DNA Hypomethylation Perturbs the Function and Survival of CNS Neurons in Postnatal Animals

Abstract

DNA methyltransferase I (Dnmt1), the maintenance enzyme for DNA cytosine methylation, is expressed at high levels in the CNS during embryogenesis and after birth. Because embryos deficient for Dnmt1 die at gastrulation, the role of Dnmt1 in the development and function of the nervous system could not be studied by using this mutation. We therefore used the cre/loxP system to produce conditional mutants that lack Dnmt1 in neuroblasts of embryonic day 12 embryos or in postmitotic neurons of the postnatal animal. Conditional deletion of the Dnmt1 gene resulted in rapid depletion of Dnmt1 proteins, indicating that the enzyme in postmitotic neurons turns over quickly. Dnmt1 deficiency in postmitotic neurons neither affected levels of global DNA methylation nor influenced cell survival during postnatal life. In contrast, Dnmt1 deficiency in mitotic CNS precursor cells resulted in DNA hypomethylation in daughter cells. Whereas mutant embryos carrying 95% hypomethylated cells in the brain died immediately after birth because of respiratory distress, mosaic animals with 30% hypomethylated CNS cells were viable into adulthood. However, these mutant cells were eliminated quickly from the brain within 3 weeks of postnatal life. Thus, hypomethylated CNS neurons were impaired functionally and were selected against at postnatal stages.

Ablation of de novo DNA methyltransferase Dnmt3a in the nervous system leads to neuromuscular defects and shortened lifespan

“…1C). Some recombination was observed in kidney and muscle, and little or none in other organs examined (ovary, lung, heart, and liver), as observed previously (Fan et al, 2001).…”

“…This breeding scheme was necessary because the Nes-Cre1 transgene is imprinted, and 95% or greater recombination in the brain is only observed when the Nes-Cre1 transgene is inherited paternally, whereas maternal transmission results in only 30% recombination (Fan et al, 2001). Dnmt3a 2lox/1lox ; Nes-Cre1 (designated as "mutant") mice obtained from this breeding scheme inherit a recombined (Dnmt3a 1lox ) allele from the father because of Nes-Cre1 activity in the male germline Dubois et al, 2006).…”

“…This methylation interferes with STAT3 binding and disallows the cells from proceeding along the astroglia lineage (Takizawa et al, 2001). Deletion of Dnmt1, which encodes the maintenance DNA methyltransferase, has no obvious effect on postnatal brain function (Fan et al, 2001), but causes neural precursor cells to differentiate prematurely into astroglia .…”

A new perspective on neural tube defects: Folic acid and microRNA misexpression

“…Methylation is essential for synthesis of membrane phospholipids, myelin basic protein, and neurotransmitters as well as for establishing proper gene expression in embryogenesis (Biniszkiewicz et al, 2002;Chen et al, 2001). During embryogenesis, all three major DNA methyltransferases (Dnmt1, Dnmt3a, and Dnmt3b) are expressed, the most important being Dnmt1 and Dnmt3b in establishing and maintaining proper genomic methylation (Fan et al, 2001;Jackson et al, 2004). In mammalian embryonic development, prior to implantation, the genome becomes demethylated by the 8-to 16-cell stage, with the inner cell mass undergoing global remethylation at discrete CpG sites before onset of gastrulation (Finnell et al, 2002;Kafri et al, 1992;Santos et al, 2002).…”

“…Dnmt1-null mice die shortly after gastrulation (between E8.0 and 10.5; Fan et al, 2001), so the use of a conditional Dnmt1 knockout is necessary to study the impact its loss has on later stages of development. In one study, primary mouse embryonic fibroblasts were extracted and analyzed for global methylation and gene expression.…”