DNA Damage Induction Alters the Expression of Ubiquitin and SUMO Regulators in Preimplantation Stage Pig Embryos

Abstract

DNA damage in early-stage embryos impacts development and is a risk factor for segregation of altered genomes. DNA damage response (DDR) encompasses a sophisticated network of proteins involved in sensing, signaling, and repairing damage. DDR is regulated by reversible post-translational modifications including acetylation, methylation, phosphorylation, ubiquitylation, and SUMOylation. While important regulators of these processes have been characterized in somatic cells, their roles in early-stage embryos remain broadly unknown. The objective of this study was to explore how ubiquitylation and SUMOylation are involved in the regulation of early development in porcine embryos by assessing the mRNA profile of genes encoding ubiquitination (UBs), deubiquitination (DUBs), SUMOylation (SUMOs) or deSUMOylation (deSUMOs) enzymes in oocyte and embryos at different stages of development, and to evaluate if the induction of DNA damage at different stages of embryo development would alter the mRNA abundance of these genes. Pig embryos were produced by in vitro fertilization and DNA damage was induced by ultraviolet (UV) light exposure for 10 s on days 2, 4 or 7 of development. The relative mRNA abundance of most UBs, DUBs, SUMOs, and deSUMOs was higher in oocytes and early-stage embryos than in blastocysts. Transcript levels for UBs (RNF20, RNF40, RNF114, RNF169, CUL5, DCAF2, DECAF13, and DDB1), DUBs (USP16), and SUMOs (CBX4, UBA2 and UBC9), were upregulated in early-stage embryos (D2 and/or D4) compared to oocytes and blastocysts. In response to UV-induced DNA damage, transcript levels of several UBs, DUBs, SUMOs, and deSUMOs decreased in D2 and D4 embryos, but increased in blastocysts. These findings revealed that transcript levels of genes encoding for important UBs, DUBs, SUMOs, and deSUMOs are regulated during early embryo development and are modulated in response to induced DNA damage. This study has also identified candidate genes controlling post-translational modifications that may have relevant roles in the regulation of normal embryo development, repair of damaged DNA, and preservation of genome stability in the pig embryo.

Simultaneous Inhibition of Histone Deacetylases and RNA Synthesis Enables Totipotency Reprogramming in Pig SCNT Embryos

“…Besides regulators of histone modifications, we observed higher mRNA levels for the ubiquitin ligase RNF114 on D2 and the SUMO-conjugating UBC9 and SUMO-activating UBA2 genes on D4 embryos that were treated with Scriptaid + DRB compared with Scriptaid only. These genes were shown to be upregulated during EGA transition and in response to DNA damage induction in pig embryos [51], and supplementation of culture medium with UBA2 improved pig embryo development [52]. In addition, we found that Scriptaid + DRB treatment increased mRNA for EIF1AX on D2, D3 and D4 embryos, and EIF2A on D4 embryos, which are important markers of EGA in porcine embryos [26,53].…”

Noval advance of histone modification in inflammatory skin diseases and related treatment methods

“…Additionally, it enhances the effectiveness of histone acetylation ( 44 , 45 ). Ubiquitin and small ubiquitin-like modifiers (SUMO) are conserved small proteins that undergo covalent attachment to an ϵ-amino group of a lysine residue on the histone ( 46 , 47 ). Ubiquitination and sumoylation are associated with DNA double-strand break repair ( 48 , 49 ).…”