Epigenetic Regulation of Motor Neuron Cell Death through DNA Methylation

Abstract

DNA methylation is an epigenetic mechanism for gene silencing engaged by DNA methyltransferase (Dnmt)-catalyzed methyl group transfer to cytosine residues in gene regulatory regions. It is unknown if aberrant DNA methylation can cause neurodegeneration. We tested the hypothesis that Dnmts can mediate neuronal cell death. Enforced expression of Dnmt3a induced degeneration of cultured NSC34 cells. During apoptosis of NSC34 cells induced by camptothecin, levels of Dnmt1 and Dnmt3a increased five-fold and two-fold, respectively, and 5-methylcytosine accumulated in nuclei. Truncation mutation of the Dnmt3a catalytic domain and Dnmt3a RNAi blocked apoptosis of cultured neurons. Inhibition of Dnmt catalytic activity with RG108 and procainamide protected cultured neurons from excessive DNA methylation and apoptosis. In vivo, Dnmt1 and Dnmt3a are expressed differentially during mouse brain and spinal cord maturation and in adulthood when Dnmt3a is abundant in synapses and mitochondria. Dnmt1 and Dnmt3a are expressed in motor neurons of adult mouse spinal cord, and, during their apoptosis induced by sciatic nerve avulsion, nuclear and cytoplasmic 5-methylcytosine immunoreactivity, Dnmt3a protein levels, and Dnmt enzyme activity increased preapoptotically. Inhibition of Dnmts with RG108 blocked completely the increase in 5-methycytosine and the apoptosis of motor neurons in mice. In human amyotrophic lateral sclerosis (ALS), motor neurons showed changes in Dnmt1, Dnmt3a, and 5-methylcytosine similar to experimental models. Thus, motor neurons can engage epigenetic mechanisms to drive apoptosis, involving Dnmt upregulation and increased DNA methylation. These cellular mechanisms could be relevant to human ALS pathobiology and disease treatment.

Aberrant Regulation of DNA Methylation in Amyotrophic Lateral Sclerosis: A New Target of Disease Mechanisms

“…Several nearby glial cells (round nuclei) have low or undetectable 5mC. The details of this study showing changes in human ALS motor cortex compared with agematched controls have been reported [70]. Scale bars: panorama 17 μm; inset 8 μm where it is thought to oppose the role of 5mC-mediated transcriptional repression by inhibiting the binding of methyl-CpG binding protein-2 and other DNA binding proteins; thus, 5hmC functions as a facilitator of geneexpression activity [44].…”

“…A possibility is that Dnmts in the adult mammalian brain re-methylate newly incorporated cytosines from G-T mismatch DNA repair after 5mC deamination [67]. We have found that Dnmt1 and Dnmt3a have distinct subcellular localizations in neurons [70], suggesting non-redundant functions. Our discovery that Dnmt3a, but not Dnmt1, is located in presynaptic axon terminals in the adult CNS is exciting because it identifies a local presynaptic possibility for Dnmt3a in regulating synaptic function.…”

“…Several studies have found high levels of Dnmt enzyme activity, mRNA, and protein in adult mammalian central nervous system (CNS) [66][67][68][69][70]. Mature neurons are enriched in Dnmts [66][67][68][69][70].…”

“…Mature neurons are enriched in Dnmts [66][67][68][69][70]. We have shown that Dnmt1 and Dnmt3a are expressed in adult mouse brain and spinal cord, and in adult human cerebral cortex and spinal cord, and that mouse and human spinal motor neurons are enriched in these proteins [70]. The functions of Dnmts in terminally-differentiated postmitotic neurons are not understood.…”

“…Mitochondrial homeostasis is another possible function of Dnmts in neurons [70]. Dnmts are present in mitochondria, but it seems controversial as to which isoform is present in mitochondria.…”