The effects of estradiol and estriol secretion pattern on estrogen receptor dynamics and gene regulation

Authors

• L Hoang
• D Stavreva
• K Wagh
• H LaPoint
• L Schiltz
• A Upadhyaya
• R Chari
• G Hager

Abstract

Previous studies have established a strong relationship between circadian rhythms and endocrine homeostasis. However, the secretion of many hormones follows both circadian and ultradian patterns. Utilizing the glucocorticoid receptor (GR) as a model system, the impacts of the hormone stimulation patterns on GR dynamics and downstream gene regulation have been thoroughly examined. (Stavreva, 2009; Stavreva, 2016, Flynn, 2018; Stavreva, 2019; Lightman, 2020; Kalafatakis, 2021).

Building upon this knowledge, we employ a battery of techniques including genomics, single molecule tracking (SMT), and high-throughput microscopy, to investigate the effects of ultradian and constant estrogen (E2) and estriol (E3) stimulation on the mobility of the estrogen receptor (ER) and the regulation of ER target genes. To do that we have tagged endogenous ER in MCF7 cells with a Halo tag, allowing us upon addition of a Halo ligand (JFX650) to study the spatiotemporal dynamics of Halo-ER upon E2 and E3 stimulation. Furthermore, we visualized RNA synthesis in response to pulsatile and constant hormone treatment at both single-cell and single-promoter levels by using a previously established cell line with 24xMS2 repeats integrated into the 3’ UTR of the ER-responsive TFF1 gene.

Our data revealed that E2 hormone fluctuations have weaker effects on ER dynamics and ER-mediated gene responses. The strong ER-E2 interactions result in continuous gene response of the ER-regulated genes on a population level, even under discontinuous (ultradian) E2 stimulation. In contrast, the less stable ER-E3 interaction allows the ER to detect E3 fluctuations, albeit with a substantial delay.

Scientific Focus Area: Cell Biology

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