Periodic Reporting for period 2 - TopoGenomics (Topological interactions as functional regulators of the eukaryotic genome: moving beyond intramolecular looping)
Okres sprawozdawczy: 2023-03-01 do 2024-08-31
Genetic information is encoded in DNA molecules, which can be very long in species with large genomes. The DNA molecules hence need to be properly organized inside the tiny volume of cells. This three-dimensional organization of DNA molecules needs to enable long-range interactions between distant DNA segments, which play fundamental roles in the function of genomes, including gene regulation, genetic recombination, and transmission of the genome across cell generations.
Key methodological advances over the last 10 years have led to tremendous progress in understanding the molecular mechanisms shaping intramolecular chromosome loop structures. In contrast, while topological interactions across separate chromosomal DNA molecules are well known to contribute to essential biological processes, limitations in our ability to distinguish specific chromosome conformations across paired molecules has hindered research into the underlying mechanisms. Our development of sister chromatid-sensitive chromosome conformation capture has recently allowed to distinguish inter-molecular contacts between sister chromatids from intra-molecular contacts forming loops on individual sister chromatids.
In this project, we aim to develop a next-generation version of sister-chromatid-conformation capture to map the conformation of sister chromatids with unprecedented resolution. We will develop a machine learning framework for integrated analysis of chromosome conformation and chromatin fiber composition. We will also develop a super-resolution imaging approach to trace sister-chromatid fibers to reveal conformational variability in individual cells.
With these new technologies, we aim to understand the molecular mechanisms shaping replicated genomes. Combining conformation analysis with profiling of genomic localization will reveal how different molecular factors organizing loops and linkages interact to shape replicated chromosomes. We also aim to uncover how sister-chromatid conformation contributes to physiological processes. Perturbations of sister chromatid conformations will reveal how trans-molecular topological interactions contribute to DNA repair, gene regulation, and resolution of sister-chromatid fibers for chromosome segregation in mitosis.
Our work will establish innovative technologies that are applicable to diverse biological studies. Furthermore, by gaining insights into the basic mechanisms of life, our study will lay the foundation for future work aiming at solving specific problems, e.g. in biomedical applications.
Key methodological advances over the last 10 years have led to tremendous progress in understanding the molecular mechanisms shaping intramolecular chromosome loop structures. In contrast, while topological interactions across separate chromosomal DNA molecules are well known to contribute to essential biological processes, limitations in our ability to distinguish specific chromosome conformations across paired molecules has hindered research into the underlying mechanisms. Our development of sister chromatid-sensitive chromosome conformation capture has recently allowed to distinguish inter-molecular contacts between sister chromatids from intra-molecular contacts forming loops on individual sister chromatids.
In this project, we aim to develop a next-generation version of sister-chromatid-conformation capture to map the conformation of sister chromatids with unprecedented resolution. We will develop a machine learning framework for integrated analysis of chromosome conformation and chromatin fiber composition. We will also develop a super-resolution imaging approach to trace sister-chromatid fibers to reveal conformational variability in individual cells.
With these new technologies, we aim to understand the molecular mechanisms shaping replicated genomes. Combining conformation analysis with profiling of genomic localization will reveal how different molecular factors organizing loops and linkages interact to shape replicated chromosomes. We also aim to uncover how sister-chromatid conformation contributes to physiological processes. Perturbations of sister chromatid conformations will reveal how trans-molecular topological interactions contribute to DNA repair, gene regulation, and resolution of sister-chromatid fibers for chromosome segregation in mitosis.
Our work will establish innovative technologies that are applicable to diverse biological studies. Furthermore, by gaining insights into the basic mechanisms of life, our study will lay the foundation for future work aiming at solving specific problems, e.g. in biomedical applications.
Our work so far has focused on technology development and their application to study how the interplay between loop-forming and cohesive activities shapes replicated chromosomes. We have developed a next-generation sequencing method to distinctly label the two replicated DNA copies of sister chromatids, and a computational framework that allows integrated analysis of three-dimensional organization of the genome and molecular composition of chromatin fibers. We have also developed an imaging-based approach to trace replicated sister chromatid fiber individually in single cells. Together, the new methods have provided detailed insights into the folding of replicated chromosomes, and their regulation by loop-forming and cohesive linkage factors. We have shown that increasing loop extrusion processivity alone is sufficient to resolve the replicated sister chromatids as normally seen only in dividing cells. We also discovered a unique architecture formed around DNA double strand breaks, which might facilitate the repair process. Our ongoing work is aimed at understanding the molecular factors regulating this new architecture, and their functional implications.
Our study will uncover an essential yet poorly understood aspect of chromosome biology: how replicated sister chromatids topologically interact to support maintenance, expression, and segregation of eukaryotic genomes. Our insights into the complex interplay between different machineries regulating topological interactions in replicated chromosomes, including loop extrusion, cohesive linkages, and chromatin compaction, will reveal how genomes dynamically adapt during cell cycle progression, gene regulation, and DNA repair. Given the broad implications of these processes in health and disease, our study will advance science far beyond the specific biological goals of this project.
Inventing sister-chromatid-sensitive chromosome conformation capture has allowed us to pioneer the analysis of intermolecular chromosome conformation. Advancing this technology towards high-resolution mapping of genome conformation and development of complementary sister chromatid fiber tracing will unleash the full potential to study the architecture of replicated genomes. By crossing the boundaries between imaging-, genomics-, and machine-learning-based approaches we will be able to probe chromosome organization across multiple dimensions: genomic position, 3D spatial position, time, and molecular composition of chromatin fibers. Our integrative and multi-disciplinary approach will help solving the mysteries underlying the complex organization of eukaryotic genomes.
Inventing sister-chromatid-sensitive chromosome conformation capture has allowed us to pioneer the analysis of intermolecular chromosome conformation. Advancing this technology towards high-resolution mapping of genome conformation and development of complementary sister chromatid fiber tracing will unleash the full potential to study the architecture of replicated genomes. By crossing the boundaries between imaging-, genomics-, and machine-learning-based approaches we will be able to probe chromosome organization across multiple dimensions: genomic position, 3D spatial position, time, and molecular composition of chromatin fibers. Our integrative and multi-disciplinary approach will help solving the mysteries underlying the complex organization of eukaryotic genomes.