Orchestrating chromosome conformation capture analysis with Bioconductor.

Genome-wide chromatin conformation capture assays provide formidable insights into the spatial organization of genomes. However, due to the complexity of the data structure, their integration in multi-omics workflows remains challenging. We present data structures, computational methods and visualization tools available in Bioconductor to investigate Hi-C, micro-C and other 3C-related data, in R. An online book ( https://bioconductor.org/books/OHCA/ ) further provides prospective end users with a number of workflows to process, import, analyze and visualize any type of chromosome conformation capture data.

Chromosome-scale assemblies of Acanthamoeba castellanii genomes provide insights into Legionella pneumophila infection-related chromatin reorganization.

The unicellular amoeba Acanthamoeba castellanii is ubiquitous in aquatic environments, where it preys on bacteria. The organism also hosts bacterial endosymbionts, some of which are parasitic, including human pathogens such as Chlamydia and Legionella spp. Here we report complete, high-quality genome sequences for two extensively studied A. castellanii strains, Neff and C3. Combining long- and short-read data with Hi-C, we generated near chromosome-level assemblies for both strains with 90% of the genome contained in 29 scaffolds for the Neff strain and 31 for the C3 strain. Comparative genomics revealed strain-specific functional enrichment, most notably genes related to signal transduction in the C3 strain and to viral replication in Neff. Furthermore, we characterized the spatial organization of the A. castellanii genome and showed that it is reorganized during infection by Legionella pneumophila Infection-dependent chromatin loops were found to be enriched in genes for signal transduction and phosphorylation processes. In genomic regions where chromatin organization changed during Legionella infection, we found functional enrichment for genes associated with metabolism, organelle assembly, and cytoskeleton organization. Given Legionella infection is known to alter its host's cell cycle, to exploit the host's organelles, and to modulate the host's metabolism in its favor, these changes in chromatin organization may partly be related to mechanisms of host control during Legionella infection.

Normalization of Chromosome Contact Maps: Matrix Balancing and Visualization.

Over the last decade, genomic proximity ligation approaches have reshaped our vision of chromosomes 3D organizations, from bacteria nucleoids to larger eukaryotic genomes. The different protocols (3Cseq, Hi-C, TCC, MicroC [XL], Hi-CO, etc.) rely on common steps (chemical fixation digestion, ligation…) to detect pairs of genomic positions in close proximity. The most common way to represent these data is a matrix, or contact map, which allows visualizing the different chromatin structures (compartments, loops, etc.) that can be associated to other signals such as transcription, protein occupancy, etc. as well as, in some instances, to biological functions.In this chapter we present and discuss the filtering of the events recovered in proximity ligation experiments as well as the application of the balancing normalization procedure on the resulting contact map. We also describe a computational tool for visualizing normalized contact data dubbed Scalogram.The different processes described here are illustrated and supported by the laboratory custom-made scripts pooled into "hicstuff," an open-access python package accessible on github ( https://github.com/koszullab/hicstuff ).

Computer vision for pattern detection in chromosome contact maps.

Chromosomes of all species studied so far display a variety of higher-order organisational features, such as self-interacting domains or loops. These structures, which are often associated to biological functions, form distinct, visible patterns on genome-wide contact maps generated by chromosome conformation capture approaches such as Hi-C. Here we present Chromosight, an algorithm inspired from computer vision that can detect patterns in contact maps. Chromosight has greater sensitivity than existing methods on synthetic simulated data, while being faster and applicable to any type of genomes, including bacteria, viruses, yeasts and mammals. Our method does not require any prior training dataset and works well with default parameters on data generated with various protocols.

Mapping of Multiple Complementary Sex Determination Loci in a Parasitoid Wasp.

Sex determination has evolved in a variety of ways and can depend on environmental and genetic signals. A widespread form of genetic sex determination is haplodiploidy, where unfertilized, haploid eggs develop into males and fertilized diploid eggs into females. One of the molecular mechanisms underlying haplodiploidy in Hymenoptera, the large insect order comprising ants, bees, and wasps, is complementary sex determination (CSD). In species with CSD, heterozygosity at one or several loci induces female development. Here, we identify the genomic regions putatively underlying multilocus CSD in the parasitoid wasp Lysiphlebus fabarum using restriction-site associated DNA sequencing. By analyzing segregation patterns at polymorphic sites among 331 diploid males and females, we identify up to four CSD candidate regions, all on different chromosomes. None of the candidate regions feature evidence for homology with the csd gene from the honey bee, the only species in which CSD has been characterized, suggesting that CSD in L. fabarum is regulated via a novel molecular mechanism. Moreover, no homology is shared between the candidate loci, in contrast to the idea that multilocus CSD should emerge from duplications of an ancestral single-locus system. Taken together, our results suggest that the molecular mechanisms underlying CSD in Hymenoptera are not conserved between species, raising the question as to whether CSD may have evolved multiple times independently in the group.

cis-Acting Complex-Trait-Associated lincRNA Expression Correlates with Modulation of Chromosomal Architecture.

Intergenic long noncoding RNAs (lincRNAs) are the largest class of transcripts in the human genome. Although many have recently been linked to complex human traits, the underlying mechanisms for most of these transcripts remain undetermined. We investigated the regulatory roles of a high-confidence and reproducible set of 69 trait-relevant lincRNAs (TR-lincRNAs) in human lymphoblastoid cells whose biological relevance is supported by their evolutionary conservation during recent human history and genetic interactions with other trait-associated loci. Their enrichment in enhancer-like chromatin signatures, interactions with nearby trait-relevant protein-coding loci, and preferential location at topologically associated domain (TAD) boundaries provide evidence that TR-lincRNAs likely regulate proximal trait-relevant gene expression in cis by modulating local chromosomal architecture. This is consistent with the positive and significant correlation found between TR-lincRNA abundance and intra-TAD DNA-DNA contacts. Our results provide insights into the molecular mode of action by which TR-lincRNAs contribute to complex human traits.

Evolution and comparative ecology of parthenogenesis in haplodiploid arthropods.

Changes from sexual reproduction to female-producing parthenogenesis (thelytoky) have great evolutionary and ecological consequences, but how many times parthenogenesis evolved in different animal taxa is unknown. We present the first exhaustive database covering 765 cases of parthenogenesis in haplodiploid (arrhenotokous) arthropods, and estimate frequencies of parthenogenesis in different taxonomic groups. We show that the frequency of parthenogenetic lineages extensively varies among groups (0-38% among genera), that many species have both sexual and parthenogenetic lineages and that polyploidy is very rare. Parthenogens are characterized by broad ecological niches: parasitoid and phytophagous parthenogenetic species consistently use more host species, and have larger, polewards extended geographic distributions than their sexual relatives. These differences did not solely evolve after the transition to parthenogenesis. Extant parthenogens often derive from sexual ancestors with relatively broad ecological niches and distributions. As these ecological attributes are associated with large population sizes, our results strongly suggests that transitions to parthenogenesis are more frequent in large sexual populations and/or that the risk of extinction of parthenogens with large population sizes is reduced. The species database presented here provides insights into the maintenance of sex and parthenogenesis in natural populations that are not taxon specific and opens perspectives for future comparative studies.