A Novel Dnmt3a Isoform Produced from an Alternative Promoter Localizes to Euchromatin and Its Expression Correlates with Activede Novo Methylation

Abstract

Previous studies have shown that the Dnmt3b gene encodes multiple variants via alternative splicing. However, only one form of Dnmt3a has been identified to date. We report here the discovery of a small form of Dnmt3a, denoted Dnmt3a2, from both human and mouse. The transcript encoding Dnmt3a2 is initiated from a downstream intronic promoter. As a result, the Dnmt3a2 protein lacks the N-terminal 223 (human) or 219 (mouse) amino acid residues of the full-length Dnmt3a. Recombinant Dnmt3a2 protein displayed similar cytosine methyltransferase activity as Dnmt3a in vitro. However, Dnmt3a and Dnmt3a2 exhibited strikingly different subcellular localization patterns. Unlike Dnmt3a, which was concentrated on heterochromatin, Dnmt3a2 displayed a localization pattern suggestive of euchromatin association. Dnmt3a2 is the predominant form in embryonic stem cells and embryonal carcinoma cells and can also be detected from testis, ovary, thymus, and spleen, whereas Dnmt3a is expressed at low levels ubiquitously. Comparison of human embryonal carcinoma cell lines with breast/ovarian cancer cell lines indicates that DNMT3A2 expression correlates with high de novo methylation activity. These findings suggest that Dnmt3a and Dnmt3a2 may have distinct DNA targets and different functions in development.

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

“…We also observed some recombination in the kidney, where Dnmt3a transcripts have been detected (Okano et al, 1998;Chen et al, 2002). Though we cannot rule out the possibility of a peripheral deficit contributing to the phenotype, we did not observe gross histological changes in the kidney.…”

“…In the nervous system, Dnmt3a is expressed largely in neurons, though some glial cells also express the enzyme (Feng et al, 2005). Expression of Dnmt3a in the brainstem and spinal cord is uncharacterized, but the enzyme is expressed in skeletal muscle (Okano et al, 1999;Chen et al, 2002). In the cerebellum, Dnmt3a protein is expressed consistently in cerebellar granule cells and in a fraction of Purkinje cells in adult mice (Feng et al, 2005).…”

Stable Methylation at Promoters Distinguishes Epiblast Stem Cells from Embryonic Stem Cells and the In Vivo Epiblasts

“…In good correlation with transcript level, DNMT3B is greater than eightfold higher in EpiSCs compared with ESCs. As expected, the total quantity of DNMT3A was similar in both cell types, but distributed over two different isoforms: the lower band of *75 kDa corresponding to DNMT3A isoform 2 (DNMT3A2, [51]) is higher in ESCs, whereas the higher band (*100 kDa; DNMT3A1) appeared only in EpiSCs and accounted for about half of the total quantity of DNMT3A in these cells. In conclusion, compared with ESCs, EpiSCs have more abundant DNMT3B and DNMT3A1, whereas Dnmt3l, Tet1, and Tet2 are much lower.…”

“…A recent study have shown that DNMT3L interacts with PRC2, the polycomb complex that tri-methylates H3K27, which helps in maintaining the bivalent domains free of DNA methylation by preventing their access by DNMT3A/3B [56]. Although the total quantity of DNMT3A remains constant in EpiSCs and ESCs, an additional splicing isoform, DNMT3A1, is present in EpiSCs [51]. Interestingly, this isoform is also upregulated upon retinoic acid ESC differentiation and exhibits distinct gene targets [57].…”

Sex-specific promoters regulate Dnmt3L expression in mouse germ cells

“…Alternative promoter usage has also been reported for the Dnmt3a (Chen et al, 2002) and Dnmt1 (Mertineit et al, 1998) genes ( Figure 5). A second promoter at the Dnmt3a locus produces a shorter, more active form of the protein known as Dnmt3a2 specifically in tissues showing de-novo methylation, including the germ cells of both sexes: however, no evidence has been found so far of any sex-specific transcripts of this gene.…”