Periodic Reporting for period 1 - MuSCel Genome (Muscle Specific C. elegans Genome in health and disease: finding novel factors in 3D organization)
Reporting period: 2017-04-01 to 2019-03-31
In MuSCel Genome we aimed to understand factors involved in 3D organization of the genome in cell nuclei and how they contribute to the maintenance of a differentiated cell state. The loss of structural integrity in the interphase nucleus is known to lead to tissue-specific pathologies in a large set of degenerative human diseases collectively called laminopathies, which are late-onset and tissue-specific, with muscle tissue being the most commonly affected. While rare, these conditions provide insight into more common disorders, including sarcopenia, muscular dystrophy, insulin resistance and aging. Understanding the mechanisms behind laminopathy related diseases may lead to new treatments for these degenerative pathologies, particularly those associated with aging. Our overall objective was to ask: How does muscle specific chromatin organization maintains tissue integrity?
Mutations in the nuclear structural protein lamin A produce rare, tissue-specific diseases called laminopathies. The introduction of a human Emery Dreifuss Muscular Dystrophy (EDMD)-inducing mutation (lamin A-Y45C) into lamin (LMN-Y59C) in worms, recapitulates many EDMD phenotypes, and correlates with hyper-sequestration of heterochromatic arrays at the nuclear periphery. Using muscle-specific emerin Dam-ID, we document the misorganization of endogenous chromatin in the LMN-Y59C mutant. We score increased perinuclear positioning along chromosome arms, and enhanced release of chromosomal centers, which accompany the disease phenotypes of reduced locomotion and compromised sarcomere integrity. By coupling the Y59C mutation with deletion of the perinuclear chromodomain protein CEC-4, which tethers H3K9-methylated chromatin, we rescue the EDMD-like physiology and ultrastructural defects in sarcomeres. Deletion of cec-4 also rescued the Y59C-induced changes in chromatin organization. Gene expression changes provoked by LMN-Y59C are also largely reversed by cec-4 deletion. The promoters of genes that change position in the LMNY59C mutant, are enriched for E2F (EFL-1/-2) binding sites, consistent with previous studies implicating the Rb-E2F interaction with lamin A in muscle dysfunction. In summary, the ablation of a perinuclear H3K9me-anchor can counteract the dominant muscle-specific defects provoked by a laminopathic mutation, implicating peripheral chromatin organization in the control of muscle integrity.
The manuscript reporting our findings has been submitted and reviewed. It is currently in revision at the journal Genes and Development. The publication of this work at a major scientific journal will be its primary mode of dissemination. It has also been presented at multiple international conferences.
The manuscript reporting our findings has been submitted and reviewed. It is currently in revision at the journal Genes and Development. The publication of this work at a major scientific journal will be its primary mode of dissemination. It has also been presented at multiple international conferences.
In this project we have used C. elegans as a model system to model a laminopathic disease and to investigate how loss of genome organization may attribute to disease phenotype. By removing a nuclear envelope associated protein and H3K9me chromatin anchor, CEC-4, we have reversed defects in chromatin organization, gene expression, muscle ultrastructure and locomotion found in an EDMD lamin mutant. This project implemented a novel muscle-specific DamID to map chromatin interactions at the nuclear envelope and show the alterations in 3D genome organization that are a consequence of a lamin mutant. Many of the regions that had an altered organization in the presence of mutant lamin, were reversed when CEC-4 was removed. Gene expression changes provoked by LMN-Y59C are also largely reversed by deletion of cec-4. In parallel with muscle-ultra structure data obtained by microscopy and locomotion assays, this work sheds light into the mechanism behind a laminopathic disease. This work is of great interest to the laminopathic community and will inform future studies delving into details of the mechanism(s) behind this class of diseases as well as potentially informing studies into future treatments.