Nuclear envelope laminopathies: evidence for developmentally inappropriate chromatin-nuclear envelope interactions

During terminal differentiation of cells, there is typically a transition of the nuclear envelope from the Lamin B protein to Lamin A/C proteins. This is commensurate with exit from the cell cycle, and maintenance of the transcriptional programs associated with the terminally differentiated cells. Dominant missense mutations in Lamin A/C cause a broad spectrum of human genetic disorders, where specific point mutations are associated with defects in specific organs or tissues. We have previously presented a model where Lamin A/C mutations disrupt developmentally appropriate interactions between chromatin and the nuclear envelope and lead to poor coordination of E2F cell cycle pathways and terminal differentiation pathways [1]. One of the phenotypes caused by Lamin A/C mutations is Emery Dreifuss Muscular Dystrophy (EDMD). An X-linked recessive phenocopy of EDMD is caused by loss of function of emerin – a binding partner to Lamin A/C at the nuclear envelope. Here, we tested the hypothesis that emerin plays a role in chromatin remodeling via stabilizing nuclear lamina-heterochromatin interactions necessary for appropriate and time dependent muscle differentiation. We used WT and emerin null mouse myogenic stem cells to study transcriptional and epigenetic changes during in vitro exit from the cell cycle and differentiation to the myogenic lineage. Specific cell cycle (E2F) and myogenic genes were analyzed by qPCR and ChlP-qPCR to determine mRNA timing and H3K9me3 enrichment on gene promoters. Nuclear lamina-chromatin colocalization was determined and quantified by confocal imaging and Matlab. Our results showed that TK1 and other cell cycle genes are inappropriately persistently expressed in emerin null cells during differentiation causing delayed exit from cell cycle. Transcripts marking commitment to the myogenic lineage (myogenin and Mef5A) showed delayed activation on both mRNA and protein level. Epigenetic imprints predicted observed deviations from transcriptional timing in emerin null cells, with persistent suppressive chromatin state on myog promoter upon myogenic induction and failure to appropriately establish repressive histone marks (H3K9me3) on Tk1 promoter (cell cycle). Finally, we showed that the early cell cycle exit and terminal differentiation of emerin null myoblasts were accompanied by decreased H3K9me3 staining at the nuclear periphery (lamin A/C immunostaining). Myogenic cells lacking emerin exhibit perturbations in terminal commitment to the myogenic lineage. Our transcriptional, chromatin remodeling and gene promoter accessibility data show that both exit from cell cycle and terminal commitment to myogenesis are disrupted due to inappropriate heterochromatin-nuclear lamina interactions in EMD myogenic cells.https://doi.org/10.1186/1756-8935-6-S1-P6

The clinical isolate Pseudomonas aeruginosa MMA83 carries two copies of the blaNDM-1 gene in a novel genetic context.

The genetic context of the bla(NDM-1) gene in the genome of Pseudomonas aeruginosa MMA83 was investigated. Sequencing of the cosmid selected for the bla(NDM-1) gene revealed the presence of two bla(NDM-1) copies in the genome of P. aeruginosa MMA83 in a unique genetic environment. Additionally, mating assays, DNA-DNA hybridization, and an S1 nuclease assay strongly suggest that the bla(NDM-1) gene in P. aeruginosa MMA83 is chromosome borne

Laminopathies disrupt epigenomic developmental programs and cell fate.

The nuclear envelope protein lamin A is encoded by the lamin A/C (LMNA) gene, which can contain missense mutations that cause Emery-Dreifuss muscular dystrophy (EDMD) (p.R453W). We fused mutated forms of the lamin A protein to bacterial DNA adenine methyltransferase (Dam) to define euchromatic-heterochromatin (epigenomic) transitions at the nuclear envelope during myogenesis (using DamID-seq). Lamin A missense mutations disrupted appropriate formation of lamin A–associated heterochromatin domains in an allele-specific manner—findings that were confirmed by chromatin immunoprecipitation–DNA sequencing (ChIP-seq) in murine H2K cells and DNA methylation studies in fibroblasts from muscular dystrophy patient who carried a distinct LMNA mutation (p.H222P). Observed perturbations of the epigenomic transitions included exit from pluripotency and cell cycle programs [euchromatin (open, transcribed) to heterochromatin (closed, silent)], as well as induction of myogenic loci (heterochromatin to euchromatin). In muscle biopsies from patients with either a gain- or change-of-function LMNA gene mutation or a loss-of-function mutation in the emerin gene, both of which cause EDMD, we observed inappropriate loss of heterochromatin formation at the Sox2 pluripotency locus, which was associated with persistent mRNA expression of Sox2. Overexpression of Sox2 inhibited myogenic differentiation in human immortalized myoblasts. Our findings suggest that nuclear envelopathies are disorders of developmental epigenetic programming that result from altered formation of lamina-associated domains