The role of the 3D genome organisation in shaping mammalian chromosome evolution

Abstract

Mammalian chromosome evolution is highly diverse since a notable variability in chromosome sizes and diploid numbers has been reported. The hierarchical 3D genome organisation (i.e., the way chromosomes are folded within the nucleus) adds another layer of complexity in the field of chromosome evolution. In this context, the study of the similarities and differences of chromosome folding and genomic interactions across species is key to understand the dynamics of genome function and speciation.

This thesis explores the dynamics of 3D chromosome folding across mammalian species representative of key phylogroups. In this context, we set two specific objectives: i) determine principles of chromatin folding, sequence conservation and evolutionary chromosomal reorganisations in mammalian somatic cells; and ii) analyse the role of chromatin remodelling in the generation of chromosomal reorganisations during the formation of germ cells. To achieve this, we employed a comprehensive integrative computational analysis, combining next generation sequencing technologies, including Hi-C, ChIP-seq and RNA-seq, with an integrative bioinformatic analysis. We generated novel Hi-C and ChIP-seq data from somatic cell lines of non-model species (i.e., Tasmanian devil, African elephant and aardvark) and compared them with available sequencing data from representative mammalian species of major phylogroups. Moreover, we integrated structural and functional data from germ cells to generate a model that integrates the role of meiosis in genome evolutionary plasticity.

Our results indicate that the 3D genome hierarchical organisation is linked to mammalian chromosomal evolution. Within mammalian species we detected two distinctive nuclear chromosomal configurations in somatic cells: i) highly compacted medium size eutherian chromosomes are organised into chromosomal territories; whereas ii) loose long marsupial chromosomes are organised in a Rabl-like configuration characterised by centromere clustering. Moreover, we detected that this variability in chromosome folding was associated with a decrease of CTCF density and DNA loop lengthening in species with long chromosomes. Finally, we showed that different chromosomal organisations do not alter TADs, confirming the essential nature of TADs for mammalian evolution. Regardless of TADs conservation, we detected contrasting patterns of chromosome evolution associated with chromosome occupancy and folding. The reconstruction of the ancestral karyotypes of marsupial and eutherian mammals (i.e., afrotherians and rodents) revealed lineage-specic evolutionary trends. While the Tasmanian devil (2n=14) exhibits only inversions from the marsupial ancestor (2n=14); afrotherian and rodent species present high complex chromosomal rearrangements (CRs) (fissions, fusions, translocations and inversion), when compared to their respective ancestral karyotypes (2n=48 for afrotheria and 2n=52 for rodentia). Moreover, we detected that in all phylogroups analysed inversions presented a significant increase of intra-chromosomal interactions and CTCF density when compared to colinear regions, suggesting that inversions can act as structural island of diversity where species specific traits evolve.

Our findings also highlight the relevance of interpreting CRs in the context of the formation of the germ line and the meiotic process. We investigated how epigenetic features and the 3D chromatin remodelling that takes place during mouse spermatogenesis influences the formation and transmission of evolutionary CRs. The reconstruction of the ancestral rodentia genome allowed us to characterise the genomic and epigenetic features of mouse-specific evolutionary breakpoint regions (EBRs) in three germ cell types: spermatogonia (pre-meiosis); primary spermatocytes (meiosis); and round spermatids (post-meiosis). We detected that EBRs showed distinctive patterns: i) they do not disrupt meiosis-essential genes; ii) they are positively associated with genomic regions that become accessible during spermatogenesis; iii) they are located at TAD boundaries in pre-meiotic and meiotic cells, but within TADs in post-meiotic cells; and iv) they co-localised with long-range spermatid-specific interactions of reproductive relevant genes that recapitulate ancestral chromosomal interactions. Overall, our results suggest that CR formation is dependent of chromatin and epigenetic remodelling in the germ line.

Mammalian chromosome evolution is highly diverse since a notable variability in chromosome sizes and diploid numbers has been reported. The hierarchical 3D genome organisation (i.e., the way chromosomes are folded within the nucleus) adds another layer of complexity in the field of chromosome evolution. In this context, the study of the similarities and differences of chromosome folding and genomic interactions across species is key to understand the dynamics of genome function and speciation.

This thesis explores the dynamics of 3D chromosome folding across mammalian species representative of key phylogroups. In this context, we set two specific objectives: i) determine principles of chromatin folding, sequence conservation and evolutionary chromosomal reorganisations in mammalian somatic cells; and ii) analyse the role of chromatin remodelling in the generation of chromosomal reorganisations during the formation of germ cells. To achieve this, we employed a comprehensive integrative computational analysis, combining next generation sequencing technologies, including Hi-C, ChIP-seq and RNA-seq, with an integrative bioinformatic analysis. We generated novel Hi-C and ChIP-seq data from somatic cell lines of non-model species (i.e., Tasmanian devil, African elephant and aardvark) and compared them with available sequencing data from representative mammalian species of major phylogroups. Moreover, we integrated structural and functional data from germ cells to generate a model that integrates the role of meiosis in genome evolutionary plasticity.

Our results indicate that the 3D genome hierarchical organisation is linked to mammalian chromosomal evolution. Within mammalian species we detected two distinctive nuclear chromosomal configurations in somatic cells: i) highly compacted medium size eutherian chromosomes are organised into chromosomal territories; whereas ii) loose long marsupial chromosomes are organised in a Rabl-like configuration characterised by centromere clustering. Moreover, we detected that this variability in chromosome folding was associated with a decrease of CTCF density and DNA loop lengthening in species with long chromosomes. Finally, we showed that different chromosomal organisations do not alter TADs, confirming the essential nature of TADs for mammalian evolution. Regardless of TADs conservation, we detected contrasting patterns of chromosome evolution associated with chromosome occupancy and folding. The reconstruction of the ancestral karyotypes of marsupial and eutherian mammals (i.e., afrotherians and rodents) revealed lineage-specic evolutionary trends. While the Tasmanian devil (2n=14) exhibits only inversions from the marsupial ancestor (2n=14); afrotherian and rodent species present high complex chromosomal rearrangements (CRs) (fissions, fusions, translocations and inversion), when compared to their respective ancestral karyotypes (2n=48 for afrotheria and 2n=52 for rodentia). Moreover, we detected that in all phylogroups analysed inversions presented a significant increase of intra-chromosomal interactions and CTCF density when compared to colinear regions, suggesting that inversions can act as structural island of diversity where species specific traits evolve.

Our findings also highlight the relevance of interpreting CRs in the context of the formation of the germ line and the meiotic process. We investigated how epigenetic features and the 3D chromatin remodelling that takes place during mouse spermatogenesis influences the formation and transmission of evolutionary CRs. The reconstruction of the ancestral rodentia genome allowed us to characterise the genomic and epigenetic features of mouse-specific evolutionary breakpoint regions (EBRs) in three germ cell types: spermatogonia (pre-meiosis); primary spermatocytes (meiosis); and round spermatids (post-meiosis). We detected that EBRs showed distinctive patterns: i) they do not disrupt meiosis-essential genes; ii) they are positively associated with genomic regions that become accessible during spermatogenesis; iii) they are located at TAD boundaries in pre-meiotic and meiotic cells, but within TADs in post-meiotic cells; and iv) they co-localised with long-range spermatid-specific interactions of reproductive relevant genes that recapitulate ancestral chromosomal interactions. Overall, our results suggest that CR formation is dependent of chromatin and epigenetic remodelling in the germ line.

Mammalian chromosome evolution is highly diverse since a notable variability in chromosome sizes and diploid numbers has been reported. The hierarchical 3D genome organisation (i.e., the way chromosomes are folded within the nucleus) adds another layer of complexity in the field of chromosome evolution. In this context, the study of the similarities and differences of chromosome folding and genomic interactions across species is key to understand the dynamics of genome function and speciation.

This thesis explores the dynamics of 3D chromosome folding across mammalian species representative of key phylogroups. In this context, we set two specific objectives: i) determine principles of chromatin folding, sequence conservation and evolutionary chromosomal reorganisations in mammalian somatic cells; and ii) analyse the role of chromatin remodelling in the generation of chromosomal reorganisations during the formation of germ cells. To achieve this, we employed a comprehensive integrative computational analysis, combining next generation sequencing technologies, including Hi-C, ChIP-seq and RNA-seq, with an integrative bioinformatic analysis. We generated novel Hi-C and ChIP-seq data from somatic cell lines of non-model species (i.e., Tasmanian devil, African elephant and aardvark) and compared them with available sequencing data from representative mammalian species of major phylogroups. Moreover, we integrated structural and functional data from germ cells to generate a model that integrates the role of meiosis in genome evolutionary plasticity.

Our results indicate that the 3D genome hierarchical organisation is linked to mammalian chromosomal evolution. Within mammalian species we detected two distinctive nuclear chromosomal configurations in somatic cells: i) highly compacted medium size eutherian chromosomes are organised into chromosomal territories; whereas ii) loose long marsupial chromosomes are organised in a Rabl-like configuration characterised by centromere clustering. Moreover, we detected that this variability in chromosome folding was associated with a decrease of CTCF density and DNA loop lengthening in species with long chromosomes. Finally, we showed that different chromosomal organisations do not alter TADs, confirming the essential nature of TADs for mammalian evolution. Regardless of TADs conservation, we detected contrasting patterns of chromosome evolution associated with chromosome occupancy and folding. The reconstruction of the ancestral karyotypes of marsupial and eutherian mammals (i.e., afrotherians and rodents) revealed lineage-specic evolutionary trends. While the Tasmanian devil (2n=14) exhibits only inversions from the marsupial ancestor (2n=14); afrotherian and rodent species present high complex chromosomal rearrangements (CRs) (fissions, fusions, translocations and inversion), when compared to their respective ancestral karyotypes (2n=48 for afrotheria and 2n=52 for rodentia). Moreover, we detected that in all phylogroups analysed inversions presented a significant increase of intra-chromosomal interactions and CTCF density when compared to colinear regions, suggesting that inversions can act as structural island of diversity where species specific traits evolve.

Our findings also highlight the relevance of interpreting CRs in the context of the formation of the germ line and the meiotic process. We investigated how epigenetic features and the 3D chromatin remodelling that takes place during mouse spermatogenesis influences the formation and transmission of evolutionary CRs. The reconstruction of the ancestral rodentia genome allowed us to characterise the genomic and epigenetic features of mouse-specific evolutionary breakpoint regions (EBRs) in three germ cell types: spermatogonia (pre-meiosis); primary spermatocytes (meiosis); and round spermatids (post-meiosis). We detected that EBRs showed distinctive patterns: i) they do not disrupt meiosis-essential genes; ii) they are positively associated with genomic regions that become accessible during spermatogenesis; iii) they are located at TAD boundaries in pre-meiotic and meiotic cells, but within TADs in post-meiotic cells; and iv) they co-localised with long-range spermatid-specific interactions of reproductive relevant genes that recapitulate ancestral chromosomal interactions. Overall, our results suggest that CR formation is dependent of chromatin and epigenetic remodelling in the germ line.