bims-cebooc 2025-10-12 papers

Issue of 2025–10–12
twelve papers selected by
Gabriele Zaffagnini, Universität zu Köln

Science. 2025 Oct 09.

Comparative analysis of human and mouse ovaries across age.

Eliza A Gaylord, Mariko H Foecke, Ryan M Samuel, Bikem Soygur, Angela M Detweiler, Tara I McIntyre, Leah C Dorman, Michael Borja, Amy E Laird, Ritwicq Arjyal, Juan Du, James M Gardner, Norma Neff, Faranak Fattahi, Diana J Laird.

The mouse is a tractable model for human ovarian biology, however its utility is limited by incomplete understanding of how transcription and signaling differ interspecifically and with age. We compared ovaries between species using three-dimensional imaging, single-cell transcriptomics, and functional studies. In mice, we mapped declining follicle numbers and oocyte competence during aging; in human ovaries, we identified cortical follicle pockets and decreases in density. Oocytes had species-specific gene expression patterns during growth that converged toward maturity. Age-related transcriptional changes were greater in oocytes than granulosa cells across species, although mature oocytes change more in humans. We identified ovarian sympathetic nerves and glia; axon density increased in aged ovaries and, when ablated in mice, perturbed folliculogenesis. This comparative atlas defines shared and species-specific hallmarks of ovarian biology.

DOI: https://doi.org/10.1126/science.adx0659

Science. 2025 Oct 09. 390(6769): 156-163

Ubiquitin-mediated mitophagy regulates the inheritance of mitochondrial DNA mutations.

Michele Frison, Brandon S Lockey, Yu Nie, Zoe Golder, Eleni Theiaspra, Cameron D Ryall, Camilla Lyons, Stephen P Burr, Malwina Prater, Lyuba V Bozhilova, Angelos Glynos, James B Stewart, Nick S Jones, Marcos R Chiaratti, Patrick F Chinnery.

Mitochondrial synthesis of adenosine triphosphate is essential for eukaryotic life but is dependent on the cooperation of two genomes: nuclear and mitochondrial DNA (mtDNA). mtDNA mutates ~15 times as fast as the nuclear genome, challenging this symbiotic relationship. Mechanisms must have evolved to moderate the impact of mtDNA mutagenesis but are poorly understood. Here, we observed purifying selection of a mouse mtDNA mutation modulated by Ubiquitin-specific peptidase 30 (Usp30) during the maternal-zygotic transition. In vitro, Usp30 inhibition recapitulated these findings by increasing ubiquitin-mediated mitochondrial autophagy (mitophagy). We also found that high mutant burden, or heteroplasmy, impairs the ubiquitin-proteasome system, explaining how mutations can evade quality control to cause disease. Inhibiting USP30 unleashes latent mitophagy, reducing mutant mtDNA in high-heteroplasmy cells. These findings suggest a potential strategy to prevent mitochondrial disorders.

DOI: https://doi.org/10.1126/science.adr5438

Elife. 2025 Oct 07. pii: RP107352. [Epub ahead of print]14

Distinct waves of ovarian follicles contribute to mouse oocyte production.

Qi Yin, Allan C Spradling.

The earliest growing mouse follicles, wave 1, rapidly develop in the ovarian medulla, while the great majority, wave 2, are stored for later use as resting primordial follicles in the cortex. Wave 1 follicles are known to mostly undergo atresia, a fate sometimes associated with the persistence of steroidogenic theca cells, but this connection is poorly understood. We characterized wave 1 follicle biology using tissue clearing, lineage tracing, and scRNA-seq to clarify their contributions to offspring and steroidogenesis. Wave 1 follicles, lineage-marked by E16.5 Foxl2 expression in granulosa cells, reach preantral stages containing theca cell layers by 2 weeks. Atresia begins about a week later, during which 80-100% of wave 1 follicles degrade their oocytes, turn over most granulosa cells, but retain theca cells which expand in number together with interstitial gland cells in the medulla. During puberty (5 weeks), these cells ultrastructurally resemble steroidogenic cells and highly express androgen biosynthetic genes. Unexpectedly, the Foxl2 lineage tag also marked about 400 primordial follicles, located near the medullary-cortical boundary, that become the earliest activated wave 2 follicles. These 'boundary' or 'wave 1.5' follicles generate 70-100% of the earliest mature oocytes, while fewer than 26 wave 1 follicles with oocytes survive. Consistent with their largely distinct fates in steroid or oocyte production, granulosa cells of antral wave 1 and 2 follicles differentially express multiple genes, including Wnt4 and Igfbp5.

Keywords: developmental biology; follicle wave; mouse; oocyte; ovarian follicle; steroid; theca cell

DOI: https://doi.org/10.7554/eLife.107352

J Cell Sci. 2025 Oct 06. pii: jcs.264161. [Epub ahead of print]

Identification of locally activated spindle-associated proteins in oocytes uncovers a phosphatase-driven mechanism.

Xiang Wan, Gera Pavlova, C Fiona Cullen, Igor Dasuzhau, Aleksandra Ciszek, Hiroyuki Ohkura.

The meiotic spindle forms only around the chromosomes in oocytes, despite the exceptionally large volume of the cytoplasm. This spatial restriction is likely to be governed by local activation of key microtubule regulators around the chromosomes in oocytes, but the identities of these microtubule regulators and the mechanisms remain unclear. To address this, we developed a novel assay to visualise spatial regulation of spindle-associated proteins in Drosophila oocytes by inducing ectopic microtubule clusters. This assay identified several proteins including the TPX2 homologue Mei-38 that localise more strongly to microtubules near the chromosomes than away from them. In Mei-38, we identified a microtubule-binding domain containing a region highly conserved also in humans. The domain itself is regulated spatially, and contains a conserved serine and a nearby PP2A-B56 docking motif. A non-phosphorylatable mutation of this serine allows the domain to localise to ectopic microtubules as well as spindle microtubules, while mutations in a PP2A-B56 docking motif greatly reduced the spindle localisation. As this phosphatase is concentrated at the kinetochores, it may act as a novel chromosomal signal spatially regulating spindle proteins within oocytes.

Keywords: Drosophila; Meiosis; Oocyte; Phosphatase; Spindle

DOI: https://doi.org/10.1242/jcs.264161

Nat Commun. 2025 Oct 07. 16(1): 8917

Ecdysone signaling-induced dumpless1 expression controls nurse cell dumping in Drosophila oogenesis.

Jingshan Li, Zishan Pan, Xiaoqi Peng, Yu Feng, Junfeng Wu, Feiying Liang, Qili Feng, XiaoQiang Yu, Huimin Deng.

Nurse cell (NC) dumping, a process essential for oocyte development, involves the rapid cytoplasmic transfer from germline-derived NCs into the oocyte. However, its regulatory mechanism remains unclear. Here, we report that ecdysone signaling in stretch follicle cells (SFCs) regulates NC dumping through dumpless1, a ZAD-C2H2 zinc finger transcription factor, in Drosophila. Ecdysone induced dumpless1 expression in SFCs, and CRISPR/Cas9-mediated knockout of dumpless1 or its functional domain ZAD suppresses NC dumping. Depletion of dumpless1 upregulates integrin βPS expression in SFC plasma membrane, while reducing cortical enrichment of Rho1 signaling-dependent phosphorylated myosin light chain (p-MLC) and disrupting actin cables organization in NCs. SFC-specific overexpression of integrin βPS reduces p-MLC enrichment in the NC cortex, whereas its knockdown in SFCs of dumpless1-/- mutants partially rescues NC dumping defect. Our findings identify dumpless1 as a critical effector of ecdysone signaling, bridging somatic-germline communication through the integrin βPS-Rho1-p-MLC axis, revealing a multicellular regulatory mechanism in Drosophila oogenesis.

DOI: https://doi.org/10.1038/s41467-025-63973-3

Anim Reprod. 2025 ;22(3): e20250090

Approaches to in vitro oocyte growth in domestic farm mammals: how and why?

Valentina Lodde, Noemi Monferini, Maria Plevridi, Pritha Dey, Ludovica Donadini, Fernanda Fagali Franchi, Federica Franciosi, Alberto Maria Luciano.

Unlocking the developmental potential of oocytes at various stages of folliculogenesis represents a major challenge in reproductive biology and assisted reproductive technologies. While in vitro maturation (IVM) of fully grown oocytes is widely applied, the vast majority of oocytes enclosed within early-stage follicles remain underutilized. This review outlines current advancements in in vitro culture systems designed to support oocyte growth and differentiation, with particular attention to the contributions of the authors. Key developments, mainly encompassing the bovine species, include the use of prematuration strategies to enhance the competence of oocytes retrieved from antral follicles, stepwise in vitro culture protocols for growing oocytes from early antral follicles, and efforts to establish defined systems for preantral follicle culture. Emerging insights into chromatin dynamics, cumulus-oocyte communication, and epigenetic regulation are shaping the design of tailored culture environments. Despite promising progress, significant challenges remain in replicating the complexity of in vivo folliculogenesis, particularly in non-rodent models. Addressing these challenges will be critical to expanding the oocyte pool available for reproductive and biotechnological applications, with broad implications for fertility preservation, livestock breeding, and fundamental research.

Keywords: early antral follicles; folliculogenesis; in vitro folliculogenesis; oocyte culture; ovarian reserve; preantral follicles; prematuration (pre-IVM)

DOI: https://doi.org/10.1590/1984-3143-AR2025-0090

MicroPubl Biol. 2025 ;2025

Exclusion of the endoplasmic reticulum from the C. elegans meiotic spindle controls spindle size.

Alma Aquino, Francis McNally.

The nuclear envelope is composed of sheet-like ER that separates chromatin from tubulin during interphase. During oocyte maturation in C. elegans , a fenestrated envelope of sheet-like ER continues to envelope the meiotic spindle through metaphase I. ER is thus excluded from the nuclear/spindle volume during spindle assembly. To test the importance of this exclusion, we forced ER into the meiotic spindle by coupling kinesin motor domains to the ER. Forcing ER into the spindle interior caused a statistically significant increase in metaphase spindle width. Exclusion of ER from the spindle thus affects spindle geometry.

DOI: https://doi.org/10.17912/micropub.biology.001812

Nat Commun. 2025 Oct 08. 16(1): 8955

Fosl2 facilitates chromatin accessibility to determine developmental events during follicular maturation.

Hongyong Zhang, Zechen Li, Yanmei Zhu, Wencong Lyu, Wenlu Wei, Haochen Wang, Shuangjie Tian, Wei Yue, Jiajing Zhong, Qing-Yuan Sun, Yiting Guan.

Granulosa cells (GCs) are the most dynamically responsive cell lineage to encourage continuous folliculogenesis; however, developmental dynamics and interplay with downstream transcription circuitry remain unclear. Here, we unravel the redistribution of genome-wide chromatin areas that drive broad developmental-related transcriptomic alterations during follicular maturation in murine and porcine GCs. Distinct GC-activated accessibility regions (GAAs) at the ovulatory phase are responsible for augmenting flanking GC-involved developmental gene (GDG) expression, which are essential for transcriptional responses to developmental cues. Mechanistically, the transcription factor Fosl2 is strongly recruited to GAAs, facilitating chromatin accessibility state transition. Elevated GAA signals driven by Fosl2 loading induce a significant upregulation of adjacent GDG expression. Additionally, GC-specific Fosl2 deletion in mice perturbs GC cellularity, leading to subfertility related to reproductive aging. Together, we highlight a dynamic chromatin accessibility landscape during follicular maturation, revealing the indispensable Fosl2 function not only controls transcriptional activation via a reconfigured chromatin state, but also orchestrates intricate signaling pathways that are fundamental for ovulation and reproduction.

DOI: https://doi.org/10.1038/s41467-025-64009-6

J Vis Exp. 2025 Sep 19.

Optimized Analysis of Proteins from Xenopus Oocytes and Embryos by Immunoblotting.

Charlotte R Kanzler, Michael D Sheets.

Analysis of proteins by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) followed by immunoblotting (western blotting) is a vital part of the molecular biologist's toolkit. This technique separates a complex protein mixture by molecular weight and then assays the presence of target proteins using specific antibodies. Immunoblotting has a variety of applications. Examples include use as a targeted approach to study protein-protein interactors or as a control to confirm expression or depletion of protein targets. However, the successful execution of immunoblotting requires complicated, multistep experiments. Protocols must be optimized for each organism, target protein, and application. Therefore, knowledge gaps exist for the use of immunoblots in many models, including the model frog Xenopus laevis. Due to their large size, abundant material for biochemical experiments, and facile handling, X. laevis oocytes and embryos have been vital for studying principles of translational control. However, this species lacks specific protocols for robust and routine immunoblotting. Here, we offer an in-depth protocol for western blotting optimized for samples from multiple Xenopus developmental stages. We then analyze translational regulators across development.

DOI: https://doi.org/10.3791/69139

Reprod Biomed Online. 2025 Jun 18. pii: S1472-6483(25)00315-3. [Epub ahead of print]51(5): 105108

Initial and cyclic recruitment of ovarian follicles: a quarter-century update.

Sezcan Mumusoglu, Qingling Yang, Kazuhiro Kawamura, Chialin Chang M, Kui Liu, Aaron J Hsueh.

A review of advancements in primordial follicle activation and cyclic recruitment research over the past 25 years is presented. The review examines the latest insights into the mechanisms of primordial follicle activation and the role of the ovarian Hippo signalling pathways, highlighting their potential for developing new infertility treatments. The concept of continuous waves of early antral follicles and their availability for cyclic recruitment is discussed. Several previously unresolved questions, such as the effect of steroid contraceptive pill usage on menopause timing, the potential risk for earlier menopause in mothers of dizygotic twins and the effects of repeated ovarian stimulation cycles on menopause onset are addressed. Controversies about the existence of female germ stem cells, the lifespan of follicles, the phenomenon of alternate ovulation between ovaries, potential changes in menopause onset in patients with polycystic ovary syndrome (PCOS) and endometriosis are discussed. In conclusion, future challenges in ovarian research, aiming to advance understanding and improve clinical practices in reproductive health are addressed.

Keywords: cyclic recruitment; folliculogenesis; hippo signalling; initial primordial follicle activation; ovarian stimulation

DOI: https://doi.org/10.1016/j.rbmo.2025.105108

STAR Protoc. 2025 Oct 07. pii: S2666-1667(25)00538-6. [Epub ahead of print]6(4): 104132

A protocol for real-time tracking of transposon products and host proteins in Drosophila ovaries.

Yaqian Xu, Kun Dou.

Drosophila ovary is an excellent model for studying material transportation and dynamics. Here, we present a protocol for live imaging of transposon products and host proteins in Drosophila ovaries. We describe steps from ovary dissection, ovariole isolation, and mounting to live tracking of fluorescently labeled proteins with single-cell resolution. Furthermore, we detail procedures of image processing for jitter correction and kymograph analysis. This protocol can be applied to studies on the dynamics of other fluorescently tagged proteins in Drosophila ovaries. For complete details on the use and execution of this protocol, please refer to Shen et al.1.

Keywords: Genetics; Microscopy; cell Biology; developmental biology; model Organisms

DOI: https://doi.org/10.1016/j.xpro.2025.104132

Hum Reprod. 2025 Oct 07. pii: deaf194. [Epub ahead of print]

In vitro system completely restores oogenesis in congenitally infertile mice.

K Yoshida, S Kimura, M Taguchi, H Morimoto, M Kanatsu-Shinohara, T Shinohara, Y Obata.

STUDY QUESTION: Can in vitro systems, combined with transient gene expression or factor supplementation, completely restore fertility in congenitally infertile mice?
SUMMARY ANSWER: Transient expression of Kitl via adeno-associated virus (AAV) vectors or supplementation with recombinant KITL in KitlSl-t/KitlSl-t mice-a model of congenital infertility caused by a mutation in the Kitl locus-resulted in the production of mature oocytes and the birth of healthy, fertile offspring.
WHAT IS KNOWN ALREADY: Although in vivo gene delivery has enabled offspring production in infertile mouse models, low efficiency, unpredictability of parturition timing, inflammatory risk, possible viral genome integration, and lack of real-time oogenesis observation remain major concerns. Despite the potential of in vitro oogenesis as an alternative, complete functional restoration of gene deficiency has not been reported.
STUDY DESIGN, SIZE, DURATION: AAV-mCherry was applied to wild-type mouse ovaries, and expression levels were compared across 15 serotypes (2.5 × 1011 viral genomes/ml; N = 4-12; 4-day infection, 20-day culture) to identify optimal AAV serotypes for ovarian gene delivery. The effects of AAV-Kitl infection (six doses; N = 3-5) and recombinant KITL supplementation (four doses; N = 5) on oocyte growth were evaluated in KitlSl-t/KitlSl-t mouse ovaries. On culture day 17 or 18, secondary follicles were isolated and cultured for an additional 16 days to evaluate oocyte competence for maturation, fertilization, and full-term development. Offspring were delivered 52-53 days after treatment initiation.
PARTICIPANTS/MATERIALS, SETTING, METHODS: Ovaries from KitlSl-t/KitlSl-t mice were dissociated into single cells and reaggregated in U-bottom wells with media containing AAV8-Kitl, AAV9-Kitl, or recombinant KITL. Reconstituted ovaries were cultured on insert membranes, thereby allowing primordial follicles to develop into secondary follicles. Isolated secondary follicles were further cultured to the antral stage, and cumulus-oocyte complexes were subjected to IVM and IVF. The resulting embryos were transferred to foster mothers. Finally, the offspring were subjected to PCR screening for AAV sequences and fertility tests.
MAIN RESULTS AND THE ROLE OF CHANCE: AAV8, AAV9, AAVrh.10, and AAVrh.32.33 induced significantly higher levels of mCherry expression in wild-type mouse ovaries than 10 of the 15 AAV evaluated serotypes in vitro (P < 0.05). AAV8-Kitl promoted primordial follicle activation in a dose-dependent manner in KitlSl-t/KitlSl-t mouse ovaries, with the highest number of secondary follicles (80 per reconstituted ovary) obtained at 1.0 × 1011 vg/ml (P < 0.05). In contrast, AAV9-Kitl required 2.5- to 10-fold higher titers to achieve comparable levels of secondary follicle formation. Contrastingly, no secondary follicles were formed in KitlSl-t/KitlSl-t mouse ovaries following mock treatment. Furthermore, supplementation with 200 ng/ml recombinant KITL supported secondary follicle formation at levels comparable to those in the wild-type mouse ovaries. More than 10% of fertilized oocytes developed to full term, regardless of the treatment method. AAV DNA was not detected in the genomes of the 47 offspring, and all tested female mice exhibited normal fertility.
LARGE SCALE DATA: N/A.
LIMITATIONS, REASONS FOR CAUTION: To date, complete in vitro oogenesis has only been achieved in mice; its applicability to other species, including humans, remains unverified.
WIDER IMPLICATIONS OF THE FINDINGS: This study establishes a novel and controllable in vitro platform to compensate for gene function through transient gene expression or factor supplementation, without permanent genomic modification. This approach provides a powerful framework for the dissection of gene functions during oogenesis, modeling of reproductive disorders, and development of fertility restoration strategies in both clinical and conservation contexts.
STUDY FUNDING/COMPETING INTEREST(S): This study was supported by KAKENHI (grant numbers 18H05547, 23K27088, and 25H01353). The authors declare no competing interests.

Keywords: in vitro oogenesis; AAV; KIT ligand; c-Kit; infertility treatment; oocyte growth

DOI: https://doi.org/10.1093/humrep/deaf194