Biochemical and Genomic Analysis of MeCP2 and Brain-Enriched DNA Methylation
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CitationKinde, Benyam Zerihun. 2016. Biochemical and Genomic Analysis of MeCP2 and Brain-Enriched DNA Methylation. Doctoral dissertation, Harvard University, Graduate School of Arts & Sciences.
AbstractDisruption of the MECP2 gene leads to Rett syndrome (RTT), a severe neurological disorder with features of autism. MECP2 encodes a methyl-CpG-binding protein that has been proposed to function as a transcriptional repressor, but despite numerous studies examining neuronal gene expression in Mecp2 mutants, no coherent model has emerged for how MeCP2 regulates gene transcription.
This dissertation presents our efforts to develop a unified, mechanistic understanding of how MeCP2 regulates gene expression in the brain by employing a combination of genomic, genetic and biochemical approaches. Using an unbiased bioinformatics screen to identify genic attributes that reliably predict gene misregulation in the absence of MeCP2, we identify a genome-wide length-dependent increase in gene transcription in multiple mouse models of RTT and in human RTT brains. We present evidence from biochemical and genomic analyses that MeCP2 tempers the expression long genes by binding to a recently identified neuronally-enriched form of DNA methylation (mCA). Building on the observation that long genes containing a high density of mCA contain the greatest possible number of MeCP2 binding sites per gene, we provide evidence that the degree of repression exerted by MeCP2 is proportional to the total number of molecules bound across the body of gene.
We also consider the role of length-dependent gene regulation by MeCP2, and we find that long genes as a population are highly enriched for neuronal functions and are selectively expressed in the brain. We demonstrate that long genes are targeted for repression not only by MeCP2 but also by the Fragile X syndrome protein FMRP, suggesting that the precise regulation of long genes as a population may be critical to brain development. Finally, we perform an initial test of the functional importance of length-dependent gene misregulation in Rett syndrome models. We show that treatment of neurons lacking MeCP2 with a topoisomerase inhibitor can decrease long gene expression, partially correcting gene misregulation in these cells, and improving one gross measure of RTT-associated cellular deficits. These results support a model in which long gene up-regulation in the absence of MeCP2 drives aspects of RTT pathophysiology.
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