bims-cebooc Biomed News
on Cell biology of oocytes
Issue of 2024–10–06
thirteen papers selected by
Gabriele Zaffagnini, Centre for Genomic Regulation
  1. An architectural role of specific RNA-RNA interactions in oskar granules.
  2. Nr5a2 is dispensable for zygotic genome activation but essential for morula development.
  3. Rise and SINE: roles of transcription factors and retrotransposons in zygotic genome activation.
  4. Regulation of outer kinetochore assembly during meiosis I and II by CENP-A and KNL-2/M18BP1 in C. elegans oocytes.
  5. Aurora B and Aurora C pools at two chromosomal regions collaboratively maintain chromosome alignment and prevent aneuploidy at the second meiotic division in mammalian oocytes.
  6. The mouse Balbiani body regulates primary oocyte quiescence via RNA storage.
  7. The TOP-2/condensin II axis silences transcription during germline specification in C. elegans.
  8. Maintaining mitochondrial DNA copy number mitigates ROS-induced oocyte decline and female reproductive aging.
  9. Female-age-dependent changes in the lipid fingerprint of the mammalian oocytes.
  10. Zebrafish Foxl2l functions in proliferating germ cells for female meiotic entry.
  11. cTAGE5 is involved in the assembly of Golgi ring in mouse primordial follicle.
  12. The Drosophila maternal-effect gene abnormal oocyte ( ao ) does not repress histone gene expression.
  13. CHK1 controls zygote pronuclear envelope breakdown by regulating F-actin through interacting with MICAL3.
  1. Nat Cell Biol. 2024 Oct 01.
    An architectural role of specific RNA-RNA interactions in oskar granules.
    Mainak Bose, Branislava Rankovic, Julia Mahamid, Anne Ephrussi.
      Ribonucleoprotein (RNP) granules are membraneless condensates that organize the intracellular space by compartmentalization of specific RNAs and proteins. Studies have shown that RNA tunes the phase behaviour of RNA-binding proteins, but the role of intermolecular RNA-RNA interactions in RNP granules in vivo remains less explored. Here we determine the role of a sequence-specific RNA-RNA kissing-loop interaction in assembly of mesoscale oskar RNP granules in the female Drosophila germline. We show that a two-nucleotide mutation that disrupts kissing-loop-mediated oskar messenger RNA dimerization impairs condensate formation in vitro and oskar granule assembly in the developing oocyte, leading to defective posterior localization of the RNA and abrogation of oskar-associated processing bodies upon nutritional stress. This specific trans RNA-RNA interaction acts synergistically with the scaffold RNA-binding protein, Bruno, in driving condensate assembly. Our study highlights the architectural contribution of an mRNA and its specific secondary structure and tertiary interactions to the formation of an RNP granule that is essential for embryonic development.
    DOI:  https://doi.org/10.1038/s41556-024-01519-3
  2. Science. 2024 Oct 04. 386(6717): eadg7325
    Nr5a2 is dispensable for zygotic genome activation but essential for morula development.
    Nicola Festuccia, Sandrine Vandormael-Pournin, Almira Chervova, Anna Geiselman, Francina Langa-Vives, Rémi-Xavier Coux, Inma Gonzalez, Guillaume Giraud Collet, Michel Cohen-Tannoudji, Pablo Navarro.
      Early embryogenesis is driven by transcription factors (TFs) that first activate the zygotic genome and then specify the lineages constituting the blastocyst. Although the TFs specifying the blastocyst's lineages are well characterized, those playing earlier roles remain poorly defined. Using mouse models of the TF Nr5a2, we show that Nr5a2-/- embryos arrest at the early morula stage and exhibit altered lineage specification, frequent mitotic failure, and substantial chromosome segregation defects. Although NR5A2 plays a minor but measurable role during zygotic genome activation, it predominantly acts as a master regulator at the eight-cell stage, controlling expression of lineage-specifying TFs and genes involved in mitosis, telomere maintenance, and DNA repair. We conclude that NR5A2 coordinates proliferation, genome stability, and lineage specification to ensure correct morula development.
    DOI:  https://doi.org/10.1126/science.adg7325
  3. Nat Rev Mol Cell Biol. 2024 Oct 02.
    Rise and SINE: roles of transcription factors and retrotransposons in zygotic genome activation.
    Pavel Kravchenko, Kikuë Tachibana.
      In sexually reproducing organisms, life begins with the fusion of transcriptionally silent gametes, the oocyte and sperm. Although initiation of transcription in the embryo, known as zygotic genome activation (ZGA), is universally required for development, the transcription factors regulating this process are poorly conserved. In this Perspective, we discuss recent insights into the mechanisms of ZGA in totipotent mammalian embryos, namely ZGA regulation by several transcription factors, including by orphan nuclear receptors (OrphNRs) such as the pioneer transcription factor NR5A2, and by factors of the DUX, TPRX and OBOX families. We performed a meta-analysis and compiled a list of pan-ZGA genes, and found that most of these genes are indeed targets of the above transcription factors. Remarkably, more than a third of these ZGA genes appear to be regulated both by OrphNRs such as NR5A2 and by OBOX proteins, whose motifs co-occur in SINE B1 retrotransposable elements, which are enriched near ZGA genes. We propose that ZGA in mice is activated by recruitment of multiple transcription factors to SINE B1 elements that function as enhancers, and discuss a potential relevance of this mechanism to Alu retrotransposable elements in human ZGA.
    DOI:  https://doi.org/10.1038/s41580-024-00772-6
  4. Curr Biol. 2024 Sep 26. pii: S0960-9822(24)01215-6. [Epub ahead of print]
    Regulation of outer kinetochore assembly during meiosis I and II by CENP-A and KNL-2/M18BP1 in C. elegans oocytes.
    Laura Bellutti, Nicolas Macaisne, Layla El Mossadeq, Thadshagine Ganeswaran, Julie C Canman, Julien Dumont.
      During cell division, chromosomes build kinetochores that attach to spindle microtubules. Kinetochores usually form at the centromeres, which contain CENP-A nucleosomes. The outer kinetochore, which is the core attachment site for microtubules, is composed of the KMN network (Knl1c, Mis12c, and Ndc80c complexes) and is recruited downstream of CENP-A and its partner CENP-C. In C. elegans oocytes, kinetochores have been suggested to form independently of CENP-A nucleosomes. Yet kinetochore formation requires CENP-C, which acts in parallel to the nucleoporin MEL-28ELYS. Here, we used a combination of RNAi and Degron-based depletion of CENP-A (or downstream CENP-C) to demonstrate that both proteins are in fact responsible for a portion of outer kinetochore assembly during meiosis I and are essential for accurate chromosome segregation. The remaining part requires the coordinated action of KNL-2 (ortholog of human M18BP1) and of the nucleoporin MEL-28ELYS. Accordingly, co-depletion of CENP-A (or CENP-C) and KNL-2M18BP1 (or MEL-28ELYS) prevented outer kinetochore assembly in oocytes during meiosis I. We further found that KNL-2M18BP1 and MEL-28ELYS are interdependent for kinetochore localization. Using engineered mutants, we demonstrated that KNL-2M18BP1 recruits MEL-28ELYS at meiotic kinetochores through a specific N-terminal domain, independently of its canonical CENP-A loading factor activity. Finally, we found that meiosis II outer kinetochore assembly was solely dependent on the canonical CENP-A/CENP-C pathway. Thus, like in most cells, outer kinetochore assembly in C. elegans oocytes depends on centromeric chromatin. However, during meiosis I, an additional KNL-2M18BP1 and MEL-28ELYS pathway acts in a non-redundant manner and in parallel to canonical centromeric chromatin.
    Keywords:  CENP-A; CENP-C; centromere; chromosome segregation; holocentric; kinetochore; microtubule; oocyte meiosis
    DOI:  https://doi.org/10.1016/j.cub.2024.09.004
  5. Front Cell Dev Biol. 2024 ;12 1470981
    Aurora B and Aurora C pools at two chromosomal regions collaboratively maintain chromosome alignment and prevent aneuploidy at the second meiotic division in mammalian oocytes.
    Anna Kouznetsova, Sonata Valentiniene, Jian-Guo Liu, Tomoya S Kitajima, Hjalmar Brismar, Christer Höög.
      Correct chromosome segregation is essential to preserve genetic integrity. The two protein kinases, Aurora B and its meiotic homolog Aurora C, regulate attachments between chromosomal kinetochores and microtubules, thereby contributing to the accuracy of the chromosome segregation process. Here we performed a detailed examination of the localization and activity of Aurora B/C kinases, their partner Incenp and the kinetochore target Hec1, during the second meiotic division in mouse oocytes. We found that a majority of Aurora B and C changed their localization from the outer kinetochore region of chromosomes at prometaphase II to an inner central region localized between sister centromeres at metaphase II. Depletion of the Aurora B/C pool at the inner central region using the haspin kinase inhibitor 5-iodotubercidin resulted in chromosome misalignments at the metaphase II stage. To further understand the role of the Aurora B/C pool at the central region, we examined the behaviour of single chromatids, that lack a central Aurora B/C pool but retain Aurora B/C at the outer kinetochores. We found that kinetochore-microtubule attachments at single chromatids were corrected at both prometaphase II and metaphase II stages, but that single chromatids compared to paired chromatids were more prone to misalignments following treatment of oocytes with the Aurora B/C inhibitory drugs AZD1152 and GSK1070916. We conclude that the Aurora B/C pool at the inner central region stabilizes chromosome alignment during metaphase II arrest, while Aurora B/C localized at the kinetochore assist in re-establishing chromosome positioning at the metaphase plate if alignment is lost. Collaboratively these two pools prevent missegregation and aneuploidy at the second meiotic division in mammalian oocytes.
    Keywords:  Aurora B; Aurora C; aneuploidy; meiosis; oocyte; second meiotic division
    DOI:  https://doi.org/10.3389/fcell.2024.1470981
  6. Commun Biol. 2024 Oct 02. 7(1): 1247
    The mouse Balbiani body regulates primary oocyte quiescence via RNA storage.
    Lei Lei, Kanako Ikami, Edgar Andres Diaz Miranda, Sooah Ko, Faith Wilson, Haley Abbott, Ronald Pandoy, Shiying Jin.
      In mammalian females, the transition from dormancy in primordial follicles to follicular development is critical for maintaining ovarian function and reproductive longevity. In mice, the quiescent primary oocyte of the primordial follicle contains a Balbiani body (B-body), an organelle aggregate comprised of a spherical structure of Golgi complexes. Here we show that the structure of the B-body is maintained by microtubules and actin. The B-body stores mRNA-capping enzyme and 597 mRNAs associated with mRNA-decapping enzyme 1 A (DCP1A). Gene ontology analysis results indicate that proteins encoded by these mRNAs function in enzyme binding, cellular component organization and packing of telomere ends. Pharmacological depolymerization of microtubules or actin led to B-body disassociation and nascent protein synthesis around the dissociated B-bodies within three hours. An increased number of activated developing follicles were observed in ovaries with prolonged culture and the in vivo mouse model. Our results indicate that the mouse B-body is involved in the activation of dormant primordial follicles likely via translation of the B-body-associated RNAs in primary oocytes.
    DOI:  https://doi.org/10.1038/s42003-024-06900-4
  7. G3 (Bethesda). 2024 Oct 03. pii: jkae236. [Epub ahead of print]
    The TOP-2/condensin II axis silences transcription during germline specification in C. elegans.
    Mezmur D Belew, Emilie Chien, Matthew Wong, W Matthew Michael.
      In C. elegans, the germline is specified via a preformation mechanism that relies on the PIE-1 protein's ability to globally silence mRNA transcription in germline precursor cells, also known as the P lineage. Recent work from our group has identified additional genome silencing events in C. elegans during oogenesis and in starved L1 larvae, and these require the condensin II complex, topoisomerase II (TOP-2), and components of the H3K9me/heterochromatin pathway. Interestingly, silencing in oocytes also requires PIE-1, but this is not the case in starved L1s. Here, we ask if additional genome silencing components besides PIE-1 are required to repress gene expression in the P lineage of early embryos, and we find that condensin II and TOP-2 are required and the H3K9me/heterochromatin pathway is not. We show that depletion of TOP-2/condensin II activates the normally suppressed RNA polymerase II to inappropriately transcribe somatic genes in the P lineage. We also present evidence that while both PIE-1 and TOP-2/condensin II are required for genome silencing in the P lineage, PIE-1 can silence transcription independently of TOP-2/condensin II when misexpressed in somatic cells. Thus, in oocytes, all three genome silencing systems (TOP-2/condensin II, H3K9me, and PIE-1) are operational while in both early embryos and starved L1s two of the three are active. Our data show that multiple, redundantly acting genome silencing mechanisms act in a mix and match manner to repress transcription at different developmental stages in the C. elegans germline.
    Keywords:  C. elegans; PIE-1; condensin; genome silencing; germline; global transcriptional repression; topoisomerase II
    DOI:  https://doi.org/10.1093/g3journal/jkae236
  8. Commun Biol. 2024 Oct 01. 7(1): 1229
    Maintaining mitochondrial DNA copy number mitigates ROS-induced oocyte decline and female reproductive aging.
    Shiyun Long, Yunchao Zheng, Xiaoling Deng, Jing Guo, Zhe Xu, Karin Scharffetter-Kochanek, Yanmei Dou, Min Jiang.
      Oocytes play a crucial role in transmitting maternal mitochondrial DNA (mtDNA), essential for the continuation of species. However, the effects of mitochondrial reactive oxygen species (ROS) on mammalian oocyte maturation and mtDNA maintenance remain unclear. We investigated this by conditionally knocking out the Sod2 gene in primordial follicles, elevating mitochondrial matrix ROS levels from early oocyte stages. Our data indicates that reproductive aging in Sod2 conditional knockout females begins at 6 months, with oxidative stress impairing oocyte quality, particularly affecting OXPHOS complex II and mtDNA-encoded mRNA levels. Despite unchanged mtDNA mutation load, mtDNA copy numbers exhibited significant variations. Strikingly, reducing mtDNA copy numbers by reducing mtSSB protein, crucial for mtDNA replication, accelerated reproductive aging onset to three months, underscoring the critical role of mtDNA copy number maintenance under oxidative stress conditions. This research provides new insights into the relationship among mitochondrial ROS, mtDNA, and reproductive aging, offering potential strategies for delaying aging-related fertility decline.
    DOI:  https://doi.org/10.1038/s42003-024-06888-x
  9. Hum Reprod. 2024 Oct 04. pii: deae225. [Epub ahead of print]
    Female-age-dependent changes in the lipid fingerprint of the mammalian oocytes.
    Simona Bisogno, Joanna Depciuch, Hafsa Gulzar, Maria Florencia Heber, Michał Kobiałka, Łukasz Gąsior, Adrianna Bereta, Anna Pieczara, Kinga Fic, Richard Musson, Gabriel Garcia Gamero, Maria Pardo Martinez, Alba Fornés Pérez, Martina Tatíčková, Zuzana Holubcova, Małgorzata Barańska, Grażyna Ewa Ptak.
       STUDY QUESTION: Can oocyte functionality be assessed by observing changes in their intracytoplasmic lipid droplets (LDs) profiles?
    SUMMARY ANSWER: Lipid profile changes can reliably be detected in human oocytes; lipid changes are linked with maternal age and impaired developmental competence in a mouse model.
    WHAT IS KNOWN ALREADY: In all cellular components, lipid damage is the earliest manifestation of oxidative stress (OS), which leads to a cascade of negative consequences for organelles and DNA. Lipid damage is marked by the accumulation of LDs. We hypothesized that impaired oocyte functionality resulting from aging and associated OS could be assessed by changes in LDs profile, hereafter called lipid fingerprint (LF).
    STUDY DESIGN, SIZE, DURATION: To investigate if it is possible to detect differences in oocyte LF, we subjected human GV-stage oocytes to spectroscopic examinations. For this, a total of 48 oocytes derived from 26 young healthy women (under 33 years of age) with no history of infertility, enrolled in an oocyte donation program, were analyzed. Furthermore, 30 GV human oocytes from 12 women were analyzed by transmission electron microscopy (TEM). To evaluate the effect of oocytes' lipid profile changes on embryo development, a total of 52 C57BL/6 wild-type mice and 125 Gnpat+/- mice were also used.
    PARTICIPANTS/MATERIALS, SETTING, METHODS: Human oocytes were assessed by label-free cell imaging via coherent anti-Stokes Raman spectroscopy (CARS). Further confirmation of LF changes was conducted using spontaneous Raman followed by Fourier transform infrared (FTIR) spectroscopies and TEM. Additionally, to evaluate whether LF changes are associated with developmental competence, mouse oocytes and blastocysts were evaluated using TEM and the lipid dyes BODIPY and Nile Red. Mouse embryonic exosomes were evaluated using flow cytometry, FTIR and FT-Raman spectroscopies.
    MAIN RESULTS AND THE ROLE OF CHANCE: Here we demonstrated progressive changes in the LF of oocytes associated with the woman's age consisting of increased LDs size, area, and number. LF variations in oocytes were detectable also within individual donors. This finding makes LF assessment a promising tool to grade oocytes of the same patient, based on their quality. We next demonstrated age-associated changes in oocytes reflected by lipid peroxidation and composition changes; the accumulation of carotenoids; and alterations of structural properties of lipid bilayers. Finally, using a mouse model, we showed that LF changes in oocytes are negatively associated with the secretion of embryonic exosomes prior to implantation. Deficient exosome secretion disrupts communication between the embryo and the uterus and thus may explain recurrent implantation failures in advanced-age patients.
    LIMITATIONS, REASONS FOR CAUTION: Due to differences in lipid content between different species' oocytes, the developmental impact of lipid oxidation and consequent LF changes may differ across mammalian oocytes.
    WIDER IMPLICATIONS OF THE FINDINGS: Our findings open the possibility to develop an innovative tool for oocyte assessment and highlight likely functional connections between oocyte LDs and embryonic exosome secretion. By recognizing the role of oocyte LF in shaping the embryo's ability to implant, our original work points to future directions of research relevant to developmental biology and reproductive medicine.
    STUDY FUNDING/COMPETING INTEREST(S): This research was funded by National Science Centre of Poland, Grants: 2021/41/B/NZ3/03507 and 2019/35/B/NZ4/03547 (to G.E.P.); 2022/44/C/NZ4/00076 (to M.F.H.) and 2019/35/N/NZ3/03213 (to Ł.G.). M.F.H. is a National Agency for Academic Exchange (NAWA) fellow (GA ULM/2019/1/00097/U/00001). K.F. is a Diamond Grant fellow (Ministry of Education and Science GA 0175/DIA/2019/28). The open-access publication of this article was funded by the Priority Research Area BioS under the program "Excellence Initiative - Research University" at the Jagiellonian University in Krakow. The authors declare no competing interest.
    TRIAL REGISTRATION NUMBER: N/A.
    Keywords:   Gnpat ; carotenoids; coherent anti-Stokes Raman spectroscopy; exosomes; lipid analysis; lipid droplets; oocyte
    DOI:  https://doi.org/10.1093/humrep/deae225
  10. Dev Biol. 2024 Sep 26. pii: S0012-1606(24)00240-9. [Epub ahead of print]517 91-99
    Zebrafish Foxl2l functions in proliferating germ cells for female meiotic entry.
    Ching-Hsin Yang, Yan-Wei Wang, Chen-Wei Hsu, Bon-Chu Chung.
      Zebrafish sex differentiation is a complicated process and the detailed mechanism has not been fully understood. Here we characterized a transcription factor, Foxl2l, which participates in female oogenesis. We show that it is expressed specifically in proliferating germ cells in juvenile gonads and mature ovaries. We have used CRISPR-Cas9 to generate zebrafish deficient in foxl2l expression. Zebrafish with foxl2l-/- are all males, and this female-to-male sex reversal cannot be reversed by tp53 mutation, indicating this sex reversal is unrelated to cell death. We have generated transgenic fish expressing GFP under the control of foxl2l promoter to track the development of foxl2l + -germ cells; these cells failed to enter meiosis and accumulated as cystic cells in the foxl2l-/- mutant. Our RNA-seq analysis also showed the reduced expression of genes in meiosis and oogenesis among other affected pathways. All together, we show that zebrafish Foxl2l is a nuclear factor controlling the expression of meiotic and oogenic genes, and its deficiency leads to defective meiotic entry and the accumulation of premeiotic germ cells.
    Keywords:  Foxl2l; Meiosis; Oogenesis; RNA-Seq; Sex differentiation; Transcriptome
    DOI:  https://doi.org/10.1016/j.ydbio.2024.09.013
  11. Sci Bull (Beijing). 2024 Sep 19. pii: S2095-9273(24)00652-2. [Epub ahead of print]
    cTAGE5 is involved in the assembly of Golgi ring in mouse primordial follicle.
    Haowei Wu, Yaqing Wang, Jiayi Li, Yan Qin, Min Chen, Zhiming Shen, Fangfang Dong, Xiuhong Cui, Liang Liu, Zhiheng Xu, Fei Gao.
      
    DOI:  https://doi.org/10.1016/j.scib.2024.09.018
  12. bioRxiv. 2024 Sep 18. pii: 2024.09.17.613536. [Epub ahead of print]
    The Drosophila maternal-effect gene abnormal oocyte ( ao ) does not repress histone gene expression.
    Risa Takenaka, Sierra M Simmerman, Casey A Schmidt, Eric H Albanese, Leila E Rieder, Harmit S Malik.
      The abnormal oocyte ( ao ) gene of Drosophila melanogaster is a maternal-effect lethal gene previously identified as encoding a transcriptional regulator of core histones. However, background genetic mutations in existing ao mutant strains could compromise their utility in manipulating histone levels. To distinguish the true ao phenotype from background effects, we created two new ao reagents: a CRISPR/Cas9-mediated knockout of the ao allele for genetic and molecular analyses and an epitope-tagged ao allele for cytological experiments. Using these reagents, we confirm previous findings that ao exhibits maternal-effect lethality, which can be rescued by either a decrease in the histone gene copy number or by Y chromosome heterochromatin. We also confirm that the Ao protein localizes to the histone locus bodies in ovaries. Our data also suggest that ao genetically interacts with the histone genes and heterochromatin, as previously suggested. However, contrary to prior findings, we find that ao does not repress core histone transcript levels. Thus, the molecular basis for ao -associated maternal-effect lethality remains unknown.
    DOI:  https://doi.org/10.1101/2024.09.17.613536
  13. EMBO Rep. 2024 Oct 02.
    CHK1 controls zygote pronuclear envelope breakdown by regulating F-actin through interacting with MICAL3.
    Honghui Zhang, Ying Cui, Bohan Yang, Zhenzhen Hou, Mengge Zhang, Wei Su, Tailai Chen, Yuehong Bian, Mei Li, Zi-Jiang Chen, Han Zhao, Shigang Zhao, Keliang Wu.
      CHK1 mutations could cause human zygote arrest at the pronuclei stage, a phenomenon that is not well understood at the molecular level. In this study, we conducted experiments where pre-pronuclei from zygotes with CHK1 mutation were transferred into the cytoplasm of normal enucleated fertilized eggs. This approach rescued the zygote arrest caused by the mutation, resulting in the production of a high-quality blastocyst. This suggests that CHK1 dysfunction primarily disrupts crucial biological processes occurring in the cytoplasm. Further investigation reveals that CHK1 mutants have an impact on the F-actin meshwork, leading to disturbances in pronuclear envelope breakdown. Through co-immunoprecipitation and mass spectrometry analysis of around 6000 mouse zygotes, we identified an interaction between CHK1 and MICAL3, a key regulator of F-actin disassembly. The gain-of-function mutants of CHK1 enhance their interaction with MICAL3 and increase MICAL3 enzymatic activity, resulting in excessive depolymerization of F-actin. These findings shed light on the regulatory mechanism behind pronuclear envelope breakdown during the transition from meiosis to the first mitosis in mammals.
    Keywords:  CHK1; F-actin; MICAL3; Pronuclear Envelope Breakdown; Zygote
    DOI:  https://doi.org/10.1038/s44319-024-00267-7
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