Abstract
Mutations in the human X-linked gene MECP2 are responsible for most Rett syndrome (RTT) cases, predominantly within its methyl-CpG-binding domain (MBD). To examine the role of MBD in the pathogenesis of RTT, we generated two MeCP2 mutant constructs, one with a deletion of MBD (MeCP2-ΔMBD), another mimicking a mutation of threonine 158 within the MBD (MeCP2-T158M) found in RTT patients. MeCP2 knockdown resulted in a decrease in total dendrite length, branching, synapse number, as well as altered spontaneous Ca2+ oscillations in vitro, which could be reversed by expression of full length human MeCP2 (hMeCP2-FL). However, the expression of hMeCP2-ΔMBD in MeCP2-silenced neurons did not rescue the changes in neuronal morphology and spontaneous Ca2+ oscillations, while expression of hMeCP2-T158M in these neurons could only rescue the decrease in dendrite length and branch number. In vivo over expression of hMeCP2-FL but not hMeCP2-ΔMBD in adult newborn neurons of the dentate gyrus also rescued the cell autonomous effect caused by MeCP2 deficiency in dendrites length and branching. Our results demonstrate that an intact and functional MBD is crucial for MeCP2 functions in cultured hippocampal neurons and adult newborn neurons.
Highlights
Rett syndrome (RTT) is a X-linked neurological disorder affecting mostly females
Full-Length MeCP2 is Essential for the Maintenance of Normal Dendritic Development in Primary Hippocampal Neurons To investigate the role of endogenous MeCP2 in dendritic growth in cultured hippocampal neurons, we knocked down the expression of MeCP2 with lentivirus carrying Short hairpin RNA (shRNA) against MeCP2 (Figure 1A)
We examined the changes in spontaneous Ca2+ oscillations after expression of human MeCP2 (hMeCP2)-FL, hMeCP2-∆methyl-CpG-binding domain (MBD) or hMeCP2-T158 mutation (T158M) in MeCP2-silenced neurons
Summary
Rett syndrome (RTT) is a X-linked neurological disorder affecting mostly females. Classical RTT is a progressive neurodevelopment disorder. Girls with RTT exhibit normal development through the first 6–18 month after birth, followed by an abrupt neuroregression and growth stagnation (Neul et al, 2010). Mutations within MECP2 (methyl-CpG binding protein 2) on the X-chromosome are responsible for nearly 95% of all RTT cases (Amir et al, 1999). MeCP2 is a member of the family of methyl-CpG binding domain (MBD) containing proteins that is the most abundant in post-mitotic neurons and it functions as a transcriptional regulator in the brain. MeCP2 contains a N-terminal domain (NTD), a methyl-binding domain (MBD), an intervening domain (ID), a transcriptionalrepressor domain (TRD) and a C-terminal domain (CTD; Hansen et al, 2011).
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