Role of MeCP2 (Meyhyl CpG-Binding Protein) during replication, epigenetic inheritance and chromocenter organization of pericentric heterochromatin.

“Epigenetic” is a term used to describe mitotically and meiotically heritable states of gene expression that are not due to changes in DNA sequence. Epigenetic events are important in all aspects of biology such as cell proliferation, development and differentiation. The maintenance of a correct epigenome is fundamental in the proper progression of silencing effect, pivotal during cellular differentiation. Multi-protein complexes required to enable the heterochromatic stable epigenetic inheritance are associated to the structural organization of heterochromatin that is determined at the time of replication, in mid-late S phase. MeCP2 is a Methyl CpG-binding protein that preferentially binds methylated DNA, localizes with pericentric heterochromatin and it has a key role to mediate large-scale chromatin organization and compaction. While the impact of loss and mutations of MeCP2 has been extensively studied as cause of RTT and other neurodevelopmental and autismspectrum diseases little is known about its function in pericentric heterochromatin replication, structure and epigenetic inheritance. To test whether MeCP2 has a role in these processes, I used proliferating cells to study the effect of MeCP2 functional ablation during cell-cycle S-phase. I found that MeCP2 is not involved in heterochromatin replication, chromocenter organization and epigenetic modifications (H3K9me3, H4K20me3 and DNA methylation). Interestingly, MeCP2 influences cell cycle progression without triggering a strong apoptotic effect. Intriguingly, low levels of LaminB, LBR and LaminA/C proteins were found, suggesting that MeCP2 could be involved in nuclear lamina organization and/or in the expression of these genes. Besides, MeCP2 silencing determines low levels of EZH2 (component of PRC2, one of the two Polycomb-Repressive Complexes). In the light of this finding is intriguing to investigate the link between MeCP2 and EZH2 due the fact that a large number of neuronal differentiation genes which are required for neuronal development cells are bound by Polycomb complexes, whereas MeCP2 is a transcriptional regulator implicated in development of the brain that is required to interpret the DNA methylation signal in neurons. In parallel, using different biochemical tools (TAP, gel filtration, mass spectrometry) I investigated the ability of MeCP2 to bind proteins involved in heterochromatin organization. A new interactor Np95, component of pHDBs (pericentric heterochromatin duplication bodies), was found. My results show that MeCP2 in addition to play a role in maturation of neurons and synaptic plasticity it is implicated in cell proliferation. Besides, MeCP2 might be involved in nuclear envelope stability and/or in the expression of lamins, and in Polycomb proteins pathway. On the basis of this work further experiments should be done to better investigate the new functions of MeCP2