Primordial ovarian follicles in mice form when somatic cells surround individual oocytes. We show that lack of Nobox, an oocyte-specific homeobox gene, accelerates postnatal oocyte loss and abolishes the transition from primordial to growing follicles in mice. Follicles are replaced by fibrous tissue in female mice lacking Nobox in a manner similar to nonsyndromic ovarian failure in women. Genes preferentially expressed in oocytes, including Oct4 and Gdf9, are down-regulated in Nobox-/- mice, whereas ubiquitous genes such as Bmp4, Kit, and Bax remain unaffected. Therefore, Nobox is critical for specifying an oocyte-restricted gene expression pattern essential for postnatal follicle development.

An abnormal embryonic karyotype was found to represent the commonest cause of recurrent miscarriage, and the percentage of cases with recurrent miscarriage of truly unexplained cause was limited to 24.5%.The two groups should be distinguished for both clinical and research purposes.

To identify novel genes involved in early mammalian folliculogenesis, we used the Unigene collection of mouse cDNA libraries to identify unique expressed sequence tags in a newborn mouse ovary cDNA library. Nobox (newborn ovary homeobox-encoding gene) was one of several genes identified by in silico (electronic database) subtraction. We cloned the mouse Nobox cDNA and characterized its genomic organization. The gene spans 14kb and is encoded by eight exons. The Nobox gene maps to proximal chromosome 6 in the mouse, and we identified a portion of the human gene encoding a NOBOX homolog which resides at a syntenic position on chromosome 7q35. Reverse transcriptase polymerase chain reaction and Northern blot analyses show that Nobox is preferentially expressed in the ovary at high levels. In situ hybridization analysis demonstrates that Nobox mRNA is present in primordial and growing oocytes. Nobox is one of the first homeobox-encoding genes preferentially expressed during mammalian folliculogenesis.

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Oocyte meiosis and early mitotic divisions in developing embryos rely on the timely production of cell cycle regulators and their clearance via proteasomal degradation. Ret Finger Protein-Like 4 (Rfpl4), encoding a RING finger-like protein with a B30.2 domain, was discovered during an in silico search for germ cell-specific genes. To study the expression and functions of RFPL4 protein, we performed immunolocalizations and used yeast two-hybrid and other protein-protein interaction assays. Immunohistochemistry and immunofluorescence showed that RFPL4 accumulates in all growing oocytes and quickly disappears during early embryonic cleavage. We used a yeast two-hybrid model to demonstrate that RFPL4 interacts with the E2 ubiquitin-conjugating enzyme HR6A, proteasome subunit ␤ type 1, ubiquitin B, as well as a degradation target protein, cyclin B1. Coimmunoprecipitation analyses of in vitro translated proteins and extracts of transiently cotransfected Chinese hamster ovary (CHO)-K1 cells confirmed these findings. We conclude that, like many RING-finger containing proteins, RFPL4 is an E3 ubiquitin ligase. The specificity of its expression and these interactions suggest that RFPL4 targets cyclin B1 for proteasomal degradation, a key aspect of oocyte cell cycle control during meiosis and the crucial oocyte-to-embryo transition to mitosis.proteolysis ͉ meiotic regulator ͉ maturation promoting factor U biquitination is the major means in eukaryotic cells for targeted protein proteolysis (1). By covalent addition of polyubiquitin to specific proteins, the ubiquitination system regulates protein levels and thereby influences diverse cellular processes. There are three well established types of enzymes involved in ubiquitination, termed E1, E2, and E3. E1 is the ubiquitin-activating enzyme, which forms a thiol-ester linkage with ubiquitin through its active site cysteine. Ubiquitin is subsequently transferred to an E2 ubiquitin-conjugating enzyme. The E3 enzyme is the ubiquitin protein ligase, which transfers ubiquitin from the E2 enzyme to lysines of a specific protein, targeting the protein for degradation by the proteasome. More recently, E4 enzymes have been described that appear to function in ubiquitin chain polymerization (2). Few E1 enzymes, several E2 enzymes, and hundreds of E3 enzymes have been identified. It is the E3 ubiquitin protein ligase that adds specificity to the process by interacting with specific target proteins. Included in the group of E3 enzymes are proteins such as the cancer-associated proteins, anaphase-promoting complex (APC), BRCA1, and MDM2, and the DNA repair proteins, RAD5 and RAD18. Many E3 ubiquitin protein ligases share a common RING (Really Interesting Novel Gene) finger consensus sequence CX 2 CX (9 -39) CX (1-3) HX (2-3) C͞HX 2 CX (4 -48) CX 2 C (reviewed in refs. 3 and 4), and a majority of RING finger-containing proteins have been shown to function as E3 ubiquitin protein ligases.A few related proteins have been identified and termed Ret Finger Protein-Like 1 (RFPL1), RFPL2, and RF...

Aneuploidy in the conceptus or fetus, occurs in 5-10% of all pregnancies and is a common reproductive problem in humans. Most aneuploid conceptuses die in utero, resulting in early pregnancy loss. Causes of recurrent miscarriage may include abnormal chromosomes in either partner, particularly translocations, antiphospholipid antibodies and uterine anomalies. Chromosomal aberrations in parents are a major pre-disposing factor and causative of abortion if carried over to the embryo. The transmission rate in the embryo can be speculated to be about 50%. Embryo morphology, developmental rates, and maternal age are correlated with chromosomal abnormalities. Translocation in either partner is one of the most important causes of recurrent miscarriage and the prognosis of subsequent pregnancy in couples with abnormal embryonic karyotype is poorer than that in couples with normal chromosome karyotypes. As for parents whose karyotypes are normal, the frequency of normal embryonic karyotypes significantly increases with the number of previous abortions and a normal karyotype in a previous pregnancy is a predictor of subsequent miscarriage. Recently, many kinds of genetic polymorphisms have also been found to be associated with recurrent miscarriages. In contrast, preimplantation genetic diagnosis for aneuploidy screening is sometimes performed in patients with unexplained recurrent miscarriages. We review genetic factors as a cause of miscarriage.

The purpose of this noninvasive prenatal testing (NIPT) study was to compare the fetal fraction of singleton gestations by gestational age, maternal characteristics and chromosome-specific aneuploidies as indicated by z-scores. This study was a multicenter prospective cohort study. Test data were collected from women who underwent NIPT by the massively parallel sequencing method. We used sequencing-based fetal fraction calculations in which we estimated fetal DNA fraction by simply counting the number of reads aligned within specific autosomal regions and applying a weighting scheme derived from a multivariate model. Relationships between fetal fractions and gestational age, maternal weight and height, and z-scores for chromosomes 21, 18 and 13 were assessed. A total of 7740 pregnant women enrolled in the study, of which 6993 met the study criteria. As expected, fetal fraction was inversely correlated with maternal weight (P<0.001). The median fetal fraction of samples with euploid result (n=6850) and trisomy 21 (n=70) were 13.7% and 13.6%, respectively. In contrast, the median fetal fraction values for samples with trisomies 18 (n=35) and 13 (n=9) were 11.0% and 8.0%, respectively. The fetal fraction of samples with trisomy 21 NIPT result is comparable to that of samples with euploid result. However, the fetal fractions of samples with trisomies 13 and 18 are significantly lower compared with that of euploid result. We conclude that it may make detecting these two trisomies more challenging.

Aim The purpose of this study was to report the 3‐year experience of a nationwide demonstration project to introduce non‐invasive prenatal testing (NIPT) of maternal plasma for aneuploidy, and review the current status of NIPT in Japan. Methods Tests were conducted to detect aneuploidy in high‐risk pregnant women, and adequate genetic counseling was provided. The clinical data, test results, and pregnancy outcomes were recorded. We discuss the problems of NIPT on the basis of published reports and meta‐analyses. Results From April 2013 to March 2016, 30 613 tests were conducted at 55 medical sites participating in a multicenter clinical study. Among the 30 613 women tested, 554 were positive (1.81%) and 30 021 were negative (98.1%) for aneuploidy. Of the 289, 128, and 44 women who tested positive for trisomies 21, 18, and 13, respectively, and underwent definitive testing, 279 (96.5%), 106 (82.8%), and 28 (63.6%) were determined to have a true‐positive result. For the 13 481 women with negative result and whose progress could be traced, two had a false‐negative result (0.02%). The tests were performed on the condition that a standard level of genetic counseling be provided at hospitals. Conclusion Here, we report on the 3‐year nationwide experience with NIPT in Japan. It is important to establish a genetic counseling system to enable women to make informed decisions regarding prenatal testing. Moreover, a welfare system is warranted to support women who decide to give birth to and raise children with chromosomal diseases.