Evidence of binding between diethylstilbestrol (DES) and the goldfish (Carassius auratus) membrane progesterone receptor α

Abstract

Background

In a previous study, diethylstilbestrol (DES) was shown to induce oocyte maturation in fish. In the present study, the interaction of DES on goldfish membrane progesterone receptor α (GmPRα) was investigated using a competitive binding assay with radiolabeled steroids. The results indicate that DES exerts its effects on membrane progesterone receptor alpha (mPRα) and induces oocyte maturation through nongenomic steroid mechanisms. This study provides empirical data that demonstrate the binding between DES and GmPRα.

Methods

Binding of DES to GmPRα was achieved by using radiolabeled DES and recombinant GmPRα expressed in culture cells or purified GmPRα proteins that coupled to graphene quantum dots (GQDs). Additionally, the competitive binding of fluorescently labeled progesterone to GmPRα-expressing cells was evaluated.

Results

Although significant nonspecific binding of radiolabeled DES to the cell membrane that expresses GmPRα has been observed, specific binding of DES to GmPRα has been successfully identified in the presence of digitonin. Furthermore, the specific binding of DES to GmPRα was confirmed by a binding assay using GQD-GmPRα. The radiolabeled DES was shown to bind to GQD-GmPRα. Additionally, the competition for the binding of fluorescently labeled progesterone to GmPRα-expressing cells was achieved with the DES.

Conclusions

The results of the experiments revealed that DES binds to GmPRα. Thus, it can be concluded that DES induces goldfish oocyte maturation by binding to GmPRα.

Keywords:

• Diethylstilbestrol (DES)
• mPRα
• goldfish
• oocyte maturation
• endocrine disrupting chemicals (EDCs)

Acknowledgments

This study was supported by Grants-in-Aid for Scientific Research in Priority Areas from the Ministry of Education, Culture, Sports, Science and JSPS KAKENHI Grant Number 23K05830 (to TT).

Author contributions

MFH performed the binding assays, analyzed the data, and drafted the manuscript. UHM conducted microscopic observation of cells. TT participated in the study design, supervised the study, and wrote the paper. All the authors have read and approved the final manuscript.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Data availability statement

The datasets generated and/or analyzed during the current study are available from the corresponding author upon reasonable request.