Functional studies on methylation of the promoter region of the NR2E1 gene regulating adipocytes in beef cattle.

Background: Methylation plays a pivotal role in the modulation of gene expression, with the occurrence of promoter and exon 1 methylation typically resulting in transcriptional repression. Lipid deposition constitutes a complex economic trait, with a multitude of genes participating in the regulatory cascade that shapes meat quality attributes. Nonetheless, the available data on genetic markers for meat quality and the corresponding functional genes remains scarce, posing significant challenges to the selection of superior breeds and the development of novel strains in beef cattle breeding programs.

Results: In the present investigation, we evaluated the methylation profile of the NR2E1 gene within the promoter CpG islands across three distinct breeds of beef cattle, and elucidated its regulatory role in modulating the proliferation, differentiation, and apoptosis of adipocytes specifically in the Guyuan cattle breed. Subsequently, the functional validation of the NR2E1 gene was conducted in the predipocyte of Guyuan cattle. The results revealed an inverse correlation between the methylation level assessments and the RNA-seq data. Specifically, the mean methylation levels in 10 CpG islands situated within the promoter region of the NR2E1 gene were observed to be highest in Wagyu cattle, with Guyuan cattle exhibiting the subsequent highest levels, and the lowest in Chinese Simmental cattle. This methylation pattern exhibited a negative correlation with the RNA-seq outcomes. The interference with the NR2E1 gene repressed the proliferation and differentiation of bovine adipocytes by down-regulating the expression of both proliferation-associated genes and differentiation-related genes. Moreover, it potentially facilitated apoptosis in bovine adipocytes through up-regulating the pro-apoptotic gene BAD and down-regulating the apoptosis suppressor gene BCL2. Further downstream pathway analysis revealed that interference with the NR2E1 gene activated the AKT pathway, whereas its over expression led to the inhibition of this pathway.

Conclusions: In the present investigation, we conducted methylation sequencing of the CpG island within the promoter region of the NR2E1 in three breeds of beef cattle, followed by a comprehensive co-analysis with transcriptome data. We further explored the influence of the NR2E1 on the proliferation, differentiation, apoptosis, and downstream signaling pathways in bovine adipocytes. The findings of this research serve as a foundational reference for advancing the molecular understanding of NR2E1 regulation in the context of bovine adipocyte biology.