bims-cebooc 2024-07-28 papers

bims-cebooc

Biomed News

on Cell biology of oocytes

Issue of 2024–07–28
fourteen papers selected by
Gabriele Zaffagnini, Centre for Genomic Regulation

Sex-specific DNA-replication in the early mammalian embryo.
Aging-associated reduction of chromosomal histones in mammalian oocytes.
Analyzing embryo dormancy at single-cell resolution reveals dynamic transcriptional responses and activation of integrin-Yap/Taz prosurvival signaling.
Tolerance thresholds underlie responses to DNA damage during germline development.
Maternal regulation of the vertebrate oocyte-to-embryo transition.
Deletion of Fbxw7 in oocytes causes follicle loss and premature ovarian insufficiency in mice.
Three-Dimensionally Printed Agarose Micromold Supports Scaffold-Free Mouse Ex Vivo Follicle Growth, Ovulation, and Luteinization.
Decoding molecular features of bovine oocyte fate during antral follicle growth via single-cell multi-omics analysis.
Mitochondria as therapeutic targets in assisted reproduction.
Mitochondria as determinants of reproductive senescence and competence: implications for diagnosis of embryo competence in assisted reproduction.
Type III adenylyl cyclase is essential for follicular development in female mice and their reproductive lifespan.
Oocyte-specific EXOC5 expression is required for mouse oogenesis and folliculogenesis.
Chromosomal missegregation and aberrant embryo development in repro57 female mice with Rnf212 homozygous mutation.
Zinc transporter ZnT3/Slc30a3 has a potential role in zinc ion influx in mouse oocytes.

Nat Commun. 2024 Jul 27. 15(1): 6323

Sex-specific DNA-replication in the early mammalian embryo.

Jason Alexander Halliwell, Javier Martin-Gonzalez, Adnan Hashim, John Arne Dahl, Eva R Hoffmann, Mads Lerdrup.

The timing of DNA replication in mammals is crucial for minimizing errors and influenced by genome usage and chromatin states. Replication timing in the newly formed mammalian embryo remains poorly understood. Here, we have investigated replication timing in mouse zygotes and 2-cell embryos, revealing that zygotes lack a conventional replication timing program, which then emerges in 2-cell embryos. This program differs from embryonic stem cells and generally correlates with transcription and genome compartmentalization of both parental genomes. However, consistent and systematic differences existed between the replication timing of the two parental genomes, including considerably later replication of maternal pericentromeric regions compared to paternal counterparts. Moreover, maternal chromatin modified by Polycomb Repressive Complexes in the oocyte, undergoes early replication, despite belonging to the typically late-replicating B-compartment of the genome. This atypical and asynchronous replication of the two parental genomes may advance our understanding of replication stress in early human embryos and trigger strategies to reduce errors and aneuploidies.

DOI: https://doi.org/10.1038/s41467-024-50727-w
Genes Cells. 2024 Jul 23.

Aging-associated reduction of chromosomal histones in mammalian oocytes.

Masashi Mori, Manami Koshiguchi, Osamu Takenouchi, Mei A Mukose, Hinako M Takase, Tappei Mishina, Hailiang Mei, Miho Kihara, Takaya Abe, Azusa Inoue, Tomoya S Kitajima.

  Mammalian oocytes undergo a long-term meiotic arrest that can last for almost the entire reproductive lifespan. This arrest occurs after DNA replication and is prolonged with age, which poses a challenge to oocytes in maintaining replication-dependent chromosomal proteins required for the completion of meiosis. In this study, we show that chromosomal histones are reduced with age in mouse oocytes. Both types of histone H3 variants, replication-dependent H3.1/H3.2 and replication-independent H3.3, decrease with age. Aging-associated histone reduction is associated with transcriptomic features that are caused by genetic depletion of histone H3.3. Neither the genetic reduction of chromosomal H3.1/H3.2 nor H3.3 accelerates the aging-associated increase in premature chromosome separation that causes meiotic segregation errors. We suggest that aging-associated reduction of chromosomal histones is linked to several transcriptomic abnormalities but does not significantly contribute to errors in meiotic chromosome segregation during the reproductive lifespan of mice.

Keywords:  aging; chromosome; meiosis; oocyte

DOI:  https://doi.org/10.1111/gtc.13146
Cell Stem Cell. 2024 Jul 18. pii: S1934-5909(24)00250-9. [Epub ahead of print]

Analyzing embryo dormancy at single-cell resolution reveals dynamic transcriptional responses and activation of integrin-Yap/Taz prosurvival signaling.

Rui Chen, Rui Fan, Fei Chen, Niraimathi Govindasamy, Heike Brinkmann, Martin Stehling, Ralf H Adams, Hyun-Woo Jeong, Ivan Bedzhov.

  Embryonic diapause is a reproductive adaptation that enables some mammalian species to halt the otherwise continuous pace of embryonic development. In this dormant state, the embryo exploits poorly understood regulatory mechanisms to preserve its developmental potential for prolonged periods of time. Here, using mouse embryos and single-cell RNA sequencing, we molecularly defined embryonic diapause at single-cell resolution, revealing transcriptional dynamics while the embryo seemingly resides in a state of suspended animation. Additionally, we found that the dormant pluripotent cells rely on integrin receptors to sense their microenvironment and preserve their viability via Yap/Taz-mediated prosurvival signaling.

Keywords:  Taz; Yap; embryo dormancy; embryonic diapause; embryonic stem cells; epiblast; extracellular matrix; integrin; pluripotency; single-cell RNA sequencing

DOI:  https://doi.org/10.1016/j.stem.2024.06.015
Genes Dev. 2024 Jul 25.

Tolerance thresholds underlie responses to DNA damage during germline development.

Gloria Jansen, Daniel Gebert, Tharini Ravindra Kumar, Emily Simmons, Sarah Murphy, Felipe Karam Teixeira.

  Selfish DNA modules like transposable elements (TEs) are particularly active in the germline, the lineage that passes genetic information across generations. New TE insertions can disrupt genes and impair the functionality and viability of germ cells. However, we found that in P-M hybrid dysgenesis in Drosophila, a sterility syndrome triggered by the P-element DNA transposon, germ cells harbor unexpectedly few new TE insertions despite accumulating DNA double-strand breaks (DSBs) and inducing cell cycle arrest. Using an engineered CRISPR-Cas9 system, we show that generating DSBs at silenced P-elements or other noncoding sequences is sufficient to induce germ cell loss independently of gene disruption. Indeed, we demonstrate that both developing and adult mitotic germ cells are sensitive to DSBs in a dosage-dependent manner. Following the mitotic-to-meiotic transition, however, germ cells become more tolerant to DSBs, completing oogenesis regardless of the accumulated genome damage. Our findings establish DNA damage tolerance thresholds as crucial safeguards of genome integrity during germline development.

Keywords:  CRISPR–Cas9; DNA damage; Drosophila melanogaster; P-element; double-strand break; genome integrity; germline development; hybrid dysgenesis; meiosis; transposable element

DOI:  https://doi.org/10.1101/gad.351701.124
PLoS Genet. 2024 Jul 25. 20(7): e1011343

Maternal regulation of the vertebrate oocyte-to-embryo transition.

Ricardo Fuentes, Florence L Marlow, Elliott W Abrams, Hong Zhang, Manami Kobayashi, Tripti Gupta, Lee D Kapp, Zachary DiNardo, Ronald Heller, Ruth Cisternas, Priscila García-Castro, Fabián Segovia-Miranda, Felipe Montecinos-Franjola, William Vought, Charles E Vejnar, Antonio J Giraldez, Mary C Mullins.

Maternally-loaded factors in the egg accumulate during oogenesis and are essential for the acquisition of oocyte and egg developmental competence to ensure the production of viable embryos. However, their molecular nature and functional importance remain poorly understood. Here, we present a collection of 9 recessive maternal-effect mutants identified in a zebrafish forward genetic screen that reveal unique molecular insights into the mechanisms controlling the vertebrate oocyte-to-embryo transition. Three genes, over easy, p33bjta, poached and black caviar, were found to control initial steps in yolk globule sizing and protein cleavage during oocyte maturation that act independently of nuclear maturation. The krang, kazukuram, p28tabj, and spotty genes play distinct roles in egg activation, including cortical granule biology, cytoplasmic segregation, the regulation of microtubule organizing center assembly and microtubule nucleation, and establishing the basic body plan. Furthermore, we cloned two of the mutant genes, identifying the over easy gene as a subunit of the Adaptor Protein complex 5, Ap5m1, which implicates it in regulating intracellular trafficking and yolk vesicle formation. The novel maternal protein Krang/Kiaa0513, highly conserved in metazoans, was discovered and linked to the function of cortical granules during egg activation. These mutant genes represent novel genetic entry points to decipher the molecular mechanisms functioning in the oocyte-to-embryo transition, fertility, and human disease. Additionally, our genetic adult screen not only contributes to the existing knowledge in the field but also sets the basis for future investigations. Thus, the identified maternal genes represent key players in the coordination and execution of events prior to fertilization.

DOI: https://doi.org/10.1371/journal.pgen.1011343
J Cell Mol Med. 2024 Jun;28(12): e18487

Deletion of Fbxw7 in oocytes causes follicle loss and premature ovarian insufficiency in mice.

Huihui Zhao, Hanbin Zhang, Yuxia Zhou, Ling Shuai, Zhenguo Chen, Liping Wang.

  Premature ovarian insufficiency (POI) is one of the important causes of female infertility. Yet the aetiology for POI is still elusive. FBXW7 (F-box with 7 tandem WD) is one of the important components of the Skp1-Cullin1-F-box (SCF) E3 ubiquitin ligase. FBXW7 can regulate cell growth, survival and pluripotency through mediating ubiquitylation and degradation of target proteins via triggering the ubiquitin-proteasome system, and is associated with tumorigenesis, haematopoiesis and testis development. However, evidence establishing the function of FBXW7 in ovary is still lacking. Here, we showed that FBXW7 protein level was significantly decreased in the ovaries of the cisplatin-induced POI mouse model. We further showed that mice with oocyte-specific deletion of Fbxw7 demonstrated POI, characterized with folliculogenic defects, early depletion of follicle reserve, disordered hormonal secretion, ovarian dysfunction and female infertility. Impaired oocyte-GCs communication, manifested as down-regulation of connexin 37, may contribute to follicular development failure in the Fbxw7-mutant mice. Furthermore, single-cell RNA sequencing and in situ hybridization results indicated an accumulation of Clu and Ccl2 transcripts, which may alter follicle microenvironment deleterious to oocyte development and accelerate POI. Our results establish the important role of Fbxw7 in folliculogenesis and ovarian function, and might provide valuable information for understanding POI and female infertility.

Keywords:   Fbxw7 ; follicle loss; ovarian dysfunction; premature ovarian insufficiency

DOI:  https://doi.org/10.1111/jcmm.18487
Bioengineering (Basel). 2024 Jul 15. pii: 719. [Epub ahead of print]11(7):

Three-Dimensionally Printed Agarose Micromold Supports Scaffold-Free Mouse Ex Vivo Follicle Growth, Ovulation, and Luteinization.

Emily J Zaniker, Prianka H Hashim, Samuel Gauthier, James A Ankrum, Hannes Campo, Francesca E Duncan.

  Ex vivo follicle growth is an essential tool, enabling interrogation of folliculogenesis, ovulation, and luteinization. Though significant advancements have been made, existing follicle culture strategies can be technically challenging and laborious. In this study, we advanced the field through development of a custom agarose micromold, which enables scaffold-free follicle culture. We established an accessible and economical manufacturing method using 3D printing and silicone molding that generates biocompatible hydrogel molds without the risk of cytotoxicity from leachates. Each mold supports simultaneous culture of multiple multilayer secondary follicles in a single focal plane, allowing for constant timelapse monitoring and automated analysis. Mouse follicles cultured using this novel system exhibit significantly improved growth and ovulation outcomes with comparable survival, oocyte maturation, and hormone production profiles as established three-dimensional encapsulated in vitro follicle growth (eIVFG) systems. Additionally, follicles recapitulated aspects of in vivo ovulation physiology with respect to their architecture and spatial polarization, which has not been observed in eIVFG systems. This system offers simplicity, scalability, integration with morphokinetic analyses of follicle growth and ovulation, and compatibility with existing microphysiological platforms. This culture strategy has implications for fundamental follicle biology, fertility preservation strategies, reproductive toxicology, and contraceptive drug discovery.

Keywords:  3D printing; agarose; bioengineering; engineered microenvironment; follicle culture; ovary; ovulation; reproductive biology; scaffolds

DOI:  https://doi.org/10.3390/bioengineering11070719
Biol Reprod. 2024 Jul 26. pii: ioae114. [Epub ahead of print]

Decoding molecular features of bovine oocyte fate during antral follicle growth via single-cell multi-omics analysis.

Qiang Zhang, Jingyao Zhang, Gang Chang, Kun Zhao, Yujun Yao, Li Liu, Zihuan Du, Yanping Wang, Xingrong Guo, Zongsheng Zhao, Weibin Zeng, Shuai Gao.

  Antral follicle size is a useful predictive marker of the competency of enclosed oocytes for yielding an embryo following in vitro maturation and fertilization. However, the molecular mechanisms underpinning oocyte developmental potential during bovine antral follicle growth are still unclear. Here, we used a modified single-cell multi-omics approach to analyze the transcriptome, DNA methylome and chromatin accessibility in parallel for oocytes and cumulus cells collected from bovine antral follicles of different sizes. Transcriptome profiling identified three types of oocytes (Small, Medium and Large) that underwent different developmental trajectories, with Large oocytes exhibiting the largest average follicle size and characteristics resembling metaphase-II oocytes. Differential expression analysis and real-time PCR assay showed that most replication-dependent histone genes were highly expressed in Large oocytes. The joint analysis of multi-omics data revealed that the transcription of 20 differentially expressed genes in Large oocytes was associated with both DNA methylation and chromatin accessibility. In addition, oocyte-cumulus interaction analysis showed that inflammation, DNA damage, and p53 signaling pathways were active in Small oocytes, which had the smallest average follicle sizes. We further confirmed that p53 pathway inhibition in in vitro maturation experiments using oocytes obtained from small antral follicles could improve the quality of oocytes and increased the blastocyte rate after in vitro fertilization and culture. Our work provides new insights into the intricate orchestration of bovine oocyte fate determination during antral folliculogenesis, which is instrumental for optimizing in vitro maturation techniques to optimize oocyte quality.

Keywords:  antral follicle; cattle; oocyte; single-cell multi-omics sequencing

DOI:  https://doi.org/10.1093/biolre/ioae114
Hum Reprod. 2024 Jul 27. pii: deae170. [Epub ahead of print]

Mitochondria as therapeutic targets in assisted reproduction.

Raziye Melike Yildirim, Emre Seli.

  Mitochondria are essential organelles with specialized functions, which play crucial roles in energy production, calcium homeostasis, and programmed cell death. In oocytes, mitochondrial populations are inherited maternally and are vital for developmental competence. Dysfunction in mitochondrial quality control mechanisms can lead to reproductive failure. Due to their central role in oocyte and embryo development, mitochondria have been investigated as potential diagnostic and therapeutic targets in assisted reproduction. Pharmacological agents that target mitochondrial function and show promise in improving assisted reproduction outcomes include antioxidant coenzyme Q10 and mitoquinone, mammalian target of rapamycin signaling pathway inhibitor rapamycin, and nicotinamide mononucleotide. Mitochondrial replacement therapies (MRTs) offer solutions for infertility and mitochondrial disorders. Autologous germline mitochondrial energy transfer initially showed promise but failed to demonstrate significant benefits in clinical trials. Maternal spindle transfer (MST) and pronuclear transfer hold potential for preventing mitochondrial disease transmission and improving oocyte quality. Clinical trials of MST have shown promising outcomes, but larger studies are needed to confirm safety and efficacy. However, ethical and legislative challenges complicate the widespread implementation of MRTs.

Keywords:   in vitro fertilization; ROS; antioxidants; embryo; maternal spindle transfer (MST); mitochondrial replacement therapies; mtDNA; oocyte; pronuclear transfer (PNT)

DOI:  https://doi.org/10.1093/humrep/deae170
Hum Reprod. 2024 Jul 27. pii: deae171. [Epub ahead of print]

Mitochondria as determinants of reproductive senescence and competence: implications for diagnosis of embryo competence in assisted reproduction.

Raziye Melike Yildirim, Emre Seli.

  Mitochondria are commonly recognized as the powerhouses of the cell, primarily responsible for energy production through oxidative phosphorylation. Alongside this vital function, they also play crucial roles in regulating calcium signaling, maintaining membrane potential, and modulating apoptosis. Their involvement in various cellular pathways becomes particularly evident during oogenesis and embryogenesis, where mitochondrial quantity, morphology, and distribution are tightly controlled. The efficiency of the mitochondrial network is maintained through multiple quality control mechanisms that are essential for reproductive success. These include mitochondrial unfolded protein response, mitochondrial dynamics, and mitophagy. Not surprisingly, mitochondrial dysfunction has been implicated in infertility and ovarian aging, prompting investigation into mitochondria as diagnostic and therapeutic targets in assisted reproduction. To date, mitochondrial DNA copy number in oocytes, cumulus cells, and trophectoderm biopsies, and fluorescent lifetime imaging microscopy-based assessment of NADH and flavin adenine dinucleotide content have been explored as potential predictors of embryo competence, yielding limited success. Despite challenges in the clinical application of mitochondrial diagnostic strategies, these enigmatic organelles have a significant impact on reproduction, and their potential role as diagnostic targets in assisted reproduction is likely to remain an active area of investigation in the foreseeable future.

Keywords:  FLIM; assisted reproduction outcomes; embryo; embryo viability; in vitro fertilization; mitochondrial dynamics; mtDNA

DOI:  https://doi.org/10.1093/humrep/deae171
iScience. 2024 Jul 19. 27(7): 110293

Type III adenylyl cyclase is essential for follicular development in female mice and their reproductive lifespan.

Baofang Zheng, Xiaoyu Hu, Yuanhui Hu, Sheng Dong, Xin Xiao, Haoming Qi, Yongdi Wang, Weina Wang, Zhenshan Wang.

  Premature ovarian failure (POF) is a complex and heterogeneous disease that causes infertility and subfertility. However, the molecular mechanism of POF has not been fully elucidated. Here, we show that the loss of adenylyl cyclase III (Adcy3) in female mice leads to POF and a shortened reproductive lifespan. We found that Adcy3 is abundantly expressed in mouse oocytes. Adcy3 knockout mice exhibited the excessive activation of primordial follicles, progressive follicle loss, follicular atresia, and ultimately POF. Mechanistically, we found that mitochondrial oxidative stress in oocytes significantly increased with age in Adcy3-deficient mice and was accompanied by oocyte apoptosis and defective folliculogenesis. In contrast, compared with wild-type female mice, humanized ADCY3 knock-in female mice exhibited improved fertility with age. Collectively, these results reveal that the previously unrecognized Adcy3 signaling pathway is tightly linked to female ovarian aging, providing potential pharmaceutical targets for preventing and treating POF.

Keywords:  molecular biology; molecular physiology; physiology; reproductive medicine

DOI:  https://doi.org/10.1016/j.isci.2024.110293
Mol Hum Reprod. 2024 Jul 22. pii: gaae026. [Epub ahead of print]

Oocyte-specific EXOC5 expression is required for mouse oogenesis and folliculogenesis.

Hongwen Wu, Hieu Nguyen, Prianka H Hashim, Ben Fogelgren, Francesca E Duncan, W Steven Ward.

  EXOC5 is a crucial component of a large multi-subunit tethering complex, the exocyst complex, that is required for fusion of secretory vesicles with the plasma membrane. Exoc5 deleted mice die as early embryos. Therefore, to determine the role of EXOC5 in follicular and oocyte development, it was necessary to produce a conditional knockout (cKO), Zp3-Exoc5-CKO, in which Exoc5 was deleted only in oocytes. The first wave of folliculogenesis appeared histologically normal and progressed to the antral stage. However, after in vitro fertilization with normal sperm, oocytes collected from the first wave (superovulated 21-day old cKO mice) were shown to be developmentally incompetent. Adult follicular waves did not progress beyond the secondary follicle stage where they underwent apoptosis. Female cKO mice were infertile. Overall, these data suggest that the first wave of folliculogenesis is less sensitive to oocyte-specific loss of Exoc5, but the resulting gametes have reduced developmental competence. In contrast, subsequent waves of folliculogenesis require oocyte-specific Exoc5 for development past the preantral follicle stage. The Zp3-Exoc5-CKO mouse provides a model for disrupting folliculogenesis that also enables the separation between the first and subsequent waves of folliculogenesis.

Keywords:  EXOC5; female infertility; follicular apoptosis; folliculogenesis; ovarian development; ovarian fibrosis

DOI:  https://doi.org/10.1093/molehr/gaae026
Reproduction. 2024 Jul 01. pii: REP-24-0030. [Epub ahead of print]

Chromosomal missegregation and aberrant embryo development in repro57 female mice with Rnf212 homozygous mutation.

Nanami Sono, Mone Takeshita, Mizuho Chikushi, Saki Nakashima, Shoko Miyawaki, Misaki Wakamatsu, Yasuhiro Fujiwara, Tetsuo Kunieda, Junko Otsuki.

Repro57 mice, induced with N-ethyl-N-nitrosourea and harboring a mutation in the Rnf212 gene, exhibit infertility in both homozygous mutant males and females. Rnf212 plays a crucial role in recombination and crossover designation. In male repro57 homozygous mutants, spermatocytes often degenerate during late prophase, and mature spermatozoa are absent in the seminiferous epithelium, indicating arrested spermatogenesis as the cause of infertility. Despite reports of infertility in Rnf212 knockout female mice, the specific mechanisms underlying infertility in female repro57 homozygous mutants remain elusive. This study investigates the chromosomal and kinetochore patterns of mature oocytes and their developmental potential following in vitro fertilization in female repro57 homozygous mutant mice. While all wild-type oocytes progress to metaphase II and exhibit euploidy, all repro57 homozygous mutant mouse oocytes display aneuploidy. Additionally, kinetochore distances in repro57 homozygous mutant oocytes exceed those observed in wild-type counterparts. Although no significant differences are noted in fertilization and early embryo development rates between wild-type and repro57 homozygous mutant mice, embryos derived from repro57 homozygous mutants exhibit significantly lower morula and blastocyst rates, accompanied by frequent cytokinesis failure and vacuole formation. These findings suggest that premature segregation of sister chromatids in repro57 homozygous mutant mice adversely impacts later stages of embryo development.

DOI: https://doi.org/10.1530/REP-24-0030
J Reprod Dev. 2024 Jul 23.

Zinc transporter ZnT3/Slc30a3 has a potential role in zinc ion influx in mouse oocytes.

Atsuko Kageyama, Jumpei Terakawa, Shunsuke Takarabe, Hibiki Sugita, Yui Kawata, Junya Ito, Naomi Kashiwazaki.

  Zinc is an essential trace element for various physiological functions, including reproduction. The influx/efflux of zinc ions is regulated by zinc transporters (Zip1-14 and ZnT1-8, 10). However, the precise roles of zinc transporters and zinc dynamics in reproductive functions are unknown. In this study, ZnT3/Slc30a3 gene knockout (KO) mice were used to analyze the role of ZnT3. In ZnT3 KO mice, intracellular zinc ions in oocytes/zygotes were significantly reduced compared to those in controls, and free zinc ions did not accumulate in the oocyte cytoplasm. However, fertilization of these oocytes and the average litter size were comparable to those of control mice. Our results suggest that ZnT3 plays an important role in the accumulation of zinc ions in oocytes but not in the developmental ability of mice. ZnT3 KO mice will be useful for examining zinc dynamics in oocytes and other tissues.

Keywords:  Fertility; Zinc; Zinc transporter; ZnT3/Slc30a3

DOI:  https://doi.org/10.1262/jrd.2024-044