the SWI/SNF family and the consequences of cancer mutations that frequently occur in these complexes.

Genomes and the origin of genetic variation

Error and chance events, random mutations, are necessary prerequisites for evolution to happen. In a perfect world with no mutations there would be no evolution because no genetic variation would be generated that natural selection or genetic drift could work upon. This chapter first reviews how DNA is organized into genomes and genes in bacteria, archaea, and, in greater detail, eukaryotes. A surprising finding is that only a small fraction of the eukaryote genome consists of coding sequence. Evolutionary processes that can explain the presence of large amounts of noncoding DNA and the repetitive structure of the genome are reviewed, with emphasis on the roles that selfish genetic elements and unequal crossing over play. The chapter further explores the mechanisms that cause mutation and how new genes and protein functions originate.

Mapping of the Retrotransposable Elements Rex1 and Rex3 in Chromosomes of Eigenmannia (Teleostei, Gymnotiformes, Sternopygidae)

Transposable elements constitute a remarkable fraction of the eukaryote genome and show particular capacity to move and insert in specific regions of the genome. This study identified the retrotransposable elements Rex1 and Rex3 in the genomes of 6 cytotypes of Eigenmannia. The sequences were isolated by PCR, sequenced and physically mapped in the chromosomes of these cytotypes, aiming to investigate the organization and distribution of these elements in this fish group, mainly in the sex chromosomes. The FISH physical mapping revealed that both Rex1 and Rex3 elements are dispersed in small clusters throughout the chromosomes of all cytotypes analyzed. However, conspicuous blocks occur in several samples, including an accentuated accumulation of the Rex3 element in X1 and X2 chromosomes of Eigenmannia sp. 2 and in the X chromosome of E. virescens. The accumulations are coincident with heterochromatin-rich regions, suggesting that Rex3 played a role in the differentiation process of the sex chromosomes.