Emerging Role of MicroRNAs in Reproductive Medicine

Nasser Chegini, Ph.D.

Early during summer 2007, I was asked by Dr. Carr to serve as the guest editor for an issue of Seminars in Reproductive Medicine. Selection of a topic relevant to my field of research that would be of interest to general readers of the journal was rather challenging considering the many outstanding articles that were published in previous issues of the journal in past years. However, I became quite convinced that the subject of microRNA (miRNA) in reproduction would be ideal because it has emerged as one of the most active research topics in various fields of biology and medicine in recent years. Comparatively, the miRNA-related research in reproductive biology and medicine is in its early stage of development but will undoubtedly evolve through in-depth investigations. These highly conserved, small, non–protein-coding RNAs have emerged as key regulators of translational efficiency of their complementary target messenger RNAs (mRNAs) from plants to viruses, nematodes, fruit flies, and animals, including humans. In this issue of Seminars in Reproductive Medicine, several articles present evidence regarding the implication of the regulatory function of miRNAs in pituitary, ovarian, uterine, and breast gene expression under normal and pathologic conditions.

During the past decade, large-scale gene expression microarray has generated comprehensive information reflecting the molecular signature of various cells and tissues under normal and disease conditions. Although the acquisition of such large genomic data has been highly informative, it has also created the challenge of how to integrate such knowledge to sort out the individual and interactive mechanisms that regulate the expression of these transcripts resulting in implementation of their appropriate biological functions. Despite considerable progress, the significance of the expression of a large number of these transcripts remains unknown, and because of their diversity, predicting their biological relevance has become more complex than was originally envisioned. Adding to this complexity was the recent discovery of miRNAs and their emergence as one of the major components of the individual and interactive mechanisms that regulate gene expression.

Since the cloning of the first miRNA, lin-4, in Caenorhabditis elegans a decade ago, several hundred more miRNAs have been identified in various species, including human.[1] Genomic analysis revealed that the genes encoding miRNAs are scattered in all chromosomes except the Y chromosome and are estimated to constitute ~1 to 4% of all known genes in higher organisms. In addition, ~50% of the miRNA genes are found in clusters and, due to a high degree of relatedness among them in each cluster, they may have derived from gene duplication. Nearly all the miRNAs identified are conserved in closely related species, or many have homologs in distant species, implying their conserved functions throughout evolution.[2] [3] The primary miRNA transcripts, or pri-miRNAs, are several kilobases long and undergo substantial processing resulting in the generation of a small, 70- to 90-nucleotide (nt) stem-loop precursor miRNA (pre-miRNA) in the nucleus. The pre-miRNAs are transported into the cytoplasm, where they undergo a second cleavage by the cytoplasmic RNase III, Dicer, generating a double-stranded miRNA duplex.[4] [5] [6] The double-stranded miRNA duplex unwinds and forms a single-strand, mature miRNA that incorporates into the RNA-induced silencing complex (RISC) and through complementary interactions with target genes regulates gene expression mostly but not always through translational repression.[7] [8]

Many putative miRNAs have been identified and/or predicted in the genomes of different species, including 580 identified and more than 1500 predicted in humans. After their identification or prediction, miRNAs are assigned a sequential number; that is, miRNA-1 (miR-1), miR-2, miR-3, and so forth. Numerical and alphabetical suffixes are used to distinguish miRNAs that are generated from more than one locus such as miR-1–1 and miR-1–2, and so forth, or to identify miRNAs that differ by a small number of bases such as miR-23a and miR-23b, respectively. miRNAs with a hairpin precursor giving rise to two miRNAs are distinguished by the arm they are generated on, that is, miR-17–5p (5′ arm) and miR-17–3p (3′ arm), and an asterisk identifies the less expressed species, such as miR-199a*.[1] [2] [3] [4] [5] [6] The specificity of miRNAs is dictated by at least 6 to 7 nt that bind to the 3′ untranslated region (3′ UTR) of their target mRNAs. As a result, a single miRNA can potentially target hundreds of genes, or a single gene could be a potential target of many different miRNAs. The biological significance of such diversity is unclear; however, it may serve as a mechanism to achieve different levels of gene regression possibly based on a cell-specific functional requirement. As such, depending on the degree of complementary sequence homology with their target genes, binding of several miRNAs may become necessary to establish full regression of specific target genes.[9] [10] Because of this complexity, to date ~70 to 100 specific gene targets have been functionally verified and experimentally confirmed.

Accumulated data has been generated regarding the expression of many miRNAs in several normal cells and tissues, with aberrant expression of a specific number of them in various disorders, more specifically cancers.[11] [12] [13] To provide insight into the diverse role of miRNAs in reproductive organs, for this issue of the journal several of my colleagues were invited to review the emerging field of miRNAs research in their respective areas of expertise. Maria Chiara Zatelli and Ettore C. degli Uberti discuss the expression and possible role of miRNAs in the pituitary. Their research team was the first to report the expression of miRNAs in pituitary adenomas and correlate miRNA expression with tumor histotype, differentiating microadenomas from macroadenomas and treated from nontreated patients compared with normal pituitary.[14] Clearly, pituitary gonadotropins are essential for many of the ovarian biological activities and subsequently other reproductive tissue physiologic functions. Han Zhao and Aleksandar Rajkovic present evidence for the expression and involvement of miRNAs during ovarian development. Their research team has recently reported that the lack of Nobox perturbs global expression of genes preferentially expressed in oocytes as well as ovarian expression of miRNA compared with that in wild-type ovaries.[15] Whereas the potential regulatory functions of miRNAs in human ovary remain to be elucidated, Tannaz Toloubeydokhti and colleagues discuss such a possibility by incorporating data obtained from mice during oogenesis and ovarian development and observe the relevance of miRNA expression to ovarian physiology and altered expression in ovarian cancer and polycystic ovary syndrome in humans. In addition, the authors discuss the expression of several of these miRNAs in granulosa/cumulus cells obtained from women undergoing oocyte retrieval for assisted reproduction and their potential regulatory function in gene expression essential for sex steroids biosynthesis and other ovarian physiologic functions.

Whether regulated independently or through an ovarian steroid–dependent manner, gene expression in uterine and other ovarian steroid target tissues is subjected to transcriptional and translational regulation. In this regard, Qun Pan discusses the result of miRNA expression signature and regulatory functions in the endometrium during normal and disease states such as endometriosis, endometrial cancer, and dysfunctional uterine bleeding.[16] This is followed by an article from Warren B. Nothnick that describes the role of miRNAs in regulation of uterine matrix metalloproteinase-9, a key enzyme in tissue turnover and angiogenesis. Uterine fibroids, which occur in a substantial number of women during their reproductive years, are also ovarian steroid sensitive tumors that tend to develop more frequently in African Americans than they do in other ethnic groups. The expression and potential regulatory function of miRNAs in the pathogenesis of leiomyomas is presented by Xiaoping Luo, and Jian-Jun Wei and Patricia Soteropoulos discuss miRNAs as a new tool for biomedical risk assessment and target identification in uterine leiomyomas.[17] [18] To conclude this issue, Brian D. Adams and colleagues present evidence for the involvement of miRNAs in pathogenesis of breast cancer with specific reference to estrogen receptor regulation by miR-206.[19]

It has become clear that miRNAs serve as key regulators of gene expression, and our understanding of their functional relevance in normal biological and disease processes is exponentially growing. We are just beginning to unveil the biological significance of miRNA expression and function in reproductive tract cells and tissues with many major challenges to uncover. These include, but are not limited to, hormonal regulation of miRNA expression and functional regulation of specific target genes with critical role in various reproductive tract tissues. Additionally, assessing the expression profile of miRNAs along with their target genes would allow us to identify if aberrantly expressed miRNA is associated with establishment and progression of a particular reproductive disorder or if the changes are the consequence of disease in comparison with normal tissue counterpart. It is also necessary to elucidate what locally expressed factors, specific or nonspecific, modulate the miRNA target recognition and repression and if these activities are cell and tissue specific. There is also a need to determine the specific expression signature of miRNAs in response to hormonal therapies for various reproductive abnormalities, reproductive aging, specifically in the case of the ovary, as well as transition from normal to cancer states. Interestingly, the miRNA signature has been a much more reliable discriminatory value compared with mRNA profiling between normal and cancer tissues and different tumor subtypes. As discussed, in pituitary and breast cancer as well as in several other tumors, the pattern of miRNA expression correlated well with pathologic features of the tumor indicating the possible prognostic use of miRNA profiling along with other well-known prognostic features.[12] [13] Collectively, such progression in miRNAs research would not only allow us to uncover the fine tuning of gene expression regulation and new regulatory mechanisms,[20] but also it may lead to complementary approaches to define the diagnostic usefulness of miRNAs in various reproductive disorders including cancers.

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