Riassunto analitico
Facioscapulohumeral muscular dystrophy (FSHD) is the third most common hereditary myopathy, associated with a reduced copy number of the D4Z4 macrosatellite at chromosome 4q35, which molecular basis remains to be elucidated. The model proposed is that epigenetic changes following D4Z4 deletion lead to disease through chromatin remodelling and inappropriate expression of nearby genes. Among 4q35 genes, FSHD Region Gene 2 (FRG2) is selectively overexpressed in FSHD patients and is stabilised upon genotoxic noxa with a magnitude inversely related to the number of D4Z4 repeats.
We recently found that FRG2 is not a protein-coding gene, but a chromatin-associated long non-coding RNA (lncRNA).
Our purpose in this work was to decipher the function of this new and unknown lncRNA. We investigated it through the integration of genomic and proteomic approaches. This approach led to the identification of thousands of FRG2 target sites in the human genome and revealed FRG2 association in trans to rDNA sequences at the short arms of all human acrocentric chromosomes. We also defined a large network of interacting proteins, including factors involved in nucleolus function and ribosomal biogenesis. Fluorescence in situ hybridization (FISH) and immuno-FISH experiments in HeLa cells and FSHD and control primary myoblasts revealed that FRG2 exerts its role within the nucleoli, specifically in the Dense Fibrillar Components (DFC), where rDNA transcription takes place. Further analyses of gene silencing in HeLa cells revealed that rDNA transcription increased following FRG2 silencing, and rDNA transcriptional evaluation assay confirmed that nucleolus function was impaired in FSHD samples showing a reduction in rRNAs production.
We concluded that FRG2 lncRNA impairs ribosome biogenesis at early stages in FSHD cells, through the binding of the rDNA arrays and the spreading of silenced chromatin.
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