Lamin-genome interactions in the development of Familial Partial Lipodystrophy

Mutations in molecules of the envelope that protects chromosomes, called lamins, cause diseases that result in a melting of fat tissue in lower parts of the body, accumulation of fat in the abdomen and metabolic complications often leading to diabetes. This disease is called a partial lipodystrophy. We have earlier reported that a gene called MIR335 is deregulated by the lamin mutation, and identified mechanisms behind this. Importantly, deregulation of this gene causes a problem with adipocyte differentiation which can be related to the lipodystrophy in patients. We now also show that in cells from these patients, the 3-dimensional (3D) conformation of chromosomes around the MIR335 gene is changed compared to control healthy individuals: the MIR335 gene interacts physically with other regions in the chromosome in patients than in healthy individuals. This means that deregulation of this gene is linked to more global abnormalities in how genes are organized and regulated in patients. This opens up new and exciting avenues of research in the area of metabolic disorders and, perhaps even more importantly, obesity.

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Familial partial lipodystrophy of Dunnigan type (FPLD2) is a laminopathy that mainly affects adipose tissue, resulting in partial lipoatrophy and severe metabolic disorders. FPLD2 is caused by the p.R482W mutation in lamin A. The mutation impairs lamin A interaction with DNA and nucleosomes in vitro, and in patient cells results in dysmorphic nuclei. We rationalize that the p.R482W mutation causes tissue-specific alterations in lamin-genome contacts which reorganize chromatin, thereby mis-positioning developmental genes, deregulating transcription, and leading to defects in adipogenesis and fat tissue metabolism. We aim to uncover the role of the LMNA p.R482W mutation on the formation of lamina-associated domains (LADs) and the impact of defective mutant LADs on the commitment of progenitor cells to adipogenesis. Using FPLD2 patient cells, patient-derived iPS cells for disease modeling, and engineered adipose stem cells (ASCs), the project combines stem cell biology, high-throughput genomics (LMNA ChIP-seq, RNA-seq, bioinformatics) and a new dCas9-EGFP locus-specific tracking system in living cells for real-time analysis of gene positioning. OBJECTIVE 1 aims to determine how the p.R482W mutation affects the position and genomic composition of LADs and the lineage specificity of dissociation of genes from the lamina on differentiation. OBJECTIVE 2 uses FPLD2 patient-derived iPS cells to identify the developmental role of the LMNA p.R482W mutation on LAD formation and adipogenesis. OBJECTIVE 3 investigates the spatio-temporal dynamics of loci with respect to wild-type and mutated lamina, using a dCas9-EGFP gene tagging strategy (perhaps the most challenging development in this project). The results will bring our understanding of laminopathies to a new level. Impact of the work extends beyond the basics of lamin-genome interactions, and aims to open doors to new possibilities for alleviating tissue-specific symptoms of laminopathies.