Meiotic Cell Cycle Progression in Mouse Oocytes: Role of Cyclins.

All eukaryotic cells, including oocytes, utilize an engine called cyclin-dependent kinase (Cdk) to drive the cell cycle. Cdks are activated by a co-factor called cyclin, which regulates their activity. The key Cdk-cyclin complex that regulates the oocyte cell cycle is known as Cdk1-cyclin B1. Recent studies have elucidated the roles of other cyclins, such as B2, B3, A2, and O, in oocyte cell cycle regulation. This review aims to discuss the recently discovered roles of various cyclins in mouse oocyte cell cycle regulation in accordance with the sequential progression of the cell cycle. In addition, this review addresses the translation and degradation of cyclins to modulate the activity of Cdks. Overall, the literature indicates that each cyclin performs unique and redundant functions at various stages of the cell cycle, while their expression and degradation are tightly regulated. Taken together, this review provides new insights into the regulatory role and function of cyclins in oocyte cell cycle progression.

Coenzyme Q10 ameliorates the quality of mouse oocytes during in vitro culture.

Oxidative stress causes several diseases and dysfunctions in cells, including oocytes. Clearly, oxidative stress influences oocyte quality during in vitro maturation and fertilization. Here we tested the ability of coenzyme Q10 (CoQ10) to reduce reactive oxygen species (ROS) and improve mouse oocyte quality during in vitro culture. Treatment with 50 µM CoQ10 efficiently reduced ROS levels in oocytes cultured in vitro. The fertilizable form of an oocyte usually contains a cortical granule-free domain (CGFD). CoQ10 enhanced the ratio of CGFD-oocytes from 35% to 45%. However, the hardening of the zona pellucida in oocytes was not affected by CoQ10 treatment. The in vitro maturation capacity of oocytes, which was determined by the first polar body extrusion, was enhanced from 48.9% to 75.7% by the addition of CoQ10 to the culture medium. During the parthenogenesis process, the number of two-cell embryos was increased by CoQ10 from 43.5% to 67.3%. Additionally, treatment with CoQ10 increased the expression of Bcl2 and Sirt1 in cumulus cells. These results suggested that CoQ10 had a positive effect on ROS reduction, maturation rate and two-cell embryo formation in mouse oocyte culture.

Differential Effects of Low and High Radiation Dose Rates on Mouse Spermatogenesis.

The adverse effects of radiation are proportional to the total dose and dose rate. We aimed to investigate the effects of radiation dose rate on different organs in mice. The mice were subjected to low dose rate (LDR, ~3.4 mGy/h) and high dose rate (HDR, ~51 Gy/h) radiation. LDR radiation caused severe tissue toxicity, as observed in the histological analysis of testis. It adversely influenced sperm production, including sperm count and motility, and induced greater sperm abnormalities. The expression of markers of early stage spermatogonial stem cells, such as Plzf, c-Kit, and Oct4, decreased significantly after LDR irradiation, compared to that following exposure of HDR radiation, in qPCR analysis. The compositional ratios of all stages of spermatogonia and meiotic cells, except round spermatid, were considerably reduced by LDR in FACS analysis. Therefore, LDR radiation caused more adverse testicular damage than that by HDR radiation, contrary to the response observed in other organs. Therefore, the dose rate of radiation may have differential effects, depending on the organ; it is necessary to evaluate the effect of radiation in terms of radiation dose, dose rate, organ type, and other conditions.

5'-UTR and ORF elements, as well as the 3'-UTR regulate the translation of Cyclin.

Mammalian oocyte maturation is wholly dependent on the translation of accumulated maternal transcripts. Therefore, measuring the translation of specific genes, especially Ccnb1 and Ccnb2, which are key regulators of the oocyte cell cycle in mice, is essential to monitor oocyte cell cycle progression. For this purpose, almost all previous research has used a reporter construct containing the 3'-untranslated region (UTR) of Ccnb. It is based on the concept that the 3'-UTR is the main modulator of translation. Here, we investigated the expression pattern of Renilla luciferase (RL) reporters combining the 5'-UTR and/or open reading frame (ORF) as well as the 3'-UTR (RL-3', 5'-RL-3', RL-ORF-3', and 5'-RL-ORF-3') of Ccnb1 and Ccnb2 in somatic cells and mouse oocytes. The addition of the 5'-UTR and/or ORF of Ccnb altered the expression of the RL-3' reporter in HEK293T cells and mouse oocytes. The ORF tended to suppress RL expression, whereas the 5'-UTR enhanced the expression in most cases. The increased rate in expression was the highest when only the 3'-UTR of Ccnb1 (RL-3') was used, whereas the 5'-RL-ORF-3' reporter showed a relatively lower increase during oocyte maturation. For Ccnb2, the RL-ORF-3' reporter showed the largest increase, and other reporters exhibited a similar increase in expression during oocyte maturation. Results show that the expression of these genes is modulated not only by the 3'-UTR but also by the 5'-UTR and ORF. Therefore, special caution should be taken when using only the 3'-UTR to monitor the expression of specific genes.

Possible association of G6PC2 and MUC6 induced by low­dose­rate irradiation in mouse intestine with inflammatory bowel disease.

Although there are several types of radiation exposure, it is debated whether low­dose­rate (LDR) irradiation (IR) affects the body. Since the small intestine is a radiation­sensitive organ, the present study aimed to evaluate how it changes when exposed to LDR IR and identify the genes sensitive to these doses. After undergoing LDR (6.0 mGy/h) γ radiation exposure, intestinal RNA from BALB/c mice was extracted 1 and 24 h later. Mouse whole genome microarrays were used to explore radiation­induced transcriptional alterations. Reverse transcription­quantitative (RT­q) PCR was used to examine time­ and dose­dependent radiation responses. The histopathological status of the jejunum in the radiated mouse was not changed by 10 mGy of LDR IR; however, 23 genes were upregulated in response to LDR IR of the jejunum in mice after 1 and 24 h of exposure. Upregulated genes were selected to validate the results of the RNA sequencing analysis for RT­qPCR detection and results showed that only Na+/K+ transporting subunit α4, glucose­6­phosphatase catalytic subunit 2 (G6PC2), mucin 6 (MUC6) and transient receptor potential cation channel subfamily V member 6 levels significantly increased after 24 h of LDR IR. Furthermore, G6PC2 and MUC6 were notable genes induced by LDR IR exposure according to protein expression via western blot analysis. The mRNA levels of G6PC2 and MUC6 were significantly elevated within 24 h under three conditions: i) Exposure to LDR IR, ii) repeated exposure to LDR IR and iii) exposure to LDR IR in the presence of inflammatory bowel disease. These results could contribute to an improved understanding of immediate radiation reactions and biomarker development to identify radiation­susceptible individuals before histopathological changes become noticeable. However, further investigation into the specific mechanisms involving G6PC2 and MUC6 is required to accomplish this.

Low dose rate radiation impairs early follicles in young mice.

Low-dose radiation is generally considered less harmful than high-dose radiation. However, its impact on ovaries remains debated. Since previous reports predominantly employed low-dose radiation delivered at a high dose rate on the ovary, the effect of low-dose radiation at a low dose rate on the ovary remains unknown. We investigated the effect of low-dose ionizing radiation delivered at a low dose rate on murine ovaries. Three- and ten-week-old mice were exposed to 0.1 and 0.5 Gy of radiation at a rate of 6 mGy/h and monitored after 3 and 30 days. While neither body weight nor ovarian area showed significant changes, ovarian cells were damaged, showing apoptosis and a decrease in cell proliferation after exposure to 0.1 and 0.5 Gy radiation. Follicle numbers decreased over time in both age groups proportionally to the radiation dose. Younger mice were more susceptible to radiation damage, as evidenced by decreased follicles in 3-week-old mice after 30 days of 0.1 Gy exposure, while 10-week-old mice showed reduced follicles only following 0.5 Gy exposure. Primordial or primary follicles were the most vulnerable to radiation. These findings suggest that even low-dose radiation, delivered at a low dose rate, can adversely affect ovarian function, particularly in the early follicles of younger mice.

The Translation of Cyclin B1 and B2 is Differentially Regulated during Mouse Oocyte Reentry into the Meiotic Cell Cycle.

Control of protein turnover is critical for meiotic progression. Using RiboTag immunoprecipitation, RNA binding protein immunoprecipitation, and luciferase reporter assay, we investigated how rates of mRNA translation, protein synthesis and degradation contribute to the steady state level of Cyclin B1 and B2 in mouse oocytes. Ribosome loading onto Ccnb1 and Mos mRNAs increases during cell cycle reentry, well after germinal vesicle breakdown (GVBD). This is followed by the translation of reporters containing 3' untranslated region of Mos or Ccnb1 and the accumulation of Mos and Cyclin B1 proteins. Conversely, ribosome loading onto Ccnb2 mRNA and Cyclin B2 protein level undergo minimal changes during meiotic reentry. Degradation rates of Cyclin B1 or B2 protein at the GV stage are comparable. The translational activation of Mos and Ccnb1, but not Ccnb2, mRNAs is dependent on the RNA binding protein CPEB1. Inhibition of Cdk1 activity, but not Aurora A kinase activity, prevents the translation of Mos or Ccnb1 reporters, suggesting that MPF is required for their translation in mouse oocytes. Conversely, Ccnb2 translation is insensitive to Cdk1 inhibition. Thus, the poised state that allows rapid meiotic reentry in mouse GV oocytes may be determined by the differential translational control of two Cyclins.

Post-transcriptional and post-translational regulation during mouse oocyte maturation.

The meiotic process from the primordial stage to zygote in female germ cells is mainly adjusted by post-transcriptional regulation of pre-existing maternal mRNA and post-translational modification of proteins. Several key proteins such as the cell cycle regulator, Cdk1/cyclin B, are post-translationally modified for precise control of meiotic progression. The second messenger (cAMP), kinases (PKA, Akt, MAPK, Aurora A, CaMK II, etc), phosphatases (Cdc25, Cdc14), and other proteins (G-protein coupled receptor, phosphodiesterase) are directly or indirectly involved in this process. Many proteins, such as CPEB, maskin, eIF4E, eIF4G, 4E-BP, and 4E-T, post-transcriptionally regulate mRNA via binding to the cap structure at the 5' end of mRNA or its 3' untranslated region (UTR) to generate a closed-loop structure. The 3' UTR of the transcript is also implicated in post-transcriptional regulation through an association with proteins such as CPEB, CPSF, GLD-2, PARN, and Dazl to modulate poly(A) tail length. RNA interfering is a new regulatory mechanism of the amount of mRNA in the mouse oocyte. This review summarizes information about post-transcriptional and post-translational regulation during mouse oocyte meiotic maturation.