BioProject

The methyl-cytosine binding protein 2 (MeCP2) is a reader of epigenetic DNA methylation marks and necessary and sufficient to reorganize 3D heterochromatin structure during cellular differentiation, e. More...

The methyl-cytosine binding protein 2 (MeCP2) is a reader of epigenetic DNA methylation marks and necessary and sufficient to reorganize 3D heterochromatin structure during cellular differentiation, e.g., myogenesis. In addition to global expression profile changes, myogenic differentiation is accompanied by 3D-heterochromatin reorganization that is dependent on MeCP2. MeCP2 is enriched at pericentric heterochromatin foci (chromocenters). During myogenesis, the total heterochromatin foci number per nucleus decreases while foci volumes and MeCP2 protein levels increase. Ectopic MeCP2 is able to mimic similar heterochromatin restructuring in the absence of differentiation. We compared expression profile changes during myogenic differentiation to changes related to MeCP2-induced heterochromatin reorganization in the absence of differentiation. We used the Affymetrix 430.2 microarray system to study the expression profile changes during myogenic differentiation (myotubes vs myoblast) and MeCP2 related expression changes in transiently transfected myoblasts (high vs. low MeCP2 protein levels) in five independent experiments for each condition. Overall design: To obtain expression profile changes during myogenic differentiation, five biological replicas of undifferentiated Pmi 28 myoblasts (MB) and in vitro differentiated Pmi 28 myotubes (MT) were cultured as described elsewhere. To obtain MeCP2 dependent and differentiation independent expression profile changes, five biological replica of undifferentiated Pmi 28 myoblasts were transiently transfected with MeCP2-YFP. Subsequently, transfected cells were FACSorted to generate MeCP2-high (R4) and -low (R5) level fractions. Total RNA was prepared for each sample out of cell pellets containing 6.5x10exp5 to 1.7x10exp6 cells and further processed according to the GeneChip Expression Analysis Technical Manual by Affymetrix and hybridized to Affymetrix 430.2 microarrays. Less...