Functional enhancers are often composed of binding

Functional enhancers are often composed of binding motifs of multiple key transcription factors to confer spatial and temporal regulation of genes in a certain context. In the uterus, the difference between the number of genes that have associated PGR occupancy and that of progesterone responsive genes implicates the need of additional transcription factors to drive functional enhancers 29, 34. Taking uterine expression of the Ihh gene as an example, an outstanding question is how PGR specifically regulates Ihh responsiveness to progesterone in the epithelium given that Ihh is expressed in many tissues at various time points. Emerging evidence suggests that a putative enhancer located 19kb upstream of the Ihh transcription start site can promote gene expression in response to progesterone stimulation 29, 34. Importantly, this enhancer is the only cis-acting element within a 200-kb vicinity of the Ihh gene body that exhibits co-occupancy of PGR, GATA2, FOXA2, and SOX17 transcription factors in uterine tissues 29, 34, 79. These observations collectively implicate the potential of this enhancer as a uterine epithelium-specific enhancer with the capacity to regulate expression in the uterus. In vivo functionality assessment on this and other enhancers by CRISPR/Cas9-mediated genome editing in mice could address this question. It would also be interesting to investigate whether the core transcription regulators that occupy the uterine-specific enhancers have the capacity to determine cell fate and be of use in the generation of in vitro uterine epithelial cell models, which are currently still lacking in the field (see Outstanding Questions). Additionally, other members of the protein complex that occupy at the enhancer of interest could be further identified by CRISPR-guided proteomic analysis at loci of interest [80]. Moreover, interaction between the enhancer of interest and other noncoding genomic elements can be explored by chromosome conformation capture or chromatin immunoprecipitation-loop assays [81]. Lastly, emerging evidence demonstrates the impact of hormone regulation on chromatin dynamics during stromal cell decidualization [82], which adds an epigenetic layer of regulation to these functional enhancers. Integration of results from these new assays would provide insights on the interaction of various regulatory mechanisms for uterine-specific gene expression. Investigating model gene transcription and reintroducing PGRs in cultured human myometrial CAY10683 sale has greatly enhanced our understanding of the functional differences and similarities of PGR isoforms. However, whether these conceptual advancements are applicable at the genome-wide scale, as well as the physiological level, remains to be addressed (see Outstanding Questions). Examining mice with myometrium-specific alteration of isoform ratios by real-time monitoring of contractility would provide in vivo evidence to support current concepts [83]. Furthermore, analyses on transcriptome and cistrome of PGR isoforms in ratio-altered mouse myometrium would help to reveal functional characteristics of PGR isoforms. Notably, both PGR-A and PGR-B isoforms are expressed in the endometrium and altered PGR-A levels have been noted in diseased conditions, such as endometriosis [84]. Functionality studies of PGR isoforms using HESCs also support that PGR-A and PGR-B not only regulate common genes but also have their own distinct downstream targets [85]. Since mice and humans share preserved molecular pathways in early pregnancy, compartmental-specific alteration of isoform expression in mouse models would shed light on the physiological roles of PGR isoforms in the endometrium.
Acknowledgements We thank Diane Cooper at the NIH Library Services for manuscript editing. This work is supported by the Intramural Research Program of the National Institute of Environmental Health Sciences: ProjectZ1AES103311 (F.J.D.).
Introduction The nomenclature of mGlu receptors is numeric beginning with mGlu1, which was first isolated by expression cloning (Houamed et al., 1991; Masu et al., 1991). The cloning of subsequent mGlu receptors resulted in their sequential numbering. The mGlu receptors are divided into three sub-families based on sequence homology, second messenger coupling, and pharmacological properties. Group I (mGlu1 & mGlu5) receptors are coupled to Gαq-like proteins and stimulate phospholipase C (PLC), leading to an elevation of intracellular Ca2+ and activation of protein kinase C (PKC). Group II (mGlu2 & mGlu3) and Group III (mGlu4, mGlu6, mGlu7 & mGlu8) receptors are coupled to Gi/Go proteins and inhibit adenylate cyclase (AC), which leads to a reduction of cAMP and inactivation of protein kinase A (PKA). The signaling cascades of mGlu receptors are far more complex than these canonical pathways (Niswender and Conn, 2010; Willard and Koochekpour, 2013). Group I mGlu receptors, for example, can modulate unconventional signaling pathways involving Gi/o and Gs (Aramori and Nakanishi, 1992; Francesconi and Duvoisin, 1998; Hermans et al., 2000; Thomsen, 1996). Furthermore, mGlu7 can stimulate PLC through the activation of a pertussis toxin (PTX)-sensitive or -resistant G proteins (Martin et al., 2010; Perroy et al., 2000), and mGlu2 or mGlu4 can activate Gα15 and PLC (Gomeza et al., 1996).