Prevention of MECP2 phosphorylation at S423 changes global transcription in primary cortical neurons co-cultured with BV2 cells upon LPS/IFNγ-induced activation.

Methyl-CpG-binding protein 2 (MECP2) is a transcriptional regulator critical for synaptic function. Dysfunction of synapses, in turn, as well as microglia-mediated neuroinflammation belong to the earliest pathological events in Alzheimer’s disease (AD). Functional versatility of MECP2, including its affinity for different binding partners, transcriptional regulation of different gene sets, and effects on neuronal plasticity, are modulated by post-translational modifications, such as phosphorylation. To characterize expression changes through blocking of MECP2 S80 and S423 phosphorylation in neuron-BV2 co-cultures upon induction of neuroinflammation, mouse primary cortical neurons were transduced with MECP2-WT and MECP2-S80A and MECP2-S423A phopsho-variants. Neuroinflammation was induced with LPS/IFNγ and the samples were subjected to RNA sequencing. In neurons co-cultured with BV2 cells, blocking of MECP2 S423 phosphorylation increased the expression of several genes important for neuron and synapse maintenance and protection upon inflammatory stress conditions. Blocking of S80 phosphorylation did not lead to major global expression changes.The results suggest that MECP2 S423 phosphorylation might play a role in activation of neuronal gene expression conveying neuroprotection under neuroinflammation related stress conditions. Cortical DIV4 neurons were transduced with lentivirus vectors encoding MECP2-WT-Myc-IRES-ZsGreen (MECP2 reference sequece: NMNM_004992), MECP2-S80A-Myc-IRES-ZsGreen, MECP-S423A-Myc-IRES-ZsGreen or ZsGreen only (Control). BV2 cells were added 48 h after transduction and inflammation was induced with LPS (200 ng/ml) and IFNγ (20 ng/ml). Four replicates of primary cortical neurons were transduced independently of each other in each of the four sample groups. 16 RNA samples were analyzed in total.