[Frontiers in Bioscience E3, 1092-1099, June 1, 2011]
Non-coding RNAs and embryo implantation
Ji-Long Liu1, Zeng-Ming Yang1,2
1
TABLE OF CONTENTS
1. ABSTRACT
In mammals, thousands of non-protein-coding RNAs (ncRNAs), including microRNAs, endogenous small interfering RNAs, PIWI-interacting RNAs and mRNA-like long ncRNAs, have been identified. These RNAs modulate gene expression at transcriptional, post-transcriptional and epigenetic levels in many developmental and metabolic processes. Increasing evidence shows that ncRNAs are also linked to embryo implantation through regulating the expression of certain key genes and pathways. In this paper, we summarized the recent literatures on the ncRNAs involved in embryo implantation.
2. INTRODUCTION
Embryo implantation is an intricate interaction between the implanting embryo and receptive uterus. This process is primed by ovarian steroid hormones for paracrine, autocrine, and juxtacrine molecular events within the uterus to produce a favorable environment for embryo implantation (1, 2). Over past decades, efforts have extensively been made to study the protein-coding genes for deciphering the biological process of embryo implantation. There is growing awareness of the importance of recently discovered non-protein-coding RNAs (ncRNAs) in the regulation of gene expression during developmental and metabolic processes (3, 4). However, the regulatory role of ncRNAs during embryo implantation is largely unexplored.
In recent years, with advances in large-scale DNA sequencing projects, the number of ncRNA sequences in eukaryotic genomes is rapidly expanding, although the number of protein-coding genes remains relatively static (5). NcRNAs are a heterogeneous group of RNA molecules, varying in size and mechanism of action. A basic classification criterion is the size. Small ncRNAs are <200 nt long and are generated from long precursor transcripts while long ncRNAs (lncRNA) are typically >200 nt long and function without major prior processing.
Small ncRNAs comprise microRNAs (miRNAs), endogenous small interfering RNAs (siRNAs) and PIWI-interacting RNAs (piRNAs). Endogenous siRNAs and piRNAs are highly and restrictively enriched in germline cells (6-9). From two-cell stage, the zygotic genome begins to produce miRNAs. A transition of the major small RNA class from siRNA/piRNA to miRNA was observed during pre-implantation embryo development (10). MiRNAs are the predominant class of small ncRNAs in developing and adult tissues. Through interacting with the 3' untranslated regions (UTRs) of protein-coding genes, miRNAs can cause mRNA degradation or translational repression (11). Several miRNAs have been identified to be differentially expressed in the uterus during embryo implantation in the human (12, 13) and mouse (14, 15) using microarrays. Little is known about their functions in embryo implantation.
The long ncRNAs are generated from intergenic region of genome as well as opposite strand and intron of protein coding genes. They range from approximately 200 nt to over 100k nt in size. Unlike miRNAs, a unified mechanism of long ncRNA action is missing. Instead, ncRNAs have been found to control gene expression at transcriptional, post-transcriptional and epigenetic levels (16). Recent studies show that some long ncRNAs also have activities for miRNA sponge (17) or miRNA masking (18), which add complexity to the functionality of long ncRNAs. Noticeably, most long ncRNAs have functions in cis, which means they merely regulate the expression of nearby protein-coding genes. The HOX transcript antisense RNA (HOTAIR), which originates from the HOXC locus and silences transcription across 40 kb of the HOXD locus (19), is one of the few examples of long ncRNAs that act in trans. During pre-implantation embryo development, the primary roles of long ncRNAs appear to be as antisense regulators of genomic imprinting (20). Some long ncRNAs were expressed in adult uterus during embryo implantation (21, 22). The function and mechanism of action of these long ncRNAs remains to be elucidated.
As discussed above, miRNAs and long ncRNAs are two major classes of ncRNAs expressed in adult uterus. In this paper, we will review the current knowledge about uterine expression, regulation, function, and mechanism of action of miRNAs and long ncRNAs during embryo implantation.
3. Role of ncRNAs in the uterus during embryo implantation
3.1. miRNAs
Mature miRNAs are synthesized through a stepwise process which concludes with the cleavage of stem-loop precursor miRNAs by the RNase III enzyme, Dicer1. Therefore, depletion of Dicer1 impairs miRNA formation. In mice, general knockout of Dicer1 leads to lethality at embryonic day 11.5 (23). In order to study the role of miRNAs in female reproduction, global Dicer1 hypomorphic mice (24) and Amhr2-Cre driven conditional Dicer1 knockout mice (25-27) have been generated. These mice are infertile because of insufficient corpus luteum (24) or disorganized oviduct (25-27). Besides serious oviduct deficiency, Amhr2-Cre driven conditional Dicer1 knockout mice also display some uterine abnormalities. Their uteri are much shorter, thinner, and lighter than wild-type uteri. Additionally, a decreased smooth muscle layer and decreased presence of uterine glands were observed (27). However, it is unknown whether the uteri of these mice are capable of embryo implantation.
Uterine miRNAs are expressed in a steroid hormone-dependent manner (28). Several groups have shown that miRNA expression profiles change dynamically in the uterus during embryo implantation (12-15). Functional studies demonstrated that a portion of these differentially expressed miRNAs are involved in the process of epithelium/stroma differentiation, inflammation, and embryo invasion.
3.1.1. miR-21
Using microarrays, miRNA expression profiles of mouse uterus at implantation sites versus inter-implantation sites on day 5 of pregnancy were compared. A total of 15 differentially expressed miRNAs were indentified, of which miR-143, miR-21, miR-20a, miR-26a, let-7a, let-7b, let-7c, and let-7d were confirmed by both Northern blot and in situ hybridization (14).
In situ hybridization analysis showed that miR-21 expression is not detectable in the mouse uterus from days 1 to 4 of pregnancy. On day 5 of pregnancy, miR-21 is strongly expressed in the subluminal stroma underlying the implanting blastocyst at implantation sites but remained absent at inter-implantation sites. From days 6 to 8 of pregnancy, miR-21 expression is also highly expressed in the decidua (14).
The expression level of miR-21 was significantly reduced by estradiol treatment and slightly down-regulated by progesterone (14). Previous studies have shown that miR-21 expression is induced by the signal transducer and activator of transcription-3 (STAT3) pathway through STAT3 binding sites located within promoter region of miR-21 in myeloma and breast cancer cell lines (29, 30). In the mouse uterus, phosphorylated STAT3 protein is strongly detected in the luminal epithelium and stroma surrounding the implanting blastocyst at implantation sites on day 5 of pregnancy (31). Nuclear localization of Stat3 in day 4 luminal epithelium is not detected in any LIF-deficient mice, indicating that LIF is the principal mediator of STAT3 activation in vivo (32). During embryo implantation, STAT3 is activated by leukemia inhibitory factor (LIF). Taken together, it seems that miR-21 at implantation sites is not modulated by estrogen directly but through the activation of LIF/STAT3 pathway.
Reck is identified as a downstream target of miR-21 using bioinformatics tools. The expression pattern of Reck is opposite to that of miR-21 during embryo implantation. Luciferase assay confirmed that Reck is a direct target of miR-21. It is reported that Reck inhibits MMP-2 and MMP-9 secretion or activity, or both (33). However, Reck only suppresses MMP-9 activity in cultured uterine stromal cells. Thus, miR-21 may participate in embryo implantation through inhibiting Reck to regulate the activity of MMP-9 (14).
3.1.2. miR-101a and miR-199a*
MiRNA microarray was also performed on day 1 (pre-receptive) versus day 4 (receptive) uteri of pregnant mice (15). Statistical analysis shows that 32 miRNAs are significantly induced on day 4 and only 5 miRNAs are expressed at higher levels on day 1.
Among up-regulated miRNAs in the receptive stage, miR-101a and miR-199a* are regulators of cyclooxygenase-2 (Cox-2). Using in situ hybridization, the spatiotemporal expression of these miRNAs in early pregnancy and delayed implantation is determined. Both miR-101a and miR-199a* are detected in the luminal epithelium on day 1 of pregnancy and become dispersed throughout the uterus on day 4. On day 5, they are localized around the blastocysts at the antimesometrial pole. From days 6 to 8, the expression of these miRNAs disappears from the antimesometrial pole, but is primarily detected at the mesometrial pole of the implantation site. In the delayed implantation model, both miRNAs are not expressed in the uterus during progesterone primed delayed implantation, but is readily induced in stromal cells surrounding the activated blastocyst after termination of the delay by estrogen. This expression pattern is very similar to that of Cox-2 mRNA (34). More importantly, Cox-2 protein is inversely correlated with miR-101a expression at each time point during artificial decidualization process after oil infusion. As Cox-2 is known to be necessary for embryo implantation and decidualization (35, 36), miR-101a and miR-199a* are likely important regulators of Cox-2 during embryo implantation and decidualization (15).
3.1.3. miR-222
MiR-222 is identified in a microarray study of miRNA expression profiles between the non-induced and induced human endometrial stromal cells (ESCs). It is strongly expressed in ESCs but down-regulated during cAMP and progesterone induced decidualization (12). Suppression of mir-222 by antisense oligonucleotides inhibits the growth of ESCs. Western blot confirms a significant increase of CDKN1C/p57kip2 protein level in the miR-222 antisense oligonucleotide-treated ESCs. A miR-222 binding site located in the 3'-UTR region of the CDKN1C/p57kip2 mRNA is validated by luciferase assay. Collectively, miR-222 is likely to be a modulator of decidualization by helping ESCs to exit the cell cycle and enter differentiation (12).
3.1.4. miR-210
In another microarray study, miRNA expression profiles of uterine epithelial cells purified from endometrium in the late proliferative and mid-secretory phase were compared (13). Among the 12 miRNAs up-regulated in the mid-secretory phase, many of them are predicted to target cell cycle related genes. As a representative example, miR-210 is increased in the mid-secretory phase, while E2F3, a known target of miR-210 (37, 38), is decreased. Thus, miRNAs such as miR-210 may participate in down-regulating the expression of cell cycle genes in the mid-secretory phase of endometrial epithelium, thereby suppressing cell proliferation (13).
3.2. Long ncRNAs
Several long ncRNAs are actively transcribed in the uterus during embryo implantation from the opposite strand of protein-coding genes, such as HOXA11 (21), Scx (22), bFGF (39), and EMX2 (40). These naturally occurring antisense RNAs are supposed to mediate transcriptional activation or transcriptional silencing of host genes. It is noteworthy that the steroid receptor RNA activator (SRA) gene, which is characterized as an intergenic ncRNA and steroid receptor co-activator, is also expressed in the uterus (41). However, gain-of-function and/or loss-of-function studies will be needed to definitely find out the regulatory effect of long ncRNAs in embryo implantation.
3.2.1. HOXA11-AS
The homeobox (Hox) genes are a set of transcription factors that determine the identity of cells and tissues during embryonic development. In mammals, Hox genes are organized into four clusters, termed A, B, C and D. Hox clusters A and D are crucial in the developing reproductive tract (42-44). Although Hox genes are typically considered to be expressed only during embryonic development, the persistent expression of HOX genes has also been noted in adult tissues (45). Specifically, HOXA10 and HOXA11 are expressed in adult human and mouse uteri.
Hoxa11 is required for uterine stromal cell and glandular cell differentiation during embryo implantation. Ablation of Hoxa11 gene in mice results in female infertility (46). Endogenous HOXA11 antisense RNA (HOXA11-AS) is present in mouse and human endometrium (21). In human uterine endometrium, the temporal expression of HOXA11-AS is complementary to that of HOXA11. HOXA11-AS mRNA level varies during the menstrual cycle, with peak expression in the mid-proliferative phase (21). HOXA11 mRNA and protein are known to rise in the mid-secretory phase, corresponding to the time of endometrial receptivity to blastocyst implantation (47). In cultured stromal cells, progesterone is able to repress HOXA11-AS mRNA, which results in up-regulation of HOXA11 mRNA. However, synthetic Hoxa11 antisense oligonucleotides transfected into the uterus fail to block Hoxa11 protein expression or Hoxa11 function in mice. Therefore, HOXA11-AS probably represses HOXA11 expression by competing for the transcription of the HOXA11 gene, rather than by directly binding HOXA11 sense mRNA (21).
3.2.2. Scx-AS
Scleraxis (Scx) is a basic helix-loop-helix type transcription factor that regulates the growth and differentiation of numerous cell types, such as osteoblastic cells (48), chondrogenic cells (49), tendon (50) and sertoli cells (51). Interestingly, Scx is located in the intron 3 region of the Bop1 gene, but on the opposite strand. Bop1 is a ribosome biogenesis protein and is involved in cell proliferation (52, 53). Thus, the Scx-Bop1 genomic locus has opposite function on both strands.
Both Scx and Bop1 are expressed in the uterine endometrial epithelial cells (22). In ovariectomized adult mice treated with estrogen, the uterine Scx mRNA is decreased at 6 h after estrogen treatment and gradually recovered by 24 h. On the contrary, Bop1 mRNA is increased by 6 folds and gradually reduced until 24 h. These two genes are reciprocally regulated by estrogen. In situ hybridization indicates that the antisense RNA of Scx (Scx-AS) is also expressed in the uterine endometrial epithelial cells. Scx-AS is increased after estrogen treatment, coinciding with a decrease in Scx mRNA. In vitro cell assay confirms that Scx-AS mRNA reduces the Scx gene expression in a trans-acting manner (22). The discovery of Scx-AS suggests that long ncRNAs may play an important role in steroid-mediated gene expression during embryo implantation.
3.2.3. EMX2-OS
Emx2 is a homeobox gene located outside of the Hox cluster. It is a transcriptional target of Hoxa10 regulation in the reproductive tract (54). EMX2 is expressed in the endometrium throughout the estrous cycle and is an important regulator of endometrial proliferation and embryo implantation (55).
Antisense transcripts at the EMX2 locus (EMX2-OS) are highly expressed in human uterus, kidney, and brain (40). In human uterine endometrium, EMX2 and EMX2-OS are identically expressed primarily in the epithelium, with a lower level of expression in the stroma. In mouse uterus, homologous EMX2 and EMX2-OS are co-expressed at high levels in the epithelium and at lower levels in the surrounding stroma during the estrous cycle (40). Recent study shows that EMX2-OS can either up-regulate or down-regulate EMX2 expression in a cell type dependent way (56). Since the expression of EMX2 and EMX2-OS are positively correlated in uterus, EMX2-OS is likely to be an activator of EMX2 during embryo implantation.
3.2.4. FGF2-AS
Fibroblast growth factor-2 (FGF2), also known as basic fibroblast growth factor (bFGF), is a heparin-binding cationic protein with mitogenic and angiogenic properties (57). The endogenous antisense mRNA of FGF2 (FGF2-AS) is present in many human tissues (58, 59). FGF2-AS can repress FGF2 expression and vice versa (60).
In endometrial stromal cells derived from women with endometriosis, FGF2-AS mRNA levels are significantly lower in late proliferative phase than those in early proliferative phase and are lower, although not significantly, compared with those in the luteal phase. This expression pattern is opposite to that of FGF2 mRNA. However, FGF2-AS mRNA level doesn't vary significantly in endometrial stromal cells from normal women during the menstrual cycle (39). It seems that the up-regulation of FGF2-AS is specific for uterine malignancies. The function of FGF2-AS in normal endometrium is unclear.
3.2.5. SRA
The steroid receptor RNA activator (SRA) is first characterized as a long ncRNA interacting with the N-terminal domain of the human progesterone receptor in a yeast two-hybrid screen (61). Further studies have demonstrated that SRA can co-activate steroid receptors (61), nuclear receptors (62) as well as other transcription factors such as MyoD (63). Although protein products derived from the short open reading frames in the SRA sequence have been detected in multiple tissues and cell lines (64), they are not required for SRA activity (65).
SRA exhibits various expression levels in pituitary gland, adrenal gland, liver, breast, ovaries, uterus and prostate. Besides, SRA is significantly up-regulated in uterine tumors compared to normal uterus (41). The uterine receptivity is established and maintained by ovarian steroid hormones, progesterone and estrogen. Most of the steroid hormone responses in the uterus are attributed to activation of classical nuclear steroid receptors (NSRs), although non-genomic mechanisms may exist (66). Thus, as a steroid receptor co-activator, SRA is potential regulator of hormone action in the uterus during embryo implantation.
4. CONCLUSIONS AND PERSPECTIVES
During the last decade, newly discovered short and long ncRNAs have dramatically changed our understanding on the transcriptome. The regulatory role of ncRNAs is attracting more and more attention. Recent evidences suggest that ncRNAs are linked to the embryo implantation process through regulating the expression of certain critical genes and pathways.
Due to severe defects in ovary or oviduct development before uterine receptivity can be established, the impact of impaired Dicer1 or miRNAs on embryo implantation cannot be directly evaluated in Dicer1 hypomorph mouse (24) or Amhr2-Cre driven conditional Dicer1 knockout mouse (25-27). However, we can still find some clues. Dicer1 hypomorph mice transplanted with wild type ovaries are fertile but deliver an average of 3.4 pups per litter, which is much less than that of wild type mice (24). Amhr2-Cre driven conditional Dicer1 knockout mice have smaller uteri with decreased smooth muscle layer and glands (27). Taken together, we suppose that impaired miRNA expression in uterus will affect embryo implantation rate and decidual development process. Good news is that chemical inhibitors of Dicer1 have been recently identified (67). These inhibitors are suitable tools for characterizing the role of miRNAs in embryo implantation. To date, much of the work has focused on miRNA profiling in the uterus. A number of miRNAs have been identified as differentially expressed ones during embryo implantation (12-15). However, only a small portion of these miRNAs have been explored further into their regulation and function during embryo implantation.
Although some long ncRNAs have been found to be dynamically expressed in the uterus during embryo implantation, these studies are rather preliminary (21, 22, 39-41). None of these ncRNAs have been proved to be indispensable in embryo implantation. There is also evidence for the existence of some other long ncRNAs in the uterus along with some other organs (68-70). The detailed expression pattern of these long ncRNAs during embryo implantation is yet to be determined. Interestingly, some well-known genes in embryo implantation, including COX-2, p53, PGR, HIFA, and MSX1, also have nearby long ncRNAs in normal or malignant tissues other than uterus (71-75). We do not know if these long ncRNAs are present and functional in the normal uterus during embryo implantation. Noticeably, although microarrays and shot-gun RNA sequencing technologies are ready for long ncRNA profiling assay (76, 77), the global expression levels of long ncRNAs in the uterus during embryo implantation is still unavailable.
Furthermore, the knowledge of ncRNAs is growing fast. Additional new classes and subclasses of ncRNAs are recently discovered, such as enhancer RNAs (78), TSS-associated RNAs (79), and miRNA embedded functional long ncRNAs (80). Presently, we have no idea about the expression or function of these novel ncRNA classes in the uterus during embryo implantation.
Undoubtedly, ncRNAs have provided new insights on the complex molecular mechanisms of embryo implantation. It is expected that understanding the role of ncRNAs in uterus during embryo implantation will allow for identification of targets in treatment of infertility and help to develop novel contraceptives.
5. ACKNOWLEDGMENT
This work was financially supported by National Basic Research Program of China (2011CB944402) and National Natural Science Foundation of China (30930013).
6. REFERENCES
1. Dey, S. K., H. Lim, S. K. Das, J. Reese, B. C. Paria, T. Daikoku & H. Wang: Molecular cues to implantation. Endocr Rev, 25, 341-73 (2004)
doi:10.1210/er.2003-0020
http://dx.doi.org/doi:10.1210/er.2003-0020
2. Wang, H. & S. K. Dey: Roadmap to embryo implantation: clues from mouse models. Nat Rev Genet, 7, 185-99 (2006)
doi:10.1038/nrg1808
http://dx.doi.org/doi:10.1038/nrg1808
3. Mattick, J. S. & I. V. Makunin: Non-coding RNA. Hum Mol Genet, 15 Spec No 1, R17-29 (2006)
doi:10.1093/hmg/ddl046
http://dx.doi.org/doi:10.1093/hmg/ddl046
4. Brosnan, C. A. & O. Voinnet: The long and the short of noncoding RNAs. Curr Opin Cell Biol, 21, 416-25 (2009)
doi:10.1016/j.ceb.2009.04.001
http://dx.doi.org/doi:10.1016/j.ceb.2009.04.001
5. Alexander, R. P., G. Fang, J. Rozowsky, M. Snyder & M. B. Gerstein: Annotating non-coding regions of the genome. Nat Rev Genet, 11, 559-71 (2010)
doi:10.1038/nrg2814
http://dx.doi.org/doi:10.1038/nrg2814
6. O'Donnell, K. A. & J. D. Boeke: Mighty Piwis defend the germline against genome intruders. Cell, 129, 37-44 (2007)
7. Klattenhoff, C. & W. Theurkauf: Biogenesis and germline functions of piRNAs. Development, 135, 3-9 (2008)
doi:10.1242/dev.006486
http://dx.doi.org/doi:10.1242/dev.006486
8. Babiarz, J. E., J. G. Ruby, Y. Wang, D. P. Bartel & R. Blelloch: Mouse ES cells express endogenous shRNAs, siRNAs, and other Microprocessor-independent, Dicer-dependent small RNAs. Genes Dev, 22, 2773-85 (2008)
doi:10.1101/gad.1705308
http://dx.doi.org/doi:10.1101/gad.1705308
9. Okamura, K., S. Balla, R. Martin, N. Liu & E. C. Lai: Two distinct mechanisms generate endogenous siRNAs from bidirectional transcription in Drosophila melanogaster. Nat Struct Mol Biol, 15, 581-90 (2008)
doi:10.1038/nsmb.1438
http://dx.doi.org/doi:10.1038/nsmb.1438
10. Ohnishi, Y., Y. Totoki, A. Toyoda, T. Watanabe, Y. Yamamoto, K. Tokunaga, Y. Sakaki, H. Sasaki & H. Hohjoh: Small RNA class transition from siRNA/piRNA to miRNA during pre-implantation mouse development. Nucleic Acids Res, 38, 5141-51 (2010)
doi:10.1093/nar/gkq229
http://dx.doi.org/doi:10.1093/nar/gkq229
11. Bartel, D. P.: MicroRNAs: genomics, biogenesis, mechanism, and function. Cell, 116, 281-97 (2004)
12. Qian, K., L. Hu, H. Chen, H. Li, N. Liu, Y. Li, J. Ai, G. Zhu, Z. Tang & H. Zhang: Hsa-miR-222 is involved in differentiation of endometrial stromal cells in vitro. Endocrinology, 150, 4734-43 (2009)
doi:10.1210/en.2008-1629
http://dx.doi.org/doi:10.1210/en.2008-1629
13. Kuokkanen, S., B. Chen, L. Ojalvo, L. Benard, N. Santoro & J. W. Pollard: Genomic profiling of microRNAs and messenger RNAs reveals hormonal regulation in microRNA expression in human endometrium. Biol Reprod, 82, 791-801 (2010)
doi:10.1095/biolreprod.109.081059
http://dx.doi.org/doi:10.1095/biolreprod.109.081059
14. Hu, S. J., G. Ren, J. L. Liu, Z. A. Zhao, Y. S. Yu, R. W. Su, X. H. Ma, H. Ni, W. Lei & Z. M. Yang: MicroRNA expression and regulation in mouse uterus during embryo implantation. J Biol Chem, 283, 23473-84 (2008)
doi:10.1074/jbc.M800406200
http://dx.doi.org/doi:10.1074/jbc.M800406200
15. Chakrabarty, A., S. Tranguch, T. Daikoku, K. Jensen, H. Furneaux & S. K. Dey: MicroRNA regulation of cyclooxygenase-2 during embryo implantation. Proc Natl Acad Sci U S A, 104, 15144-9 (2007)
doi:10.1073/pnas.0705917104
http://dx.doi.org/doi:10.1073/pnas.0705917104
16. Mercer, T. R., M. E. Dinger & J. S. Mattick: Long non-coding RNAs: insights into functions. Nat Rev Genet, 10, 155-9 (2009)
doi:10.1038/nrg2521
http://dx.doi.org/doi:10.1038/nrg2521
17. Poliseno, L., L. Salmena, J. Zhang, B. Carver, W. J. Haveman & P. P. Pandolfi: A coding-independent function of gene and pseudogene mRNAs regulates tumour biology. Nature, 465, 1033-8 (2010)
doi:10.1038/nature09144
http://dx.doi.org/doi:10.1038/nature09144
18. Faghihi, M. A., M. Zhang, J. Huang, F. Modarresi, M. P. Van der Brug, M. A. Nalls, M. R. Cookson, G. St-Laurent, 3rd & C. Wahlestedt: Evidence for natural antisense transcript-mediated inhibition of microRNA function. Genome Biol, 11, R56 (2010)
doi:10.1186/gb-2010-11-5-r56
http://dx.doi.org/doi:10.1186/gb-2010-11-5-r56
19. Rinn, J. L., M. Kertesz, J. K. Wang, S. L. Squazzo, X. Xu, S. A. Brugmann, L. H. Goodnough, J. A. Helms, P. J. Farnham, E. Segal & H. Y. Chang: Functional demarcation of active and silent chromatin domains in human HOX loci by noncoding RNAs. Cell, 129, 1311-23 (2007)
20. Koerner, M. V., F. M. Pauler, R. Huang & D. P. Barlow: The function of non-coding RNAs in genomic imprinting. Development, 136, 1771-83 (2009)
doi:10.1242/dev.030403
http://dx.doi.org/doi:10.1242/dev.030403
21. Chau, Y. M., S. Pando & H. S. Taylor: HOXA11 silencing and endogenous HOXA11 antisense ribonucleic acid in the uterine endometrium. J Clin Endocrinol Metab, 87, 2674-80 (2002)
doi:10.1210/jc.87.6.2674
http://dx.doi.org/doi:10.1210/jc.87.6.2674
22. Arao, Y., K. Carpenter, S. Hewitt & K. S. Korach: Estrogen down-regulation of the Scx gene is mediated by the opposing strand-overlapping gene Bop1. J Biol Chem, 285, 4806-14 (2010)
doi:10.1074/jbc.M109.036681
http://dx.doi.org/doi:10.1074/jbc.M109.036681
23. Bernstein, E., S. Y. Kim, M. A. Carmell, E. P. Murchison, H. Alcorn, M. Z. Li, A. A. Mills, S. J. Elledge, K. V. Anderson & G. J. Hannon: Dicer is essential for mouse development. Nat Genet, 35, 215-7 (2003)
doi:10.1038/ng1253
http://dx.doi.org/doi:10.1038/ng1253
24. Otsuka, M., M. Zheng, M. Hayashi, J. D. Lee, O. Yoshino, S. Lin & J. Han: Impaired microRNA processing causes corpus luteum insufficiency and infertility in mice. J Clin Invest, 118, 1944-54 (2008)
doi:10.1172/JCI33680
http://dx.doi.org/doi:10.1172/JCI33680
25. Nagaraja, A. K., C. Andreu-Vieyra, H. L. Franco, L. Ma, R. Chen, D. Y. Han, H. Zhu, J. E. Agno, P. H. Gunaratne, F. J. DeMayo & M. M. Matzuk: Deletion of Dicer in somatic cells of the female reproductive tract causes sterility. Mol Endocrinol, 22, 2336-52 (2008)
doi:10.1210/me.2008-0142
http://dx.doi.org/doi:10.1210/me.2008-0142
26. Gonzalez, G. & R. R. Behringer: Dicer is required for female reproductive tract development and fertility in the mouse. Mol Reprod Dev, 76, 678-88 (2009)
doi:10.1002/mrd.21010
http://dx.doi.org/doi:10.1002/mrd.21010
27. Hong, X., L. J. Luense, L. K. McGinnis, W. B. Nothnick & L. K. Christenson: Dicer1 is essential for female fertility and normal development of the female reproductive system. Endocrinology, 149, 6207-12 (2008)
doi:10.1210/en.2008-0294
http://dx.doi.org/doi:10.1210/en.2008-0294
28. Nothnick, W. B. & C. Healy: Estrogen induces distinct patterns of microRNA expression within the mouse uterus. Reprod Sci, 17, 987-94 (2010)
doi:10.1177/1933719110377472
http://dx.doi.org/doi:10.1177/1933719110377472
29. Loffler, D., K. Brocke-Heidrich, G. Pfeifer, C. Stocsits, J. Hackermuller, A. K. Kretzschmar, R. Burger, M. Gramatzki, C. Blumert, K. Bauer, H. Cvijic, A. K. Ullmann, P. F. Stadler & F. Horn: Interleukin-6 dependent survival of multiple myeloma cells involves the Stat3-mediated induction of microRNA-21 through a highly conserved enhancer. Blood, 110, 1330-3 (2007)
doi:10.1182/blood-2007-03-081133
http://dx.doi.org/doi:10.1182/blood-2007-03-081133
30. Iliopoulos, D., S. A. Jaeger, H. A. Hirsch, M. L. Bulyk & K. Struhl: STAT3 activation of miR-21 and miR-181b-1 via PTEN and CYLD are part of the epigenetic switch linking inflammation to cancer. Mol Cell, 39, 493-506 (2010)
doi:10.1016/j.molcel.2010.07.023
http://dx.doi.org/doi:10.1016/j.molcel.2010.07.023
31. Teng, C. B., H. L. Diao, X. H. Ma, L. B. Xu & Z. M. Yang: Differential expression and activation of Stat3 during mouse embryo implantation and decidualization. Mol Reprod Dev, 69, 1-10 (2004)
doi:10.1002/mrd.20149
http://dx.doi.org/doi:10.1002/mrd.20149
32. Cheng, J. G., J. R. Chen, L. Hernandez, W. G. Alvord & C. L. Stewart: Dual control of LIF expression and LIF receptor function regulate Stat3 activation at the onset of uterine receptivity and embryo implantation. Proc Natl Acad Sci U S A, 98, 8680-5 (2001)
doi:10.1073/pnas.151180898
http://dx.doi.org/doi:10.1073/pnas.151180898
33. Clark, J. C., D. M. Thomas, P. F. Choong & C. R. Dass: RECK--a newly discovered inhibitor of metastasis with prognostic significance in multiple forms of cancer. Cancer Metastasis Rev, 26, 675-83 (2007)
doi:10.1007/s10555-007-9093-8
http://dx.doi.org/doi:10.1007/s10555-007-9093-8
34. Chakraborty, I., S. K. Das, J. Wang & S. K. Dey: Developmental expression of the cyclo-oxygenase-1 and cyclo-oxygenase-2 genes in the peri-implantation mouse uterus and their differential regulation by the blastocyst and ovarian steroids. J Mol Endocrinol, 16, 107-22 (1996)
doi:10.1677/jme.0.0160107
http://dx.doi.org/doi:10.1677/jme.0.0160107
35. Lim, H., B. C. Paria, S. K. Das, J. E. Dinchuk, R. Langenbach, J. M. Trzaskos & S. K. Dey: Multiple female reproductive failures in cyclooxygenase 2-deficient mice. Cell, 91, 197-208 (1997)
36. Ye, X., K. Hama, J. J. Contos, B. Anliker, A. Inoue, M. K. Skinner, H. Suzuki, T. Amano, G. Kennedy, H. Arai, J. Aoki & J. Chun: LPA3-mediated lysophosphatidic acid signalling in embryo implantation and spacing. Nature, 435, 104-8 (2005)
doi:10.1038/nature03505
http://dx.doi.org/doi:10.1038/nature03505
37. Giannakakis, A., R. Sandaltzopoulos, J. Greshock, S. Liang, J. Huang, K. Hasegawa, C. Li, A. O'Brien-Jenkins, D. Katsaros, B. L. Weber, C. Simon, G. Coukos & L. Zhang: miR-210 links hypoxia with cell cycle regulation and is deleted in human epithelial ovarian cancer. Cancer Biol Ther, 7, 255-64 (2008)
38. Biswas, S., S. Roy, J. Banerjee, S. R. Hussain, S. Khanna, G. Meenakshisundaram, P. Kuppusamy, A. Friedman & C. K. Sen: Hypoxia inducible microRNA 210 attenuates keratinocyte proliferation and impairs closure in a murine model of ischemic wounds. Proc Natl Acad Sci U S A, 107, 6976-81 (2010)
doi:10.1073/pnas.1001653107
http://dx.doi.org/doi:10.1073/pnas.1001653107
39. Mihalich, A., M. Reina, S. Mangioni, E. Ponti, L. Alberti, P. Vigano, M. Vignali & A. M. Di Blasio: Different basic fibroblast growth factor and fibroblast growth factor-antisense expression in eutopic endometrial stromal cells derived from women with and without endometriosis. J Clin Endocrinol Metab, 88, 2853-9 (2003)
doi:10.1210/jc.2002-021434
http://dx.doi.org/doi:10.1210/jc.2002-021434
40. Noonan, F. C., P. J. Goodfellow, L. J. Staloch, D. G. Mutch & T. C. Simon: Antisense transcripts at the EMX2 locus in human and mouse. Genomics, 81, 58-66 (2003)
doi:10.1016/S0888-7543(02)00023-X
http://dx.doi.org/doi:10.1016/S0888-7543(02)00023-X
41. Lanz, R. B., S. S. Chua, N. Barron, B. M. Soder, F. DeMayo & B. W. O'Malley: Steroid receptor RNA activator stimulates proliferation as well as apoptosis in vivo. Mol Cell Biol, 23, 7163-76 (2003)
doi:10.1128/MCB.23.20.7163-7176.2003
http://dx.doi.org/doi:10.1128/MCB.23.20.7163-7176.2003
42. Taylor, H. S., G. B. Vanden Heuvel & P. Igarashi: A conserved Hox axis in the mouse and human female reproductive system: late establishment and persistent adult expression of the Hoxa cluster genes. Biol Reprod, 57, 1338-45 (1997)
doi:10.1095/biolreprod57.6.1338
http://dx.doi.org/doi:10.1095/biolreprod57.6.1338
43. Warot, X., C. Fromental-Ramain, V. Fraulob, P. Chambon & P. Dolle: Gene dosage-dependent effects of the Hoxa-13 and Hoxd-13 mutations on morphogenesis of the terminal parts of the digestive and urogenital tracts. Development, 124, 4781-91 (1997)
44. Block, K., A. Kardana, P. Igarashi & H. S. Taylor: In utero diethylstilbestrol (DES) exposure alters Hox gene expression in the developing mullerian system. FASEB J, 14, 1101-8 (2000)
45. Morgan, R.: Hox genes: a continuation of embryonic patterning? Trends Genet, 22, 67-9 (2006)
doi:10.1016/j.tig.2005.11.004
http://dx.doi.org/doi:10.1016/j.tig.2005.11.004
46. Gendron, R. L., H. Paradis, H. M. Hsieh-Li, D. W. Lee, S. S. Potter & E. Markoff: Abnormal uterine stromal and glandular function associated with maternal reproductive defects in Hoxa-11 null mice. Biol Reprod, 56, 1097-105 (1997)
doi:10.1095/biolreprod56.5.1097
http://dx.doi.org/doi:10.1095/biolreprod56.5.1097
47. Taylor, H. S., P. Igarashi, D. L. Olive & A. Arici: Sex steroids mediate HOXA11 expression in the human peri-implantation endometrium. J Clin Endocrinol Metab, 84, 1129-35 (1999)
doi:10.1210/jc.84.3.1129
http://dx.doi.org/doi:10.1210/jc.84.3.1129
48. Liu, Y., A. Nifuji, M. Tamura, J. M. Wozney, E. N. Olson & M. Noda: Scleraxis messenger ribonucleic acid is expressed in C2C12 myoblasts and its level is down-regulated by bone morphogenetic protein-2 (BMP2). J Cell Biochem, 67, 66-74 (1997)
doi:10.1002/(SICI)1097-4644(19971001)67:1<66::AID-JCB7>3.0.CO;2-U
http://dx.doi.org/doi:10.1002/(SICI)1097-4644(19971001)67:1<66::AID-JCB7>3.0.CO;2-U
49. Kawa-uchi, T., A. Nifuji, N. Mataga, E. N. Olson, J. Bonaventure, K. Shinomiya, Y. Liu & M. Noda: Fibroblast growth factor downregulates expression of a basic helix-loop-helix-type transcription factor, scleraxis, in a chondrocyte-like cell line, TC6. J Cell Biochem, 70, 468-77 (1998)
doi:10.1002/(SICI)1097-4644(19980915)70:4<468::AID-JCB4>3.0.CO;2-H
http://dx.doi.org/doi:10.1002/(SICI)1097-4644(19980915)70:4<468::AID-JCB4>3.0.CO;2-H
50. Murchison, N. D., B. A. Price, D. A. Conner, D. R. Keene, E. N. Olson, C. J. Tabin & R. Schweitzer: Regulation of tendon differentiation by scleraxis distinguishes force-transmitting tendons from muscle-anchoring tendons. Development, 134, 2697-708 (2007)
doi:10.1242/dev.001933
http://dx.doi.org/doi:10.1242/dev.001933
51. Muir, T., I. Sadler-Riggleman & M. K. Skinner: Role of the basic helix-loop-helix transcription factor, scleraxis, in the regulation of Sertoli cell function and differentiation. Mol Endocrinol, 19, 2164-74 (2005)
doi:10.1210/me.2004-0473
http://dx.doi.org/doi:10.1210/me.2004-0473
52. Pestov, D. G., T. M. Grzeszkiewicz & L. F. Lau: Isolation of growth suppressors from a cDNA expression library. Oncogene, 17, 3187-97 (1998)
doi:10.1038/sj.onc.1202260
http://dx.doi.org/doi:10.1038/sj.onc.1202260
53. Strezoska, Z., D. G. Pestov & L. F. Lau: Functional inactivation of the mouse nucleolar protein Bop1 inhibits multiple steps in pre-rRNA processing and blocks cell cycle progression. J Biol Chem, 277, 29617-25 (2002)
doi:10.1074/jbc.M204381200
http://dx.doi.org/doi:10.1074/jbc.M204381200
54. Troy, P. J., G. S. Daftary, C. N. Bagot & H. S. Taylor: Transcriptional repression of peri-implantation EMX2 expression in mammalian reproduction by HOXA10. Mol Cell Biol, 23, 1-13 (2003)
doi:10.1128/MCB.23.1.1-13.2003
http://dx.doi.org/doi:10.1128/MCB.23.1.1-13.2003
55. Taylor, H. S. & X. Fei: Emx2 regulates mammalian reproduction by altering endometrial cell proliferation. Mol Endocrinol, 19, 2839-46 (2005)
doi:10.1210/me.2005-0130
http://dx.doi.org/doi:10.1210/me.2005-0130
56. Spigoni, G., C. Gedressi & A. Mallamaci: Regulation of Emx2 expression by antisense transcripts in murine cortico-cerebral precursors. PLoS One, 5, e8658 (2010)
doi:10.1371/journal.pone.0008658
http://dx.doi.org/doi:10.1371/journal.pone.0008658
57. Gospodarowicz, D., N. Ferrara, L. Schweigerer & G. Neufeld: Structural characterization and biological functions of fibroblast growth factor. Endocr Rev, 8, 95-114 (1987)
doi:10.1210/edrv-8-2-95
http://dx.doi.org/doi:10.1210/edrv-8-2-95
58. Knee, R. S., S. E. Pitcher & P. R. Murphy: Basic fibroblast growth factor sense (FGF) and antisense (gfg) RNA transcripts are expressed in unfertilized human oocytes and in differentiated adult tissues. Biochem Biophys Res Commun, 205, 577-83 (1994)
doi:10.1006/bbrc.1994.2704
http://dx.doi.org/doi:10.1006/bbrc.1994.2704
59. Murphy, P. R. & R. S. Knee: Identification and characterization of an antisense RNA transcript (gfg) from the human basic fibroblast growth factor gene. Mol Endocrinol, 8, 852-9 (1994)
doi:10.1210/me.8.7.852
http://dx.doi.org/doi:10.1210/me.8.7.852
60. MacFarlane, L. A., Y. Gu, A. G. Casson & P. R. Murphy: Regulation of fibroblast growth factor-2 by an endogenous antisense RNA and by argonaute-2. Mol Endocrinol, 24, 800-12 (2010)
doi:10.1210/me.2009-0367
http://dx.doi.org/doi:10.1210/me.2009-0367
61. Lanz, R. B., N. J. McKenna, S. A. Onate, U. Albrecht, J. Wong, S. Y. Tsai, M. J. Tsai & B. W. O'Malley: A steroid receptor coactivator, SRA, functions as an RNA and is present in an SRC-1 complex. Cell, 97, 17-27 (1999)
62. Shi, Y., M. Downes, W. Xie, H. Y. Kao, P. Ordentlich, C. C. Tsai, M. Hon & R. M. Evans: Sharp, an inducible cofactor that integrates nuclear receptor repression and activation. Genes Dev, 15, 1140-51 (2001)
doi:10.1101/gad.871201
http://dx.doi.org/doi:10.1101/gad.871201
63. Caretti, G., R. L. Schiltz, F. J. Dilworth, M. Di Padova, P. Zhao, V. Ogryzko, F. V. Fuller-Pace, E. P. Hoffman, S. J. Tapscott & V. Sartorelli: The RNA helicases p68/p72 and the noncoding RNA SRA are coregulators of MyoD and skeletal muscle differentiation. Dev Cell, 11, 547-60 (2006)
doi:10.1016/j.devcel.2006.08.003
http://dx.doi.org/doi:10.1016/j.devcel.2006.08.003
64. Chooniedass-Kothari, S., E. Emberley, M. K. Hamedani, S. Troup, X. Wang, A. Czosnek, F. Hube, M. Mutawe, P. H. Watson & E. Leygue: The steroid receptor RNA activator is the first functional RNA encoding a protein. FEBS Lett, 566, 43-7 (2004)
doi:10.1016/j.febslet.2004.03.104
http://dx.doi.org/doi:10.1016/j.febslet.2004.03.104
65. Lanz, R. B., B. Razani, A. D. Goldberg & B. W. O'Malley: Distinct RNA motifs are important for coactivation of steroid hormone receptors by steroid receptor RNA activator (SRA). Proc Natl Acad Sci U S A, 99, 16081-6 (2002)
doi:10.1073/pnas.192571399
http://dx.doi.org/doi:10.1073/pnas.192571399
66. Losel, R. M., E. Falkenstein, M. Feuring, A. Schultz, H. C. Tillmann, K. Rossol-Haseroth & M. Wehling: Nongenomic steroid action: controversies, questions, and answers. Physiol Rev, 83, 965-1016 (2003)
67. Watashi, K., M. L. Yeung, M. F. Starost, R. S. Hosmane & K. T. Jeang: Identification of small molecules that suppress microRNA function and reverse tumorigenesis. J Biol Chem, 285, 24707-16 (2010)
doi:10.1074/jbc.M109.062976
http://dx.doi.org/doi:10.1074/jbc.M109.062976
68. Seim, I., S. L. Carter, A. C. Herington & L. K. Chopin: Complex organisation and structure of the ghrelin antisense strand gene GHRLOS, a candidate non-coding RNA gene. BMC Mol Biol, 9, 95 (2008)
doi:10.1186/1471-2199-9-95
http://dx.doi.org/doi:10.1186/1471-2199-9-95
69. Loebel, D. A., B. Tsoi, N. Wong & P. P. Tam: A conserved noncoding intronic transcript at the mouse Dnm3 locus. Genomics, 85, 782-9 (2005)
doi:10.1016/j.ygeno.2005.02.001
http://dx.doi.org/doi:10.1016/j.ygeno.2005.02.001
70. Ingraham, S. E., R. A. Lynch, U. Surti, J. L. Rutter, A. J. Buckler, S. A. Khan, A. G. Menon & P. Lepont: Identification and characterization of novel human transcripts embedded within HMGA2 in t(12;14)(q15;q24.1) uterine leiomyoma. Mutat Res, 602, 43-53 (2006)
doi:10.1016/j.mrfmmm.2006.07.007
http://dx.doi.org/doi:10.1016/j.mrfmmm.2006.07.007
71. Guttman, M., I. Amit, M. Garber, C. French, M. F. Lin, D. Feldser, M. Huarte, O. Zuk, B. W. Carey, J. P. Cassady, M. N. Cabili, R. Jaenisch, T. S. Mikkelsen, T. Jacks, N. Hacohen, B. E. Bernstein, M. Kellis, A. Regev, J. L. Rinn & E. S. Lander: Chromatin signature reveals over a thousand highly conserved large non-coding RNAs in mammals. Nature, 458, 223-7 (2009)
doi:10.1038/nature07672
http://dx.doi.org/doi:10.1038/nature07672
72. Mahmoudi, S., S. Henriksson, M. Corcoran, C. Mendez-Vidal, K. G. Wiman & M. Farnebo: Wrap53, a natural p53 antisense transcript required for p53 induction upon DNA damage. Mol Cell, 33, 462-71 (2009)
doi:10.1016/j.molcel.2009.01.028
http://dx.doi.org/doi:10.1016/j.molcel.2009.01.028
73. Yue, X., J. C. Schwartz, Y. Chu, S. T. Younger, K. T. Gagnon, S. Elbashir, B. A. Janowski & D. R. Corey: Transcriptional regulation by small RNAs at sequences downstream from 3' gene termini. Nat Chem Biol, 6, 621-9 (2010)
doi:10.1038/nchembio.400
http://dx.doi.org/doi:10.1038/nchembio.400
74. Thrash-Bingham, C. A. & K. D. Tartof: aHIF: a natural antisense transcript overexpressed in human renal cancer and during hypoxia. J Natl Cancer Inst, 91, 143-51 (1999)
doi:10.1093/jnci/91.2.143
http://dx.doi.org/doi:10.1093/jnci/91.2.143
75. Coudert, A. E., L. Pibouin, B. Vi-Fane, B. L. Thomas, M. Macdougall, A. Choudhury, B. Robert, P. T. Sharpe, A. Berdal & F. Lezot: Expression and regulation of the Msx1 natural antisense transcript during development. Nucleic Acids Res, 33, 5208-18 (2005)
doi:10.1093/nar/gki831
http://dx.doi.org/doi:10.1093/nar/gki831
76. Wu, S. C., E. M. Kallin & Y. Zhang: Role of H3K27 methylation in the regulation of lncRNA expression. Cell Res, 20, 1109-16 (2010)
doi:10.1038/cr.2010.114
http://dx.doi.org/doi:10.1038/cr.2010.114
77. Haas, B. J. & M. C. Zody: Advancing RNA-Seq analysis. Nat Biotechnol, 28, 421-3 (2010)
doi:10.1038/nbt0510-421
http://dx.doi.org/doi:10.1038/nbt0510-421
78. Kim, T. K., M. Hemberg, J. M. Gray, A. M. Costa, D. M. Bear, J. Wu, D. A. Harmin, M. Laptewicz, K. Barbara-Haley, S. Kuersten, E. Markenscoff-Papadimitriou, D. Kuhl, H. Bito, P. F. Worley, G. Kreiman & M. E. Greenberg: Widespread transcription at neuronal activity-regulated enhancers. Nature, 465, 182-7 (2010)
doi:10.1038/nature09033
http://dx.doi.org/doi:10.1038/nature09033
79. Seila, A. C., J. M. Calabrese, S. S. Levine, G. W. Yeo, P. B. Rahl, R. A. Flynn, R. A. Young & P. A. Sharp: Divergent transcription from active promoters. Science, 322, 1849-51 (2008)
doi:10.1126/science.1162253
http://dx.doi.org/doi:10.1126/science.1162253
80. Trujillo, R. D., S. B. Yue, Y. Tang, W. E. O'Gorman & C. Z. Chen: The potential functions of primary microRNAs in target recognition and repression. EMBO J, 29, 3272-85 (2010)
doi:10.1038/emboj.2010.208
http://dx.doi.org/doi:10.1038/emboj.2010.208
Abbreviations: ncRNAs (non-coding RNAs), miRNAs (microRNAs), siRNAs (small interfering RNAs), piRNAs (PIWI-interacting RNAs), ESCs (endometrial stromal cells), NSRs (nuclear steroid receptors).
Key Words: Implantation, Uterus, ncRNA, microRNA, Review
Send correspondence to: Zeng-Ming Yang, School of Life Science, Xiamen University, Xiamen 361005, China. Tel.: 86-592-2186823, Fax: 86-592-2186823, E-mail: zmyang@xmu.edu.cn