Alternative splicing and expression analysis of bovine DNA methyltransferase 1

Abstract

Methylation of specific CG residues in the mammalian genome results in tissue-specific patterns of gene expression, which are critical for cell differentiation. Embryos that fail to establish and maintain proper DNA methylation patterns show severe developmental abnormalities as is the case of DNA methyltransferase 1 (Dnmt1) -deficient embryos. Dnmt1 is the main maintenance methyltransferase in the mouse and its expression is regulated by a splicing mechanism that dictates the expression of stage-specific isoforms. Little is known about Dnmt1 expression in the cow and isoforms of Dnmt1 are yet unknown in this species. Here we demonstrate that the previously described bovine Dnmt1 transcript is ubiquitously expressed in embryos and fetal tissue. In addition, we report the identification of a splice variant of the bovine Dnmt1, which shows a ubiquitous expression pattern. This new transcript was detected using 5 RACE and genomic mapping and its expression pattern was shown to be consistent with a tissue-specific mode of regulation. Furthermore, our analysis shows that the expression of an oocyte-specific isoform of Dnmt1 is unlikely to occur in cattle. The newly reported isoform of Dnmt1 was demonstrated to be, similarly to Dnmt1a, polyadenylated and if translated possess the functional domains necessary for maintenance and de novo methyltransferase activity.

Localization of DNA methyltransferase-1 during oocyte differentiation, in vitro maturation and early embryonic development in cow

“…In fact, a growing body of evidence indicates that the chromatin structure together with DNA methylation mutually reinforce the transcriptional repressive state of DNA (Bird, 2002;Fuks et al, 2003) their limited competences as it is becoming evident that genome wide epigenetic modifications must be set in place during gametogenesis in order to allow a correct developmental program (Albertini et al, 2003;Bao et al, 2000;De La Fuente, 2006;Li, 2002). RT-PCR analysis confirmed the presence of the somatic form of Dnmt1 transcripts in immature bovine oocytes as previously observed (Golding and Westhusin, 2003;Russell and Betts, 2008). Moreover this transcript was always detectable throughout the final phase of oocyte differentiation, independently on the chromatin configuration.…”

“…While Dnmt1s protein is ubiquitously expressed, Dnmt1o is predominantly expressed in the mouse oocyte and preimplantation embryo (Bestor, 2000;Mertineit et al, 1998).The oocyte specific isoform of Dnmt1 has been found in monkey (Vassena et al, 2005) and pig (Jeong et al, 2009) and a novel Dnmt1 isoform was found to be expressed in sheep oocyte, embryo and early fetal lineage, but not in the adult tissue (Taylor et al, 2009). On the contrary, two independent studies indicate that the expression of an oocyte specific isoform of Dnmt1 is unlikely to occur in cattle (Golding and Westhusin, 2003;Russell and Betts, 2008). In mouse, Dnmt1o is thought to be implicated in the maintenance of DNA methylation of imprinted genes at the 8-cells stage of development, when it translocates into the nuclei for one cell cycle (Carlson et al, 1992;Howell et al, 2001;Ratnam et al, 2002).…”

“…Moreover, in the cow the oocyte specific isoform of Dnmt1 (Dnmt1o) was not found to be expressed in oocytes and embryos (Golding and Westhusin, 2003;Russell and Betts, 2008).…”

Effects of Downregulating DNA Methyltransferase 1 Transcript by RNA Interference on DNA Methylation Status of the Satellite I Region and In Vitro Development of Bovine Somatic Cell Nuclear Transfer Embryos

“…These variants are derived by alternative splicing of the Dnmt1 gene [38,39]. Recently, Russell et al [40] indicated that the expression of DNMT1o, which is an oocyte-specific isoform of DNMT1, was not detected in cattle. They also demonstrated that there were two variants of DNMT1, namely, DNMT1a and DNMT1b, by alternative splicing of the DNMT1 gene in cattle.…”

Tasks of some dietary essential nutrients on epigenetics and one carbon metabolism in livestock during production

“…by in vitro trials, all the -DNA methyltransferase mRNA-sequences cloned in bovines, sheep and chicken [10]participate very strong homology with those of mouse and human, that shows a conservative role of DNA methyltransferase in various animals. Different isoforms of Dnmt1, Dnmt3a, and Dnmt3b were also found in bovine; even so the functions of these isoforms in DNA methylation stay unknown [11,12].…”