The Distribution, Expression Patterns and Functional Analysis of NR1D1 and NR4A2 in the Reproductive Axis Tissues of the Male Tianzhu White Yak

Abstract

Nuclear hormone receptors NR1D1 and NR4A2 play important roles in the synthesis and metabolism of hormones that are thought to be strictly regulated by the hypothalamus-pituitary-gonad axis (HPG) tissues via gene expression. However, in the yak, the function and regulatory mechanisms of NR1D1 and NR4A2 are not clearly understood. The current study is aimed to investigate the expression patterns, distribution and functions of these two receptors in HPG tissues in male Tianzhu white yaks. Immunohistochemical staining showed NR1D1 and NR4A2 proteins were present in all yak HPG tissues with differential expression patterns and degrees of staining, particularly in Leydig cells that were strongly positive in accordance with the immunofluorescence results. qRT-PCR and Western blot results suggested that the highest expression levels of NR1D1 and NR4A2 mRNA were present in the hypothalamus, while the expression levels of NR1D1 and NR4A2 proteins were higher in the testis and epididymis than in the hypothalamus or pituitary gland. In addition, expression levels of NR1D1 and NR4A2 mRNA and protein in testicular tissues differed by age. Expression levels were significantly higher at 6 years of age. Gene ontology (GO) and pathway analysis enrichment revealed that NR1D1 may directly regulate the synthesis and metabolism of steroid hormones via interaction with different targets, while NR4A2 may indirectly regulate the synthesis and metabolism of steroid hormones. These results showed that NR1D1 and NR4A2, as important mediators, are involved in the regulation of male yak reproduction, and especially of steroid hormones and androgen metabolism. These results will be helpful for the further understanding of the regulatory mechanisms of NR1D1 and NR4A2 in yak reproduction.