Role of Mecp2 in the High-Order Chromatin Structure During Neural Differentiation

The methyl-CpG binding protein 2 (MeCP2) is a transcription factor ubiquitously expressed in mammalian tissues, mutated in Rett syndrome (OMIM #321750), a progressive neurodevelopmental disorder. MeCP2 is known to bind methylated DNA and translate epigenetic information through the recruitment of co-repressor complexes, thereby modulating gene expression and, importantly, regulating higher order heterochromatin structure. At genome wide level, neuronal MeCP2 abundance is comparable to histone-octamer level and its binding tracks methyl-CpG density, suggesting a global regulatory role of MeCP2. To understand the effects of MeCP2 deficiency on heterochromatin organization during neural differentiation, we established a versatile model for murine embryonic stem cells (mESCs) in vitro differentiation. Therefore, we modified Mecp2 deficient (Mecp2-/y) mESCs to generate a cell line expressing green fluorescent protein upon neural differentiation. Both wild type and Mecp2-/y were able to differentiate toward different neuronal and glial cell types. We found that MeCP2 protein levels increase significantly during neural differentiation and accumulate at constitutive heterochromatin (chromocenters). Then, we quantitatively analyzed heterochromatin organization (chromocenter clustering) during neural differentiation in wild type and in Mecp2 deficient cells. Statistical analysis of Mecp2 wild type neurons revealed a significant clustering of chromocenters per nucleus during neural differentiation. In contrast Mecp2 deficient neurons and astroglia cells showed significant impairment in heterochromatin reorganization. Furthermore, the re-expression of MeCP2A and MeCP2B, the two splicing isoforms of MeCP2, separately, in the deficient cell line revealed that MeCP2B is able to almost completely recover the heterochromatin clustering during neural differentiation, whereas the rescue after the re-insertion of MeCP2A is only partial. Our results introduce a new and handy cellular model to study the molecular effects of Mecp2 deficiency. Furthermore, this work supports the view of MeCP2 as a multifunctional and chromatin structure organizing factor and highlight a main role of MeCP2B isoform in the phenomenon.