New Insights Into Progesterone Receptor Signaling in the Endometrium Required for Embryo Implantation

65 1

Journal of Molecular F J DeMayo and J P Lyson Progesterone signaling in the 65:1 T1–T14 Endocrinology endometrium THEMATIC REVIEW 90 YEARS OF PROGESTERONE New insights into progesterone receptor signaling in the endometrium required for embryo implantation

Francesco J DeMayo1 and John P Lydon2

1Reproductive and Developmental Biology Laboratory, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina, USA 2Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, Texas, USA

Correspondence should be addressed to J P Lydon: [email protected]

This review forms part of a special section on 90 years of progesterone. The guest editors for this section are Dr Simak Ali, Imperial College London, UK and Dr Bert W O’Malley, Baylor College of Medicine, USA

Abstract

Progesterone’s ability to maintain pregnancy in eutherian mammals highlighted this Key Words steroid as the ‘hormone of pregnancy’. It was the unique ‘pro-gestational’ bioactivity of ff progesterone progesterone that enabled eventual purification of this ovarian steroid to crystalline ff nuclear receptor form by Willard Myron Allen in the early 1930s. While a functional connection between ff isoforms normal progesterone responses (’progestational proliferation’) of the uterus with the ff endometrium maintenance of pregnancy was quickly appreciated, an understanding of progesterone’s ff receptivity involvement in the early stages of pregnancy establishment was comparatively less ff decidualization well understood. With the aforementioned as historical backdrop, this review focuses ff mediators on a selection of key advances in our understanding of the molecular mechanisms by ff modifiers which progesterone, through its nuclear receptor (the progesterone receptor), drives the development of endometrial receptivity, a transient uterine state that allows for embryo implantation and the establishment of pregnancy. Highlighted in this review are the significant contributions of advanced mouse engineering and genome-wide transcriptomic and cistromic analytics which reveal the pivotal molecular mediators and modifiers that are essential to progesterone-dependent endometrial receptivity and decidualization. With a clearer understanding of the molecular landscape that underpins uterine responsiveness to progesterone during the periimplantation period, we predict that common gynecologic morbidities due to abnormal progesterone responsiveness will Journal of Molecular be more effectively diagnosed and/or treated in the future. Endocrinology (2020) 65, T1–T14

Introduction

Sitting on top of the world… the newly minted physician not only became a father but successfully crystallized the corpus luteum hormone, May 1933 was a ‘glorious month’ for a young Willard progesterone – in his words ‘My friends gave me double Myron Allen (1904–1993) (Allen 2005). For the first time, congratulations and I was sitting on top of the world’ (Allen

-19-0212 Journal of Molecular F J DeMayo and J P Lyson Progesterone signaling in the 65:1 T2 Endocrinology endometrium

2005). Only a few years previously, during his medical in structure with clearly demarcated functional domains training (and as part of his two-year research fellowship (Fig. 1). The PGR is composed of an N-terminal domain for a master’s degree (1927–1929)), Allen along with (NTD), a central DNA-binding domain (DBD) with two George Washington Corner (1889–1981), his anatomy highly conserved zinc fingers, a hinge (H) region, followed professor and mentor at the University of Rochester, by a ligand-binding domain (LBD) near the C-terminus, demonstrated through a series of landmark experiments reviewed in (Diep et al. 2015, Grimm et al. 2016) (Fig. 1). that alcohol extracts of the ovarian corpus luteum were Within this functional domain organization, the PGR sufficient to maintain pregnancy in rabbits that were also contains three activational domains or functions ovariectomized 18 h following copulation (Allen & (AFs) that interface with coregulator proteins within the Corner 1929, Corner & Allen 1929). With the isolation, transcriptional complex; AF1 and AF3 are located within bioassay and official naming of progesterone, the first the NTD, whereas the AF2 resides in the LBD (Fig. 1). chapter of the progesterone story was concluded. The next The PGR comprises two primary receptor isoforms: the chapter would focus on identifying and characterizing the full-length PGR-B (~116 kDa) and the truncated PGR-A ‘receptor’ for this hormone and elucidating its mechanism (~94 kDa); the PGR-A lacks the first 164 amino acid residues of action, and this chapter of study remains a relentless of the NTD (termed the B upstream segment (BUS) (Bain preoccupation for many molecular endocrinologists to et al. 2000)) (Fig. 1). Absence of the AF3 in the PGR-A isoform this day. Inspired by the pioneering research of Jacob is thought to explain, in part, the different transactivational and Monod in the early 1960s, which established the properties reported for these two receptor isoforms in basic regulatory principles of gene expression (Jacob & vitro. The PGR isoforms are frequently co-expressed in the Monod 1961), investigations on the rat uterus by Jensen majority of target tissues and are encoded by a single gene and Gorski (Jensen 1962, Toft & Gorski 1966, Jensen et al. that uses a PGR-B distal promoter and a PGR-A proximal 1968), and on the chick oviduct model by O’Malley et al. promoter in tandem to regulate expression of the respective (1969, 1970) and O’Malley and Schrader (1972) provided receptor isoforms (Kastner et al. 1990). Given the cellular the necessary evidential support for the existence of a coexistence of both isoforms, the physiological response specific intracellular binding protein (’the receptor’) for of a target tissue to progesterone exposure is considered estrogen and progesterone respectively. to be dependent on the net stoichiometry of the PGR-B In honor of the 90th anniversary commemorating the and PGR-A isoforms within a cell population, in which discovery of progesterone, this review focuses on recent the formation of homodimers and/or heterodimers project advances in our understanding of progesterone action distinct transactivational controls on the expression of through its receptor in the uterus that enables embryo select gene sets. implantation and the establishment of pregnancy. As a The classical mechanism of PGR action entails prequel, however, a review of the fundamental structural/ progesterone traversing the lipid bilayer of the target cell functional properties of the intracellular receptor for plasma membrane, interacting with the LBD of the PGR progesterone is instructive. to trigger a conformational change (’the activation step’ (O’Malley et al. 1969, 1970, O’Malley & Schrader 1972)), which results in ligand-bound receptor translocating to The progesterone receptor: an isoform duo the nucleus and directly binding as a dimer to a canonical Whether during the luteal (or secretory) phase of the progesterone response element (PRE) within the promoter menstrual cycle or with pregnancy onset, most of the or distant enhancer of a target gene (Diep et al. 2015, physiological responses to ovarian-derived progesterone Grimm et al. 2016). As part of this process, coregulators are mediated by its specific intracellular receptor, the (i.e. coactivators or corepressors) and associated cofactors progesterone receptor (PGR) (Graham & Clarke 1997). are recruited to the nucleating transcriptional complex Early structural characterization of the PGR demonstrated to activate or silence gene expression, the net result of that the receptor belongs to the nuclear receptor which manifests the progesterone physiological response superfamily of transcription factors (Conneely et al. 1986, at the molecular level. In many physiological contexts, Jeltsch et al. 1986), which includes cognate receptors for PGR expression is induced by estrogen through its nuclear other steroid hormones (including estrogen) as well as receptor, estrogen receptor-α (ESR1) (Graham & Clarke for retinoids, prostanoids, thyroid hormone, and vitamin 1997). D3 (Tsai & O’Malley 1994, Mangelsdorf et al. 1995). As a Studies on human breast cancer cell lines in particular nuclear receptor superfamily member, the PGR is modular revealed that PGR’s expression levels, transcriptional

Figure 1 Functional domain organization of the human progesterone receptor. The progesterone receptor (PGR), a member of the NR3C3 subfamily of the nuclear receptor superfamily (Tsai & O’Malley 1994, Mangelsdorf et al. 1995), is comprised of two isoforms: the full-length PGR-B and the truncated PGR-A; the same gene – located on human chromosome 11 (11q22-q23) – encodes both isoforms. The N-terminal domain (NTD), DNA-binding domain (DBD), hinge (H) region, and ligand-binding domain (LBD) are indicated. The positions of activation functional domains (AF1, AF2, and AF3) are shown; note: the PGR-A isoform lacks AF-3 and multiple phosphorylation events. Phosphorylation sites are denoted by (P); locations of sumoylation, acetylation, and methylation sites are also highlighted. Detailed information on these PTMs is comprehensively described in previous, excellent reviews, such as (Hagan & Lange 2014, Diep et al. 2015, Grimm et al. 2016). The schematic was reproduced and modified from Grimm( et al. 2016) with permission from the Journal of Molecular Biology (license number: 4591510498146). activity, trafficking, and target gene selection are modulated and global 3D organization of the chromatin, which can by post-translational modifications (PTMs) (Hagan & Lange control transcriptional output both in ligand-dependent 2014), which include phosphorylation, ubiquitination, and -independent context (Le Dily et al. 2019). sumoylation, acetylation, and methylation (Fig. 1). Collectively, these mechanisms of action studies Related studies have shown that PGR can also have furnished critical conceptual advances – within an indirectly modulate gene expression through a non- in vitro context – on the diverse mechanisms by which classical mode of action which dispenses with the need PGR markedly expands its signaling bandwidth beyond for direct DNA binding. Instead, PGR tethers to other the classical mode of action. However, findings from these transcription factors bound to DNA, such as activator studies raise important questions to be addressed in the protein 1 (AP1), SP1, NFkB, and signal transducer activator future: (1.) how are extranuclear and nuclear actions of of transcription 3 (Stat3) (Grimm et al. 2016). In addition PGR integrated and coordinated? (2.) is this regulatory to genomic actions, PGR has been implicated in rapid complexity observed in non-cancerous cells, such as non-genomic effects that operate beyond the strictures of uterine cells? and (3.) can these various PGR signaling the nucleus. In this non-classical or extranuclear signaling events be functionally linked to in vivo physiological and/ context, PGR has been shown to bind kinases sequestered or pathophysiological endpoints? with growth factor receptors at the plasma membrane, Having provided an abbreviated account of the which trigger rapid activation of kinase signaling cascades structural and functional properties of the PGR (much of that frequently phosphorylate a myriad of transcription the information gleaned from in vitro studies), the ensuing factors (including PGR) to influence downstream gene- sections will focus on the significant advances in our expression programs (Hagan & Lange 2014, Grimm et al. understanding of uterine PGR action in an in vivo context, 2016). Intriguingly, recent genome topology studies thanks in large part to studies on the mouse and on the support the involvement of PGR (along with ESR1) in local genetically engineered mouse in particular.

Progesterone and its uterine receptor are and economic hardship for women and their families required for pregnancy establishment worldwide. Given progesterone is the quintessential ‘hormone of With a perhydrocyclopentanophenanthrene ring pregnancy’, a comprehensive understanding of the pivotal structure, progesterone received its apt appellation by signaling mechanisms by which progesterone, through its Allen and colleagues (Allen et al. 1935) primarily because receptor, governs endometrial periimplantation biology the steroid hormone maintained gestation (hence ‘pro- is essential if future goals to develop more effective gestation’) in pregnant rabbits previously ovariectomized diagnostics, prognostics, and/or therapeutics to manage (Allen & Corner 1929, Corner & Allen 1929). In the early pregnancy loss are to be realized. However, due decades since, however, rodent studies in particular to obvious ethical restrictions on investigative studies demonstrated that progesterone via PGR in the uterus on the human endometrium, most of our mechanistic is essential not only for the maintenance but also for understanding of endometrial periimplantation responses the establishment of pregnancy. In viviparous placental to progesterone at the cellular and molecular level has mammals (eutherians, including humans), pregnancy primarily been obtained from investigations on the establishment is achieved by successful implantation of murine uterus. the activated blastocyst into the receptive endometrium of the uterus (Psychoyos 1973, Schlafke & Enders 1975, Enders 1976). Held within the pelvis by the endopelvic The mouse: a surrogate for the human fascia, the uterus is a complex composite organ which In the human and mouse, embryo implantation is is composed of the inner endometrium and outer dependent on the endometrium transitioning from a ‘pre- myometrium. The endometrium, which comprises receptive’ to a ‘receptive’ state within a short time-frame the luminal and glandular epithelium as well as the (the window of receptivity) during which the luminal stromal compartment, supports embryo implantation epithelium of the endometrium is transiently permissive and pregnancy progression to term. At parturition, the to blastocyst attachment, adhesion, and subsequent contractile force of the surrounding myometrial circular invasion (Cha et al. 2012b, Pawar et al. 2014). In both and longitudinal smooth muscle layers is required for live species, synchronized development of the endometrial birth delivery. receptive state with the on-time arrival of the activated Advancing our understanding of progesterone’s blastocyst is controlled by ovarian-derived progesterone involvement in early pregnancy establishment is of in tight coordination with estrogen. Although the utmost clinical importance as embryo implantation is an endometrial cellular changes that enable embryo inefficient reproductive process, with implantation failure implantation in humans are interstitial as opposed to and early embryo miscarriage still representing significant eccentric in the murine endometrium, the execution etiologic factors for preclinical pregnancy loss even for of the basic developmental steps that lead to uterine healthy women (Macklon et al. 2002). Previous studies receptivity are shared by both species, suggesting that estimate that for each menstrual cycle in a healthy woman, many of the key molecular signaling mechanisms which the chance for natural conception is only 30% (Zinaman mediate progesterone (and estrogen) control of these et al. 1996, Wilcox et al. 1999), and the low conception developmental steps are also conserved (Lim & Wang rate is due in large part to implantation failure. While 2010). advancements in our understanding of oocyte and embryo development have significantly improved pregnancy Endometrial receptivity and decidualization success rates using assisted reproductive technologies require progesterone (ARTs), periimplantation failure due to a nonreceptive endometrium continues to limit the full potential of these Prior to embryo implantation in the mouse, proliferation technologies (Blesa et al. 2014). Moreover, implantation of the endometrial luminal epithelium occurs in response failure is frequently implicated in recurrent pregnancy to elevating serum levels of preovulatory estrogen (Cha loss after parental chromosomal abnormalities, maternal et al. 2012b, Namiki et al. 2018). Following ovulation, thrombophilic disorders, and anatomical uterine defects fertilization, and progressive development of the resultant are first eliminated as causal factors Rai( & Regan 2006). zygote to the activated blastocyst-stage with its two distinct In addition to the clear clinical challenges, such early cell lineages (the inner cell mass and outer trophectoderm), pregnancy failures contribute to serious emotional distress rising levels of systemic progesterone synthesized and

https://jme.bioscientifica.com © 2020 Society for Endocrinology https://doi.org/10.1530/JME-19-0212 Published by Bioscientifica Ltd. Printed in Great Britain Downloaded from Bioscientifica.com at 09/23/2021 01:12:14PM via free access Journal of Molecular F J DeMayo and J P Lyson Progesterone signaling in the 65:1 T5 Endocrinology endometrium secreted from newly formed ovarian corpora lutea result invasion, which ultimately establishes the hemochorial in endometrial stromal cell proliferation, which is further and endotheliochorial placenta in the mouse and human augmented by a small nidatory spike in estrogen levels. respectively. Importantly, uNK cells curtail excessive In parallel, progesterone suppresses estrogen-induced trophectodermal invasion into the uterine compartment proliferation of the endometrial epithelia, which results by triggering apoptosis of the advancing trophoblastic in the transition of the epithelium from a proliferative to cellular front of the invading conceptus. While the a differentiative state to become temporarily permissive to mouse requires the conceptus or an artificial stimulus to embryo attachment and invasion. In response to embryo trigger endometrial stromal decidualization, the human attachment (’the attachment reaction’), underlying endometrial stromal compartment can decidualize during fibroblastic stromal cells undergo extensive localized the progesterone-dominant early secretory phase of a progesterone-dependent proliferation and differentiation non-conception cycle (Gellersen & Brosens 2014). Also, to become polygonal epithelioid decidual cells, a cellular cultured human endometrial stromal cells require only transformation process termed decidualization. As estrogen, progesterone, and cAMP to decidualize in vitro endometrial decidualization manifests, luminal epithelial (Brosens et al. 1999). cells undergo entosis at the implantation site thereby Progesterone-dependent uterine receptivity and allowing the blastocyst’s trophoblastic cells to breach the decidualization represent critical reproductive events underlying basal lamina and invade the decidualizing along a tightly choreographed procession of uterine stromal compartment of the endometrium. Early cellular changes in which the successful execution of studies by Lydon and colleagues using the Pgr knockout one event is predicated on the successful completion of (PRKO) mouse model, in which both Pgr isoforms were events that occured before. Because the establishment simultaneously ablated (Lydon et al. 1995), underscored of the fetoplacental interface is inextricably linked to the indispensability of nuclear receptor-mediated successful decidualization, defective decidualization progesterone signaling in the development of the receptive has been causally connected to a number of obstetric and decidualized uterus. Subsequent studies using Pgr disorders, such as early fetal miscarriage due to placental isoform specific knockout mice demonstrated that PGR-A insufficiency, placenta accreta, preeclampsia, fetal growth alone – and not PGR-B or PGR isoform heterodimerization restriction, and preterm birth (Cha et al. 2012b). – is required for murine pregnancy establishment and Although progesterone’s pivotal role as an apex maintenance (Mulac-Jericevic et al. 2000). In humans, hormone regulator of uterine peri-implantation biology however, both PGR isoforms are important for pregnancy is well recognized, our fundamental understanding of the success, with the PGR-B isoform playing a predominant nuclear receptor-mediated signaling mechanisms by which role in decidualization (Kaya et al. 2015). progesterone exerts its action in the endometrium that Surrounding the invading conceptus, decidual cells lead to pregnancy establishment is only now emerging. furnish histotrophic nutrition, an immunotolerant Without question, the ongoing efforts to identify the key microenvironment, a negative selection process for non- molecular mediators and modifiers of uterine progesterone viable embryos, and protection against physiological action will provide deeper mechanistic insight into early stressors as well as excessive invasion by the embryo into pregnancy establishment and uncover new molecular the uterine compartment (Gellersen & Brosens 2014). signals that may be used in novel and more efficient Progesterone-driven decidualization is accompanied clinical solutions to address periimplantation failure, by angiogenesis and the influx of leukocytes, such gestational complications that occur in later trimesters, as uterine natural killer (uNK) cells and monocytes. and common reproductive clinicopathologies arising due Furthermore, molecular studies provide strong support to defective responsiveness to progesterone. for progesterone induction of interleukin 15 (and its receptor) in endometrial stromal cells as an essential trans- Molecular mediators and modifiers of uterine presentation signal in the differentiation of immature progesterone receptor action uNK cells to mature cells (Okada et al. 2000, Wilkens et al. 2013). Concentrated around maternal spiral arterioles, Just as signaling crosstalk between the blastocyst and uterine uNK cells are important innate immune cells that compartment is essential for embryo implantation, so too promote an immunosuppressive milieu for the invading is the reciprocal communication between the epithelial hemiallogeneic conceptus and promote remodeling of and stromal cellular compartments of the endometrium spiral arterioles to facilitate endovascular trophoblast critical for the development of uterine receptivity and

The Indian hedgehog-signaling axis is a pivotal PGR molecular mediator in the endometrium

Separate investigations disclosed that Indian hedgehog (Ihh) is transcriptionally induced by progesterone in the luminal epithelium of the murine endometrium, just prior to embryo implantation (Matsumoto et al. 2002, Takamoto et al. 2002). Later cistromic studies would demonstrate that Ihh is a direct molecular target of PGR (Wang et al. 2018). A secreted morphogen, IHH is a member of the highly conserved hedgehog family (Ng & Curran 2011), which includes sonic and desert hedgehog. The hedgehog triad of diffusible morphogens regulates cell proliferation, differentiation, and short-range cell-cell communication, cellular processes that are essential for organogenesis, tissue homeostasis, and oncogenesis. Investigations on the Figure 2 PRKO and a conditional Ihh-knockout mouse respectively A propensity for complexity: molecular signaling required for progesterone-dependent uterine receptivity and decidualization. (A) The demonstrated that progesterone induction of Ihh was progesterone-PGR-IHH-COUP-TFII signaling pathway, which spans the via the PGR (Takamoto et al. 2002), and that IHH was epithelial and stromal cellular compartments of the endometrium, essential for uterine receptivity and decidualization (Lee controls ESR1 activity in the endometrial epithelium. Such ESR1 control is required for epithelial differentiation and development of the receptive et al. 2006). Absence of Ihh in the murine endometrium endometrium for embryo implantation. For clarity, other signals (i.e. Msx resulted in derailed cell-cycle progression, significantly 1 and 2), which are important for uterine receptivity, are not be included decreased EGF signaling, and the display of overt in this schematic. (B) A selection of progesterone-induced signaling pathways required for endometrial stromal cell decidualization during the histological hallmarks of a persistent estrogenized uterus periimplantation period. Adapted with permission in modified form from (i.e. the presence of numerous cystic glandular ducts and Wu et al. (2018). a hylalinized stroma) (Lee et al. 2006, Franco et al. 2010). Crossing the epithelial-stromal divide as a paracrine and stromal cell decidualization (Kurihara et al. 2007, Lee secreted factor, epithelial-derived IHH activates the et al. 2010), which is caused by a persistent estrogenized canonical hedgehog effector pathway in the underlying uterus (Kurihara et al. 2007, Lee et al. 2010). Of clinical stroma (Fig. 2A); the hedgehog pathway includes the significance, aberrant downregulation of bothIHH and cognate IHH receptor patched-1 (PTCH1), the activated COUP-TFII in the human endometrium is linked to intracellular transducer: Smoothened (SMO) and the endometriosis (Lin et al. 2014), a uterine pathology that glioma-associated oncogene homolog (GLI) transcription is dependent in part on a heightened estrogenic response. factors (Fig. 2A). Activation of the hedgehog pathway Recent clinical data also reveal that selective progesterone promotes the expression of the orphan nuclear receptor modulators can significantly perturb the normal receptor: chicken ovalbumin upstream promoter expression levels of these progesterone-responsive factors transcription factor II (COUP-TFII; NR2F2) in the stroma in the human endometrium (Whitaker et al. 2017). (Takamoto et al. 2002) (Fig. 2A). A key mesenchymal Enhanced COUP-TFII expression by the PGR-IHH differentiation factor, COUP-TFII modulates a myriad axis is associated with increased stromal PGR expression of cellular processes from angiogenesis, organogenesis, as well as the induction of stromal heart and neural inflammation, and metabolism to cell adhesion and cell- crest derivatives expressed transcript 2 (Hand2) (Li et al. fate specification (Wu et al. 2016). Conditional ablation 2011), a basic helix-loop-helix transcription factor. of Coup-tfII results in a block in embryo implantation While it is not clear whether COUP-TFII directly or

https://jme.bioscientifica.com © 2020 Society for Endocrinology https://doi.org/10.1530/JME-19-0212 Published by Bioscientifica Ltd. Printed in Great Britain Downloaded from Bioscientifica.com at 09/23/2021 01:12:14PM via free access Journal of Molecular F J DeMayo and J P Lyson Progesterone signaling in the 65:1 T7 Endocrinology endometrium indirectly (i.e. through stromal PGR) increases stromal been shown to be important for epithelial-stromal Hand2 levels, HAND2 normally inhibits the expression crosstalk required for the elaboration of the receptive of several stromal fibroblast growth factor (fgf) family uterus (Daikoku et al. 2011, Nallasamy et al. 2012). In members (fgf1, -2, -9, and -18) in the endometrium (Li parallel, enhanced stromal EGFR-mediated signaling et al. 2011). Accordingly, molecular phenotyping of the phosphorylates and activates the serine/threonine protein conditional Hand2 knockout mouse demonstrated that kinase: With-No-Lysine (K) 1 (WNK1) (Large et al. 2014), absence of Hand2 results in constitutive induction of a kinase which is essential for decidualization of human stromal fgf expression in the endometrium (Li et al. 2011). endometrial stromal cells in culture (Adams et al. 2017). As paracrine signals within the normal endometrium, In a remarkably short period of time, the confluence stromal FGFs engage their transmembrane tyrosine of independent molecular investigations have furnished kinase receptors (FGFRs with associated docking factors) invaluable insights into the tremendous molecular located in epithelial cells to trigger phosphorylation (and complexity that underlies progesterone regulation activation) of extracellular signal regulated kinases 1 and of endometrial receptivity and decidualization. At a 2 (ERK1/2), which in turn phosphorylate, stabilize, and minimum, the innumerable genes, pathways, and activate the ESR1 (Li et al. 2011). Apart from epithelial networks identified to date are merely a prologue for the proliferation, activated ESR1 maintains expression of immense molecular complexity that is to come. Given the Muc1 (MUC1), a glycoprotein expressed on the apical predicted scale and complexity of the multicomponent surface of luminal epithelial cells that prevents embryo molecular networks that mediate progesterone control attachment (Surveyor et al. 1995). Moreover, persistent of uterine receptivity and decidualization, an immediate proliferation of the glandular epithelium results in a challenge will be to delineate the molecular mechanisms block in the expression of the leukemia inhibitory factor by which these various signal transduction pathways ((LIF) an interleukin 6 family cytokine member) and are spatiotemporally integrated and coordinated in a the Forkhead box A1 (Fox A1) transcription factor, both hierarchical manner. As detailed previously, many of these pivotal signaling cues for embryo implantation (Stewart progesterone mediators are critical for endometrial stromal et al. 1992, Kelleher et al. 2017). Therefore, uncoupling of cell decidualization, a type of mesenchymal-epithelial the progestero ne-PG R-IHH -COUP -TFII -HAND 2 regulatory transformation (MET) cellular process. However, which axis can cause persistent activation of epithelial ESR1 (Fig. factors are also important for other cellular processes that 2A), which results in failure of the luminal epithelial cell are required for successful stromal decidualization – that to exit the cell cycle and undergo differentiation, the is, angiogenesis, local immunosuppression, and immune net result of which is to prevent the development of the cell influx – remain an open question. Because premature receptive state. senescence of decidual cells is an established cause of Uncovering the complex molecular circuitries that preterm birth (Cha et al. 2012a), another open question mediate progesterone-driven endometrial receptivity also is whether all or a subset of these factors are continually revealed how uterine receptivity is tightly coordinated required to maintain healthy decidual tissue throughout with progesterone-dependent endometrial decidualization gestation. While much remains unknown, expanding our at the molecular level. For example, induction of stromal knowledge in this area will undoubtedly provide a broader COUP-TFII by the PGR-IHH axis results in the induction conceptual foundation for understanding progesterone of bone morphogenetic protein 2 (Bmp2) (Kurihara et al. action not only in normal endometrial function but also 2007), a member of the transforming growth factor beta in common gynecologic morbidities in which normal (TGFβ) superfamily of cytokines. As a consequence of progesterone responsiveness is markedly attenuated or elevated Bmp2 levels, wingless-type MMTV integration site abrogated (Al-Sabbagh et al. 2012). (WNT) family member 4 (Wnt 4) is induced in the stroma. Both BMP2 and WNT 4 – along with COUP-TFII and IHH Modifiers of endometrial PGR action – are essential for PGR-dependent endometrial stromal cell decidualization in both human and mouse (Lee et al. Loosely defined, transcriptional modifiers of PGR are 2007, Li et al. 2007, 2013, Franco et al. 2011) (Fig. 2B). This factors that regulate the transcriptional output mediated regulatory complexity is further increased by the finding by ligand-activated PGR through control of PGR levels and/ that BMP2 upregulates members of the muscle segment or by participating directly or indirectly as a coregulator homeobox (Msx) family of transcription factors (Msx1 in PGR-containing transcriptional complexes that and 2) (Nallasamy et al. 2019), which have previously govern target gene expression. Modifier control of PGR

https://jme.bioscientifica.com © 2020 Society for Endocrinology https://doi.org/10.1530/JME-19-0212 Published by Bioscientifica Ltd. Printed in Great Britain Downloaded from Bioscientifica.com at 09/23/2021 01:12:14PM via free access Journal of Molecular F J DeMayo and J P Lyson Progesterone signaling in the 65:1 T8 Endocrinology endometrium functionality operates in tandem with PGR PTMs, another both human and mouse uterine datasets highlighted regulatory process that modifies PGR transcriptional the existence of a conserved GATA2-PGR regulatory activity and turnover (Fig. 1). Recent transcriptomic network that relies on a cooperative relationship between and cistromic analyses together with follow-up in vivo GATA2 and PGR (as well as with other transcription validation in engineered mouse models have identified factors important for uterine receptivity (i.e. SOX17 (see crucial modifiers of uterine PGR activity that modulate below)) in controlling normal endometrial epithelial the levels and/or the transcriptional activity of PGR in the transcriptional responsiveness to progesterone during endometrium. the periimplantation period. As a prototypic example, chromatin co-occupancy of GATA2 and PGR has been mapped to a putative enhancer located 19 kb upstream The GATA2 transcription factor is a master of the promoter controlling Ihh transcription (Rubel et al. modifier of PGR expression and transcriptional 2016), which is required for uterine receptivity. Note: Ihh activity in the uterine epithelium is not induced by progesterone in the murine uterine Through their highly conserved dual zinc finger domains, epithelium in the absence of GATA2 (Rubel et al. 2016). the six member GATA-binding transcription factor family Similar to findings from studies on the androgen bind the evolutionary conserved consensus sequence receptor ((AR); a close relative of PGR) in the prostate (Wu 5′-(A/T)GATA(A/G)-3′ to control diverse physiological et al. 2014), data support epithelial GATA2 serving as a processes from hematopoiesis, adipocyte differentiation pioneer factor as well as a coregulator of PGR target gene to pituitary function (Tremblay et al. 2018). To execute expression in the uterine epithelium (Rubel et al. 2016) (Fig. these processes at the molecular level, GATA transcription 2A). Indeed, extensive studies on the prostate demonstrate factors can act as ‘pioneer factors’ endowed with special that GATA2 can: (a.) bind upstream regulatory elements properties that allow for their binding to heterochromatin to increase AR expression; (b.) establish chromatin to promote – through recruitment of epigenetic modifiers accessibility at AR enhancer regions through recruitment – chromatin opening. In turn, open chromatin facilitates of p300 histone acetyltransferase that creates active local combinatorial assembly at a promoter or an enhancer chromatin by acetylating lysine 27 in histone 3 (H3K27); of other transcriptional regulators (i.e. coregulators and and (c.) generate and maintain regulatory chromatin tissue-specific transcription factors) that ultimately loops between AR-bound distal enhancers and AR target modulate target gene-expression programs. gene promoters through enlistment of the mediator Expression studies in the murine uterus revealed that coregulator complex (MED1) (Wu et al. 2014). Whether the temporal expression profile of theGata2 transcription GATA2 modifiesPgr expression and its signaling program factor and Pgr is similar in the epithelium, in which through similar mechanisms in the uterine epithelium expression levels for both transcription factors peak just awaits further investigation. What governs GATA2 before and precipitously decline immediately after entry expression in the uterine epithelium and whether there into the window of receptivity (Rubel et al. 2012a). That are fundamental commonalities between the GATA2-PGR GATA2 and PGR might be mechanistically linked is also network in the uterus and GATA regulatory networks supported by similar infertility phenotypes – a block in operative in other physiological systems also remain open uterine receptivity and decidualization – exhibited by questions. female mice in which the uterine epithelium is selectively devoid of Gata2 or Pgr (Franco et al. 2012, Rubel et al. A potent coregulator of uterine PGR action: SOX17 2016). Abrogation of Gata2 also results in uterine epithelial signatures of unopposed estrogen signaling Mining the PGR cistrome dataset from the mouse uterus (Rubel et al. 2016), such as aberrant epithelial integrity and (Rubel et al. 2012b), the SOX17 transcription factor was stratification. Further mouse phenotypic investigations, quickly identified as a putative direct target of PGR and in conjunction with data sourced from genome-wide GATA2 and possibly important for mediating progesterone- chromatin occupancy and transcriptomic analytics of dependent uterine receptivity. Initial in vivo support for murine and human uterine tissue, revealed that GATA2 is this proposal came from the observation that female responsible for the expression of epithelial PGR as well as mice with whole-body Sox17 haploinsufficiency display the ability of PGR to activate its downstream transcriptional a severe subfertility defect due to impaired implantation program that drives uterine receptivity (Rubel et al. 2016). (Hirate et al. 2016). Subsequent investigations on mice In particular, gene signature and pathway analysis of with Sox17 selectively ablated in PGR positive cells of the

https://jme.bioscientifica.com © 2020 Society for Endocrinology https://doi.org/10.1530/JME-19-0212 Published by Bioscientifica Ltd. Printed in Great Britain Downloaded from Bioscientifica.com at 09/23/2021 01:12:14PM via free access Journal of Molecular F J DeMayo and J P Lyson Progesterone signaling in the 65:1 T9 Endocrinology endometrium uterus or in the uterine epithelium confirmed not only The PGR and FOXO1 relationship: the essential role of epithelial SOX17 in the development a ‘Yin-Yang’ dynamic of the receptive endometrium but also the requirement for this transcription factor in postnatal endometrial A member of the O-class of Forkhead box (FOX) proteins, gland development (adenogenesis) (Guimaraes-Young FOXO1 is critical for a broad spectrum of fundamental et al. 2016, Wang et al. 2018). cellular processes, including metabolism, oxidative At least 20 members comprise the SRY-determining stress avoidance, cell proliferation, and apoptosis (Link region Y-related high-mobility group (HMG) box 2019). Conditional Foxo1 knockout (Foxo1d/d) mouse (SOX) family of transcription factors, which are critical studies demonstrated that FOXO1 is essential for embryo determinants of cell-fate reprogramming by pioneering implantation (Vasquez et al. 2018). Absence of FOXO1 in epigenetic remodeling of the genome (Grimm et al. 2019). the uterine epithelium resulted in an aberrant alteration Mechanistically, SOX transcription factors can bind DNA in epithelial cell polarity and an impaired apoptotic through the HMG domain to cause DNA to bend over long program that blocked embryo invasion through the distances in the genome that enables the juxtaposition luminal epithelial compartment (Vasquez et al. 2018). of enhancer bound factors to interact with members of Transcriptomic analysis revealed that epithelial FOXO1 the transcriptional holocomplex at target gene promoters. regulates expression programs associated with cell Accordingly, the fundamental regulatory functions of invasion, molecular transport, apoptosis, β-catenin SOX family members (including SOX17) encompass signaling, and an increase in PGR signaling (Vasquez the specification and morphogenesis of the primitive et al. 2018). Intriguingly, the enhanced PGR signaling and definitive endoderm, the ontogenesis of germ persisting in the endometrial epithelium of the Foxo1d/d cells, maintenance of fetal and neonatal hematopoietic mouse is reflected by the constitutive expression of PGR stem cells, and the development and function of the during the window of receptivity; note: previous studies cardiovascular system. showed that epithelial PGR levels must markedly decrease Comparative analyses of transcription factor cistrome to enable normal uterine receptivity to occur (Wetendorf datasets from the mouse uterus disclosed a discrete et al. 2017) (Fig. 3). That there may be a mutually exclusive regulatory element containing a cluster of binding sites or ‘Yin-Yang’ reciprocal relationship between FOXO1 for PGR, GATA2, FOXA2, and SOX17 (Wang et al. 2018). and PGR expression in the murine uterine epithelium is Importantly, this cis regulatory element is associated with further supported by the absence of FOXO1 expression in over 700 uterine genes, many of which are important for the uterine epithelium of mice engineered to continually uterine epithelial function (Wang et al. 2018). As described express epithelial PGR (Wetendorf et al. 2017). Importantly, previously, this cis-regulatory element was identified 19 kb this Yin-Yang relationship between FOXO1 and PGR from the promoter of the Ihh gene, a gene that is critical was observed in the glandular epithelium of human for progesterone-dependent uterine receptivity (Fig. 2A). endometrial tissue biopsied during the proliferative and As a testament to the singular importance of SOX17 as a secretory phase of the cycle (Vasquez et al. 2018). These coregulator of PGR driven Ihh gene transcription, Crispr/ studies showed that while FOXO1 is not expressed in Cas9-mediated in vivo deletion of the SOX17 binding the glandular epithelium during the proliferative and site within this element renders the murine endometrial late-secretory phase of the cycle, FOXO1 is strongly epithelium incapable of exiting the cell cycle to enter expressed in the glandular epithelial compartment during the differentiation program that is necessary for uterine the mid-secretory phase of the cycle (considered the receptivity (Wang et al. 2018). Collectively, these studies window of receptivity). In contrast, glandular epithelial underscore a pivotal role for SOX17 in uterine epithelial PGR expression is strongest during proliferative phase transcriptional responses required for progesterone of the cycle and undetectable during the cycle’s mid- controlled uterine receptivity. To be addressed by future secretory phase. Interestingly, this reciprocal expression studies is the cellular and molecular mechanisms by relationship between PGR and FOXO1 in the glandular which SOX17 regulates postnatal adenogenesis, the role epithelium of the human endometrium was also reported of SOX17 in the uterine vasculature during the embryo by the Critchley group (Whitaker et al. 2017). Together, implantation process, and the involvement SOX17 as a these findings support the proposal that the gain and loss mutated cancer driver gene in endometrial carcinomas of FOXO1 and PGR expression respectively are interrelated (Senna Tan et al. 2019). and that the expression switch of these two transcription

cells exhibiting replication stress through programming these cells for senescence and ultimately for uNK mediated clearance (Brighton et al. 2017). It should be noted that there are numerous important PGR modifiers – that is, the FKBP52 immunophilin (Tranguch et al. 2005) and the steroid receptor coactivator (SRC) 2 (Kommagani et al. 2013) just to name two – of endometrial PGR action that could not be described here due to space limitations.

Figure 3 Progesterone receptor and FOXO1 display a ‘Yin-Yang’ expression relationship in the epithelium of the murine endometrium during the Perspectives window of implantation. (A) Robust luminal epithelial (LE) expression of FOXO 1 (brown arrowhead) in the uterus of a control mouse (Foxo1f/f) at Engineering the murine Pgr gene as a null allele (Lydon day 4.5 of pregnancy (the morning of vaginal plug detection is designated et al. 1995), a floxed allele (Fernandez-Valdivia et al. 2010), pregnancy day 0.5). (B) Serial section of uterine tissue shown in (A) stained or a cre driver (Soyal et al. 2005) has significantly expanded for PGR. As expected (Wetendorf et al. 2017), note the absence of PGR expression in the luminal epithelium (blue arrowhead) with marked our understanding of uterine PGR action in vivo. With the expression in the stroma. (C) Staining for FOXO1 expression in the uterus advent of precise Crispr/Cas9 gene editing (Doudna & of a Foxo1d/d mouse at 4.5 days following the detection of the vaginal plug. Charpentier 2014), high-resolution structure functional Unlike control uterus (A), note the clear absence of FOXO1 expression in the luminal epithelium (blue arrowhead) in the Foxo1d/d uterus. (D) analysis is now possible in vivo not only for the PGR but Staining for PGR expression in a serial section of uterine tissue shown in also for its mediators and modulators. Application of (C) reveals abnormal constitutive PGR expression in the luminal this fast and effective mouse engineering methodology epithelium (brown arrowhead) in the Foxo1d/d uterus. (E) The immunohistochemical findings shown in A, B, C, and D are schematically is predicted to allow expeditious functional validation summarized as a proposed model for the suppression of PGR expression of PTM events on the PGR in an in vivo context. Also, in the endometrial luminal epithelium by FOXO1 within the window of this technological advance will provide opportunities to receptivity. The immunohistochemical data shown in panels A, B, C, and D and the modified schematic model shown in (E) are presented in adapted elucidate the functional importance in vivo of defined form from Vasquez et al. (2018). protein-protein interaction sites on PGR and its modifiers within a transcriptional complex. In addition to protein factors represents an evolutionary, conserved, and pivotal structure-function analysis, this mouse technology will be determinant of endometrial receptivity. invaluable in validating the in vivo functional relevance A critical question to be addressed in the future is: of cis-regulatory elements within enhancers or promoters what is the regulatory mechanism(s) that orchestrated the of predicted target genes of PGR or of its mediators and expression switch of FOXO1 and PGR in the endometrial modifiers; the first example of applying this approach epithelium just prior to the window of receptivity? Also, in uterine studies was recently published (Wang et al. previous in vitro studies using cultured human endometrial 2018). In addition to the mouse, this powerful and stromal cells revealed an important coregulator role versatile gene-editing tool will enable broad application for FOXO1 in progesterone-dependent transcriptional of precise genome manipulation in other experimental programs necessary for human endometrial stromal models of uterine biology and pathobiology, such as the cell decidualization (Vasquez et al. 2015). Although the simple monolayer cell-culture model, the more recent mouse studies did not provide support for FOXO1 in the 3D organoid cell-culture system, and animal models that initiation of progesterone-dependent endometrial stromal are closer to human on the evolutionary scale. Especially cell decidualization in vivo (Vasquez et al. 2018), these noteworthy are the technological advances in 3D organoid studies did not rule out a role for FOXO1 in maintaining cultures of human endometrium that can recapitulate a normal decidual response over time. In the future, it – in response to hormone – the dynamic endometrial will be interesting to address whether continued FOXO1 cellular and molecular changes that occur throughout the expression is necessary to prevent premature senescence menstrual cycle and early pregnancy (Turco et al. 2017). of the decidua as decidual premature senescence causes From a clinical perspective, this powerful cell-culture adverse gestational outcomes in later trimesters (Cha et al. approach in combination with Crispr/Cas9 gene editing is 2012a,b). In relation, recent studies revealed a key role predicted to accelerate and expand our understanding of for FOXO1 in maintaining a healthy human decidual cell endometrial PGR signaling in human uterine receptivity population by removing a subset of endometrial stromal and decidualization that is free from ethical constraints.

To date, the use of engineered mouse models, human Acknowledgements biopsy tissue, and primary cells in conjunction with the The authors apologize in advance to investigators whose exceptional analysis and integration of genome-wide transcriptome research did not get cited due to space limitations set by the journal. and cistrome datasets have now exposed to molecular dissection the complex signaling cascades – comprising transcription factors, growth factors, morphogens, and References cytokines – that underpin progesterone-driven uterine Adams NR, Vasquez YM, Mo Q, Gibbons W, Kovanci E & DeMayo FJ receptivity and decidualization. Due to space limitations, 2017 WNK lysine deficient protein kinase 1 regulates human this review showcased only a select number of molecular endometrial stromal cell decidualization, proliferation, and migration in part through mitogen-activated protein kinase 7. Biology of mediators and modifiers along the chain of signal Reproduction 97 400–412. (https://doi.org/10.1093/biolre/iox108) transmission that starts with initial progesterone exposure Allen WM 1974 Recollections of my life with progesterone. Gynecologic and ends with the development of the uterine receptive Investigation 5 142–182. (https://doi.org/10.1159/000301649) Allen WM 2005 My life with progesterone∗. American Journal of state. Going forward, a formidable challenge will be to Obstetrics and Gynecology 193 1575–1577. (https://doi.org/10.1016/j. understand how these numerous progesterone-responsive ajog.2005.06.007) genes, pathways, and networks operate dynamically and Allen WM & Corner GW 1929 Physiology of the corpus luteum, III: normal growth and implantation of embryos after early ablation of spatially in concert with signaling networks governed the ovaries, and under the influence of extracts of the corpus by other extrinsic cues such as systemic estrogen. luteum. American Journal of Physiology: Legacy Content 88 340–346. Meeting this challenge will be a clinical imperative, as (https://doi.org/10.1152/ajplegacy.1929.88.2.340) Allen WM, Butenandt A, Corner GW & Slotta KH 1935 Nomenclature of derailment of the majority of mediators and modifiers of corpus luteum hormone. Science 82 153. (https://doi.org/10.1126/ progesterone signaling lead to a persistent estrogenized science.82.2120.153-a) uterus, an abnormal uterine state that (if persistent) can Al-Sabbagh M, Lam EW & Brosens JJ 2012 Mechanisms of endometrial progesterone resistance. Molecular and Cellular Endocrinology 358 cause not only infertility but also the etiopathogenesis 208–215. (https://doi.org/10.1016/j.mce.2011.10.035) of common uterine morbidities, such as endometriosis, Bain DL, Franden MA, McManaman JL, Takimoto GS & Horwitz KB endometrial hyperplasia, and cancer (Al-Sabbagh et al. 2000 The N-terminal region of the human progesterone A-receptor. Structural analysis and the influence of the DNA binding domain. 2012). Journal of Biological Chemistry 275 7313–7320. (https://doi. In conclusion, our review ends as it began, with Willard org/10.1074/jbc.275.10.7313) Myron Allen. Toward the end of his scientific career, Blesa D, Ruiz-Alonso M & Simon C 2014 Clinical management of endometrial receptivity. Seminars in Reproductive Medicine 32 Willard Myron Allen authored a brief career retrospective, 410–413. (https://doi.org/10.1055/s-0034-1376360) entitled ‘Recollections of My Life with Progesterone’ (Allen Brighton PJ, Maruyama Y, Fishwick K, Vrljicak P, Tewary S, Fujihara R, 1974). Within the concluding paragraph, Allen wrote: Muter J, Lucas ES, Yamada T, Woods L, et al. 2017 Clearance of senescent decidual cells by uterine natural killer cells in cycling ‘These were golden years for both of us (referring to Allen and human endometrium. eLife 6 e31274. (https://doi.org/10.7554/ Corner). For a few fleeting years progesterone was ours. My eLife.31274) story ends on a happy note; there are still unsolved problems Brosens JJ, Hayashi N & White JO 1999 Progesterone receptor regulates decidual prolactin expression in differentiating human endometrial relating to progesterone’. Written many decades ago, the stromal cells. Endocrinology 140 4809–4820. (https://doi.org/10.1210/ concluding line is prescient as it is understated. endo.140.10.7070) Cha J, Hirota Y & Dey SK 2012a Sensing senescence in preterm birth. Cell Cycle 11 205–206. (https://doi.org/10.4161/cc.11.2.18781) Cha J, Sun X & Dey SK 2012b Mechanisms of implantation: strategies for successful pregnancy. Nature Medicine 18 1754–1767. (https://doi. Declaration of interest org/10.1038/nm.3012) The authors declare that there is no conflict of interest that could be Conneely OM, Sullivan WP, Toft DO, Birnbaumer M, Cook RG, perceived as prejudicing the impartiality of this review. Maxwell BL, Zarucki-Schulz T, Greene GL, Schrader WT & O’Malley BW 1986 Molecular cloning of the chicken progesterone receptor. Science 233 767–770. (https://doi.org/10.1126/ science.2426779) Funding Corner GW & Allen WM 1929 Physiology of the corpus luteum, II: Supported in part by the Intramural Research Program of the National production of a special uterine reaction (progestational proliferation) Institute of Environmental Health Sciences: Project Z1AES103311-01 (F J D) by extracts of the corpus luteum. American Journal of Physiology: and by National Institutes of Health/NICHD: R01HD042311 (J P L). Legacy Content 88 326–339. (https://doi.org/10.1152/ ajplegacy.1929.88.2.326) Daikoku T, Cha J, Sun X, Tranguch S, Xie H, Fujita T, Hirota Y, Lydon J, DeMayo F, Maxson R, et al. 2011 Conditional deletion of Msx Author contribution statement homeobox genes in the uterus inhibits blastocyst implantation by Drs DeMayo and Lydon contributed equally to the preparation of this altering uterine receptivity. Developmental Cell 21 1014–1025. review. (https://doi.org/10.1016/j.devcel.2011.09.010)

Diep CH, Daniel AR, Mauro LJ, Knutson TP & Lange CA 2015 Jensen EV 1962 On the mechanism of estrogen action. Perspectives in Progesterone action in breast, uterine, and ovarian cancers. Journal of Biology and Medicine 6 47–59. (https://doi.org/10.1353/pbm.1963.0005) Molecular Endocrinology 54 R31–R53. (https://doi.org/10.1530/JME-14- Jensen EV, Suzuki T, Kawashima T, Stumpf WE, Jungblut PW & 0252) DeSombre ER 1968 A two-step mechanism for the interaction of Doudna JA & Charpentier E 2014 Genome editing. The new frontier of estradiol with rat uterus. PNAS 59 632–638. (https://doi.org/10.1073/ genome engineering with CRISPR-Cas9. Science 346 1258096. pnas.59.2.632) (https://doi.org/10.1126/science.1258096) Kastner P, Krust A, Turcotte B, Stropp U, Tora L, Gronemeyer H & Enders AC 1976 Anatomical aspects of implantation. Journal of Chambon P 1990 Two distinct estrogen-regulated promoters Reproduction and Fertility. Supplement 1–15. generate transcripts encoding the two functionally different human Fernandez-Valdivia R, Jeong J, Mukherjee A, Soyal SM, Li J, Ying Y, progesterone receptor forms A and B. EMBO Journal 9 1603–1614. Demayo FJ & Lydon JP 2010 A mouse model to dissect (https://doi.org/10.1002/j.1460-2075.1990.tb08280.x) progesterone signaling in the female reproductive tract and Kaya HS, Hantak AM, Stubbs LJ, Taylor RN, Bagchi IC & Bagchi MK mammary gland. Genesis 48 106–113. (https://doi.org/10.1002/ 2015 Roles of progesterone receptor A and B isoforms during human dvg.20586) endometrial decidualization. Molecular Endocrinology 29 882–895. Franco HL, Lee KY, Broaddus RR, White LD, Lanske B, Lydon JP, (https://doi.org/10.1210/me.2014-1363) Jeong JW & DeMayo FJ 2010 Ablation of Indian hedgehog in the Kelleher AM, Peng W, Pru JK, Pru CA, DeMayo FJ & Spencer TE 2017 murine uterus results in decreased cell cycle progression, aberrant Forkhead box a2 (FOXA2) is essential for uterine function and fertility. epidermal growth factor signaling, and increased estrogen signaling. PNAS 114 E1018–E1026. (https://doi.org/10.1073/pnas.1618433114) Biology of Reproduction 82 783–790. (https://doi.org/10.1095/ Kommagani R, Szwarc MM, Kovanci E, Gibbons WE, Putluri N, Maity S, biolreprod.109.080259) Creighton CJ, Sreekumar A, DeMayo FJ, Lydon JP, et al. 2013 Franco HL, Dai D, Lee KY, Rubel CA, Roop D, Boerboom D, Jeong JW, Acceleration of the glycolytic flux by steroid receptor coactivator-2 is Lydon JP, Bagchi IC, Bagchi MK, et al. 2011 WNT4 is a key regulator essential for endometrial decidualization. PLoS Genetics 9 e1003900. of normal postnatal uterine development and progesterone (https://doi.org/10.1371/journal.pgen.1003900) signaling during embryo implantation and decidualization in the Kurihara I, Lee DK, Petit FG, Jeong J, Lee K, Lydon JP, DeMayo FJ, mouse. FASEB Journal 25 1176–1187. (https://doi.org/10.1096/fj.10- Tsai MJ & Tsai SY 2007 COUP-TFII mediates progesterone regulation 175349) of uterine implantation by controlling ER activity. PLoS Genetics 3 Franco HL, Rubel CA, Large MJ, Wetendorf M, Fernandez-Valdivia R, e102. (https://doi.org/10.1371/journal.pgen.0030102) Jeong JW, Spencer TE, Behringer RR, Lydon JP & Demayo FJ 2012 Large MJ, Wetendorf M, Lanz RB, Hartig SM, Creighton CJ, Mancini MA, Epithelial progesterone receptor exhibits pleiotropic roles in uterine Kovanci E, Lee KF, Threadgill DW, Lydon JP, et al. 2014 The development and function. FASEB Journal 26 1218–1227. (https:// epidermal growth factor receptor critically regulates endometrial doi.org/10.1096/fj.11-193334) function during early pregnancy. PLoS Genetics 10 e1004451. Gellersen B & Brosens JJ 2014 Cyclic decidualization of the human (https://doi.org/10.1371/journal.pgen.1004451) endometrium in reproductive health and failure. Endocrine Reviews Le Dily F, Vidal E, Cuartero Y, Quilez J, Nacht AS, Vicent GP, Carbonell- 35 851–905. (https://doi.org/10.1210/er.2014-1045) Caballero J, Sharma P, Villanueva-Canas JL, Ferrari R, et al. 2019 Graham JD & Clarke CL 1997 Physiological action of progesterone in Hormone-control regions mediate steroid receptor-dependent target tissues. Endocrine Reviews 18 502–519. (https://doi. genome organization. Genome Research 29 29–39. (https://doi. org/10.1210/edrv.18.4.0308) org/10.1101/gr.243824.118) Grimm SL, Hartig SM & Edwards DP 2016 Progesterone receptor Lee K, Jeong J, Kwak I, Yu CT, Lanske B, Soegiarto DW, Toftgard R, signaling mechanisms. Journal of Molecular Biology 428 3831–3849. Tsai MJ, Tsai S, Lydon JP, et al. 2006 Indian hedgehog is a major (https://doi.org/10.1016/j.jmb.2016.06.020) mediator of progesterone signaling in the mouse uterus. Nature Grimm D, Bauer J, Wise P, Kruger M, Simonsen U, Wehland M, Genetics 38 1204–1209. (https://doi.org/10.1038/ng1874) Infanger M & Corydon TJ 2019 The role of SOX family members in Lee KY, Jeong JW, Wang J, Ma L, Martin JF, Tsai SY, Lydon JP & solid tumours and metastasis. Seminars in Cancer Biology In press. DeMayo FJ 2007 Bmp2 is critical for the murine uterine decidual (https://doi.org/10.1016/j.semcancer.2019.03.004) response. Molecular and Cellular Biology 27 5468–5478. (https://doi. Guimaraes-Young A, Neff T, Dupuy AJ & Goodheart MJ 2016 org/10.1128/MCB.00342-07) Conditional deletion of Sox17 reveals complex effects on uterine Lee DK, Kurihara I, Jeong JW, Lydon JP, DeMayo FJ, Tsai MJ & Tsai SY adenogenesis and function. Developmental Biology 414 219–227. 2010 Suppression of ERalpha activity by COUP-TFII is essential for (https://doi.org/10.1016/j.ydbio.2016.04.010) successful implantation and decidualization. Molecular Endocrinology Hagan CR & Lange CA 2014 Molecular determinants of context- 24 930–940. (https://doi.org/10.1210/me.2009-0531) dependent progesterone receptor action in breast cancer. BMC Li Q, Kannan A, Wang W, Demayo FJ, Taylor RN, Bagchi MK & Bagchi IC Medicine 12 32. (https://doi.org/10.1186/1741-7015-12-32) 2007 Bone morphogenetic protein 2 functions via a conserved Hantak AM, Bagchi IC & Bagchi MK 2014 Role of uterine stromal- signaling pathway involving Wnt4 to regulate uterine decidualization epithelial crosstalk in embryo implantation. International Journal of in the mouse and the human. Journal of Biological Chemistry 282 Developmental Biology 58 139–146. (https://doi.org/10.1387/ 31725–31732. (https://doi.org/10.1074/jbc.M704723200) ijdb.130348mb) Li Q, Kannan A, DeMayo FJ, Lydon JP, Cooke PS, Yamagishi H, Hirate Y, Suzuki H, Kawasumi M, Takase HM, Igarashi H, Naquet P, Srivastava D, Bagchi MK & Bagchi IC 2011 The antiproliferative Kanai Y & Kanai-Azuma M 2016 Mouse Sox17 haploinsufficiency action of progesterone in uterine epithelium is mediated by Hand2. leads to female subfertility due to impaired implantation. Scientific Science 331 912–916. (https://doi.org/10.1126/science.1197454) Reports 6 24171. (https://doi.org/10.1038/srep24171) Li Q, Kannan A, Das A, Demayo FJ, Hornsby PJ, Young SL, Taylor RN, Jacob F & Monod J 1961 Genetic regulatory mechanisms in the Bagchi MK & Bagchi IC 2013 WNT4 acts downstream of BMP2 and synthesis of proteins. Journal of Molecular Biology 3 318–356. (https:// functions via beta-catenin signaling pathway to regulate human doi.org/10.1016/s0022-2836(61)80072-7) endometrial stromal cell differentiation. Endocrinology 154 446–457. Jeltsch JM, Krozowski Z, Quirin-Stricker C, Gronemeyer H, Simpson RJ, (https://doi.org/10.1210/en.2012-1585) Garnier JM, Krust A, Jacob F & Chambon P 1986 Cloning of the Lim HJ & Wang H 2010 Uterine disorders and pregnancy complications: chicken progesterone receptor. PNAS 83 5424–5428. (https://doi. insights from mouse models. Journal of Clinical Investigation 120 org/10.1073/pnas.83.15.5424) 1004–1015. (https://doi.org/10.1172/JCI41210)

Lin SC, Li YH, Wu MH, Chang YF, Lee DK, Tsai SY, Tsai MJ & Tsai SJ Rai R & Regan L 2006 Recurrent miscarriage. Lancet 368 601–611. 2014 Suppression of COUP-TFII by proinflammatory cytokines (https://doi.org/10.1016/S0140-6736(06)69204-0) contributes to the pathogenesis of endometriosis. Journal of Clinical Rubel CA, Franco HL, Jeong JW, Lydon JP & DeMayo FJ 2012a GATA2 is Endocrinology and Metabolism 99 E427–E437. (https://doi. expressed at critical times in the mouse uterus during pregnancy. org/10.1210/jc.2013-3717) Gene Expression Patterns 12 196–203. (https://doi.org/10.1016/j. Link W 2019 Introduction to FOXO biology. Methods in Molecular Biology gep.2012.03.004) 1890 1–9. (https://doi.org/10.1007/978-1-4939-8900-3_1) Rubel CA, Lanz RB, Kommagani R, Franco HL, Lydon JP & DeMayo FJ Lydon JP, DeMayo FJ, Funk CR, Mani SK, Hughes AR, Montgomery Jr 2012b Research resource: genome-wide profiling of progesterone CA, Shyamala G, Conneely OM & O’Malley BW 1995 Mice lacking receptor binding in the mouse uterus. Molecular Endocrinology 26 progesterone receptor exhibit pleiotropic reproductive abnormalities. 1428–1442. (https://doi.org/10.1210/me.2011-1355) Genes and Development 9 2266–2278. (https://doi.org/10.1101/ Rubel CA, Wu SP, Lin L, Wang T, Lanz RB, Li X, Kommagani R, gad.9.18.2266) Franco HL, Camper SA, Tong Q, et al. 2016 A Gata2-dependent Macklon NS, Geraedts JP & Fauser BC 2002 Conception to ongoing transcription network regulates uterine progesterone responsiveness pregnancy: the ‘black box’ of early pregnancy loss. Human and endometrial function. Cell Reports 17 1414–1425. (https://doi. Reproduction Update 8 333–343. (https://doi.org/10.1093/ org/10.1016/j.celrep.2016.09.093) humupd/8.4.333) Schlafke S & Enders AC 1975 Cellular basis of interaction between Mangelsdorf DJ, Thummel C, Beato M, Herrlich P, Schutz G, trophoblast and uterus at implantation. Biology of Reproduction 12 Umesono K, Blumberg B, Kastner P, Mark M, Chambon P, et al. 1995 41–65. (https://doi.org/10.1095/biolreprod12.1.41) The nuclear receptor superfamily: the second decade. Cell 83 Senna Tan D, Holzner M, Weng M, Srivastava Y & Jauch R 2019 SOX17 835–839. (https://doi.org/10.1016/0092-8674(95)90199-x) in cellular reprogramming and cancer. Seminars in Cancer Biology In Matsumoto H, Zhao X, Das SK, Hogan BL & Dey SK 2002 Indian press. (https://d oi.or g/10. 1016/ j.sem cance r.201 9.08. 008) hedgehog as a progesterone-responsive factor mediating epithelial- Soyal SM, Mukherjee A, Lee KY, Li J, Li H, DeMayo FJ & Lydon JP 2005 mesenchymal interactions in the mouse uterus. Developmental Biology Cre-mediated recombination in cell lineages that express the 245 280–290. (https://doi.org/10.1006/dbio.2002.0645) progesterone receptor. Genesis 41 58–66. (https://doi.org/10.1002/ Mulac-Jericevic B, Mullinax RA, DeMayo FJ, Lydon JP & Conneely OM gene.20098) 2000 Subgroup of reproductive functions of progesterone mediated Stewart CL, Kaspar P, Brunet LJ, Bhatt H, Gadi I, Kontgen F & by progesterone receptor-B isoform. Science 289 1751–1754. (https:// Abbondanzo SJ 1992 Blastocyst implantation depends on maternal doi.org/10.1126/science.289.5485.1751) expression of leukaemia inhibitory factor. Nature 359 76–79. Nallasamy S, Li Q, Bagchi MK & Bagchi IC 2012 Msx homeobox genes (https://doi.org/10.1038/359076a0) critically regulate embryo implantation by controlling paracrine Surveyor GA, Gendler SJ, Pemberton L, Das SK, Chakraborty I, Julian J, signaling between uterine stroma and epithelium. PLoS Genetics 8 Pimental RA, Wegner CC, Dey SK & Carson DD 1995 Expression and e1002500. (https://doi.org/10.1371/journal.pgen.1002500) steroid hormonal control of Muc-1 in the mouse uterus. Nallasamy S, Kaya Okur HS, Bhurke A, Davila J, Li Q, Young SL, Endocrinology 136 3639–3647. (https://doi.org/10.1210/ Taylor RN, Bagchi MK & Bagchi IC 2019 Msx homeobox genes act endo.136.8.7628404) downstream of BMP2 to regulate endometrial decidualization in Takamoto N, Zhao B, Tsai SY & DeMayo FJ 2002 Identification of Indian mice and in humans. Endocrinology 160 1631–1644. (https://doi. hedgehog as a progesterone-responsive gene in the murine uterus. org/10.1210/en.2019-00131) Molecular Endocrinology 16 2338–2348. (https://doi.org/10.1210/ Namiki T, Ito J & Kashiwazaki N 2018 Molecular mechanisms of me.2001-0154) embryonic implantation in mammals: lessons from the gene Toft D & Gorski J 1966 A receptor molecule for estrogens: isolation from manipulation of mice. Reproductive Medicine and Biology 17 331–342. the rat uterus and preliminary characterization. PNAS 55 1574–1581. (https://doi.org/10.1002/rmb2.12103) (https://doi.org/10.1073/pnas.55.6.1574) Ng JM & Curran T 2011 The Hedgehog’s tale: developing strategies for Tranguch S, Cheung-Flynn J, Daikoku T, Prapapanich V, Cox MB, Xie H, targeting cancer. Nature Reviews: Cancer 11 493–501. (https://doi. Wang H, Das SK, Smith DF & Dey SK 2005 Cochaperone org/10.1038/nrc3079) immunophilin FKBP52 is critical to uterine receptivity for embryo Okada H, Nakajima T, Sanezumi M, Ikuta A, Yasuda K & Kanzaki H 2000 implantation. PNAS 102 14326–14331. (https://doi.org/10.1073/ Progesterone enhances interleukin-15 production in human pnas.0505775102) endometrial stromal cells in vitro. Journal of Clinical Endocrinology Tremblay M, Sanchez-Ferras O & Bouchard M 2018 GATA transcription and Metabolism 85 4765–4770. (https://doi.org/10.1210/ factors in development and disease. Development 145 dev164384. jcem.85.12.7023) (https://doi.org/10.1242/dev.164384) O’Malley BW & Schrader WT 1972 Progesterone receptor components: Tsai MJ & O’Malley BW 1994 Molecular mechanisms of action of identification of subunits binding to the target-cell genome. Journal steroid/thyroid receptor superfamily members. Annual Review of of Steroid Biochemistry 3 617–629. (https://doi.org/10.1016/0022- Biochemistry 63 451–486. (https://doi.org/10.1146/annurev. 4731(72)90107-0) bi.63.070194.002315) O’Malley BW, McGuire WL, Kohler PO & Korenman SG 1969 Studies on Turco MY, Gardner L, Hughes J, Cindrova-Davies T, Gomez MJ, Farrell L, the mechanism of steroid hormone regulation of synthesis of Hollinshead M, Marsh SGE, Brosens JJ, Critchley HO, et al. 2017 specific proteins. Recent Progress in Hormone Research 25 105–160. Long-term, hormone-responsive organoid cultures of human (https://doi.org/10.1016/b978-0-12-571125-8.50006-5) endometrium in a chemically defined medium. Nature Cell Biology O’Malley BW, Sherman MR & Toft DO 1970 Progesterone ‘receptors’ in 19 568–577. (https://doi.org/10.1038/ncb3516) the cytoplasm and nucleus of chick oviduct target tissue. PNAS 67 Vasquez YM, Mazur EC, Li X, Kommagani R, Jiang L, Chen R, Lanz RB, 501–508. (https://doi.org/10.1073/pnas.67.2.501) Kovanci E, Gibbons WE & DeMayo FJ 2015 FOXO1 is required for Pawar S, Hantak AM, Bagchi IC & Bagchi MK 2014 Minireview: steroid- binding of PR on IRF4, novel transcriptional regulator of regulated paracrine mechanisms controlling implantation. Molecular endometrial stromal decidualization. Molecular Endocrinology 29 Endocrinology 28 1408–1422. (https://doi.org/10.1210/me.2014-1074) 421–433. (https://doi.org/10.1210/me.2014-1292) Psychoyos A 1973 Hormonal control of ovoimplantation. Vitamins and Vasquez YM, Wang X, Wetendorf M, Franco HL, Mo Q, Wang T, Hormones 31 201–256. (https://doi.org/10.1016/s0083- Lanz RB, Young SL, Lessey BA, Spencer TE, et al. 2018 FOXO1 6729(08)60999-1) regulates uterine epithelial integrity and progesterone receptor

expression critical for embryo implantation. PLoS Genetics 14 Wilkens J, Male V, Ghazal P, Forster T, Gibson DA, Williams AR, Brito- e1007787. (https://doi.org/10.1371/journal.pgen.1007787) Mutunayagam SL, Craigon M, Lourenco P, Cameron IT, et al. 2013 Wang X, Li X, Wang T, Wu SP, Jeong JW, Kim TH, Young SL, Lessey BA, Uterine NK cells regulate endometrial bleeding in women and are Lanz RB, Lydon JP, et al. 2018 SOX17 regulates uterine epithelial- suppressed by the progesterone receptor modulator asoprisnil. stromal cross-talk acting via a distal enhancer upstream of Ihh. Journal of Immunology 191 2226–2235. (https://doi.org/10.4049/ Nature Communications 9 4421. (https://doi.org/10.1038/s41467-018- jimmunol.1300958) 06652-w) Wu D, Sunkel B, Chen Z, Liu X, Ye Z, Li Q, Grenade C, Ke J, Zhang C, Wetendorf M, Wu SP, Wang X, Creighton CJ, Wang T, Lanz RB, Blok L, Chen H, et al. 2014 Three-tiered role of the pioneer factor GATA2 in Tsai SY, Tsai MJ, Lydon JP, et al. 2017 Decreased epithelial promoting androgen-dependent gene expression in prostate cancer. progesterone receptor A at the window of receptivity is required for Nucleic Acids Research 42 3607–3622. (https://doi.org/10.1093/nar/ preparation of the endometrium for embryo attachment. Biology of gkt1382) Reproduction 96 313–326. (https://doi.org/10.1095/ Wu SP, Yu CT, Tsai SY & Tsai MJ 2016 Choose your destiny: make a cell biolreprod.116.144410) fate decision with COUP-TFII. Journal of Steroid Biochemistry and Whitaker LH, Murray AA, Matthews R, Shaw G, Williams AR, Molecular Biology 157 7–12. (https://doi.org/10.1016/j. Saunders PT & Critchley HO 2017 Selective progesterone receptor jsbmb.2015.11.011) modulator (SPRM) ulipristal acetate (UPA) and its effects on the Wu SP, Li R & DeMayo FJ 2018 Progesterone receptor regulation of human endometrium. Human Reproduction 32 531–543. (https://doi. uterine adaptation for pregnancy. Trends in Endocrinology and org/10.1093/humrep/dew359) Metabolism 29 481–491. (https://doi.org/10.1016/j.tem.2018.04.001) Wilcox AJ, Baird DD & Weinberg CR 1999 Time of implantation of the Zinaman MJ, Clegg ED, Brown CC, O’Connor J & Selevan SG 1996 conceptus and loss of pregnancy. New England Journal of Medicine Estimates of human fertility and pregnancy loss. Fertility and Sterility 340 1796–1799. (https://doi.org/10.1056/NEJM199906103402304) 65 503–509. (https://doi.org/10.1016/S0015-0282(16)58144-8)

Received in final form 25 November 2019 Accepted 6 December 2019 Accepted Manuscript published online 6 December 2019