The estrogen receptor α cistrome in human endometrium and epithelial organoids

Abstract

ContextEndometrial health is impacted by molecular processes that underlie estrogen responses.ObjectiveTo define estrogen regulation of endometrial function by integrating the estrogen receptor alpha (ESR1) cistrome and transcriptome of endometrial biopsies taken from the proliferative and midsecretory phases of the menstrual cycle and hormonally stimulated endometrial epithelial organoids.DesignESR1 ChIPseq and RNAseq were performed on proliferative or mid-secretory endometrial biopsies and on hormone treated organoid cultures.SettingEndometrial samples were obtained from volunteers at outpatient research clinics for ChIPseq and for organoid culture.Patients or Other ParticipantsParticipants were fertile, reproductive aged women with normal cycle length, and without any history of infertility or irregular cycles. In total, 5 new endometrial biopsies obtained from 5 women were used in this study and were analyzed together with previously published cycle stage endometrial RNAseq data.Intervention(s)There were no interventions in this study.Main Outcome Measure(s)The cycle stage specific ESR1 binding sites and gene expression identification of human endometrium and organoid cultures were integrated with changes in gene expression.ResultsGenes with ESR1 binding in whole endometrium were enriched for chromatin modification and regulation of cell proliferation. The distribution of ESR1 binding sites in organoids was more distal to the gene promoter when compared to primary endometrium. Organoid estrogen/ESR1 candidate target genes impacted formation of cellular protrusions, and chromatin modification,ConclusionsAnalysis of the ESR1 cistromes and transcriptomes from endometrium and organoids provides important resources for understanding how estrogen impacts endometrial health and function.

Progesterone Signaling in Endometrial Epithelial Organoids

“…We previously described our use of endometrial organoids as an estrogen-responsive model to characterize the ESR1 transcriptome and cistrome in human endometrial epithelial cells [ 26 ]. To test the progesterone response of the organoids, we added analysis of organoids that were treated with estradiol (E2) to increase expression of PGR and then treated with E2 together with the progesterone agonist, medroxyprogesterone acetate (MPA).…”

“…We previously observed that donor two derived organoids were less responsive to estrogen than donor one derived organoids [ 26 ]. Similarly, E2+MPA produces fewer DEG from donor two ( Figure S1b, Table S1a,b ).…”

“…Organoid culture, isolation, and analysis of RNA and chromatin were described previously [ 26 ]. Organoids were plated and allowed to form for 4 days.…”

“…RNA was isolated using Trizol (Life Technologies Inc., Carlsbad, CA, USA) and analyzed by RT-PCR or RNAseq as previously described [ 26 ]. For RT-PCR, cDNA was prepared using Superscript II (Invitrogen, Carlsbad, CA, USA) with Random Hexamers (Invitrogen, Waltham, MA, USA), as previously described [ 27 , 28 ].…”

“…Here, we assessed the progesterone-induced responses of human endometrial epithelial organoids by examining their transcriptome and PGR cistrome. We integrated these findings, and our previously published estrogen-induced transcriptional responses and ESR1 cistrome of human endometrial epithelial organoids [ 26 ], with HiC analysis of the 3D arrangement of human endometrial epithelial organoid chromatin to find candidate ESR1 or PGR gene interactions.…”