Alpha-ketoglutarate affects murine embryo development through metabolic and epigenetic modulations

Abstract

α-Ketoglutarate (α-KG) is an intermediary metabolite in the tricarboxylic acid (TCA) cycle and functions to inhibit ATPase and maintain the pluripotency of embryonic stem cells (ESCs); however, little is known regarding the effects of α-KG on the development of preimplantation embryos. Herein, we report that α-KG (150 μM) treatment significantly promoted the blastocyst rate, the number of inner cell mass (ICM) cells and foetal growth after embryo transfer. Mechanistic studies revealed two important pathways involved in the α-KG effects on embryo development. First, α-KG modulates mitochondria function by inducing relatively low ATP production without modification of mitochondrial copy number. The relatively low energy metabolism preserves the pluripotency and competence of the ICM. Second, α-KG modifies epigenetics in embryos cultured in vitro by affecting the activity of the DNA demethylation enzyme TET and the DNA methylation gene Dnmt3a to increase the ratio of 5hmC/5mC ratio. Elevation of the 5hmC/5mC ratio not only promotes the pluripotency of the ICM but also leads to a methylation level in an in vitro embryo close to that in an in vivo embryo. All these functions of α-KG collectively contribute to an increase in the number of ICM cells, leading to greater adaptation of cultured embryos to in vitro conditions and promoting foetal growth after embryo transfer. Our findings provide basic knowledge regarding the mechanisms by which α-KG affects embryo development and cell differentiation.

Tricarboxylic Acid Cycle Metabolites as Mediators of DNA Methylation Reprogramming in Bovine Preimplantation Embryos

“…This is due to the fact that fast and slow embryos present consistent differences in terms of gene expression, stress response and energy production mechanisms [10,11], which is particularly intriguing as they are morphologically similar. These molecular and metabolic observations led us to hypothesize that the said differences could be a consequence of the constant crosstalk between energy metabolism and epigenetic reprogramming, i.e., metabolites modulate epigenetics through energy metabolism enzymes, but epigenetic mechanisms can also impact the metabolic profile [2,5,16,17].…”

“…The consequences of α-KG supplementation during in vitro culture were recently described in murine embryos [5], in which α-KG treatment affected TET activity leading to an increase in the 5hmC/5mC ratio, improving cell differentiation and embryo development. In the present study, blastocyst rates and total cell number were barely affected by the modulation of metabolites.…”

“…One example of how metabolism and epigenetics are interconnected in these cells is how a high intracellular α-ketoglutarate (α-KG)/succinate ratio allows the maintenance of high TET activity and reduced DNA methylation, whereas cells at more differentiated states, such as somatic cells, present a lower α-KG/succinate ratio and consequently lower TET activity and higher DNA methylation levels [2]. Similarly, a recent study demonstrated that α-KG levels affect Tet and Dnmt3 activity, increasing the ratio of 5-hydroxymethylcytosine/5-methylcytosine (5hmC/5mC) in murine embryos [5].…”

Metabolic–Epigenetic Axis in Pluripotent State Transitions

“…These processes have been recently investigated in vivo, and the role of αKG in developing embryo has been reported [79]. Specifically, the treatment with αKG improves the blastocyst rate, the number of inner cell mass (ICM) and the fetal growth after embryo transfer, and this occurs, at least in part by modulating the activity of Tet and increasing the 5hmC/5mC ratio [79]. of Tet target genes as well as of several markers of inner cell mass (ICM) and germline cells.…”

Erasing gametes to write blastocysts: metabolism as the new player in epigenetic reprogramming

“…Cellular metabolites have been described as enzyme co-factors, responsible for epigenetic changes or even as substrates for chemical modifications in nucleic acids or structural chromatin proteins, once again reinforcing the relationship between metabolism and the molecular profile of the cells ( Figure 3 ). The characterization of these phenomena in embryos is even more recent ( Ispada et al., 2018 , 2020 ; Zhang et al., 2019b ). Some of these mechanisms are better described below.…”