Decoding the Molecular Landscape of Prepubertal Oocyte Maturation: GTPBP4 as a Key Driver of In Vitro Developmental Competence

Abstract

The intricate mechanisms driving oocyte maturation remain only partially understood, especially within the domains of domestic animal reproduction and translational medicine. In the case of prepubertal girls, the clinical challenge is especially pronounced, as ovarian tissue cryopreservation-though promising-remains an experimental technique necessitating rigorous scientific validation to guarantee the developmental potential of preserved materials and facilitate broader clinical adoption. To address these knowledge gaps, while considering the ethical implications, we applied transcriptome and translatome sequencing to comprehensively profile the transcriptional and translational dynamics of oocyte maturation in adult and prepubertal goats. Our analyses uncovered a sequential transition in gene expression regulation, shifting from cytoplasmic processes to chromosome segregation during the maturation process. Comparative profiling between adult and prepubertal goat oocytes revealed critical regulatory factors essential for prepubertal oocyte maturation. These include genes involved in organelle function (GTPBP4 and TOMM7), spindle organisation (CKS2, CCP110, CKAP5 and ESCO1) and chromosome segregation (CENPE, CENPF, CENPN and SGO2). Functional validation through in vitro maturation experiments demonstrated that GTPBP4 significantly enhances the developmental competence of prepubertal goat oocytes. This enhancement occurs through mechanisms that promote cell cycle progression, organelle maturation and mRNA translation. These findings provide a detailed map of the molecular events underpinning goat oocyte maturation and offer new perspectives on the developmental strategies required for oocyte competence in prepubertal females. Translating these insights to humans, this research highlights potential fertility preservation strategies for prepubertal girls, such as ovarian tissue cryopreservation and transplantation, in vitro follicle culture, meiotic maturation and artificial ovary technologies. Moreover, the identified mechanisms have significant implications for improving reproductive efficiency in domestic animal breeding, bridging basic research and applied science.

Keywords

adult goat / GTPBP4 / oocyte maturation / prepubertal goat / transcriptome / translatome

References

[1] D. Keefe, M. Kumar, and K. Kalmbach, “Oocyte Competency Is the Key to Embryo Potential,” Fertility and Sterility 103, no. 2 (2015): 317–322, https://doi.org/10.1016/j.fertnstert.2014.12.115.

[2] Y. Yuan, L. D. Spate, B. K. Redel, et al., “Quadrupling Efficiency in Production of Genetically Modified Pigs Through Improved Oocyte Maturation,” Proceedings of the National Academy of Sciences of the United States of America 114, no. 29 (2017): E5796–E5804, https://doi.org/10.1073/pnas.1703998114.

[3] M. Zhang, J. Zhang, D. Wang, et al., “C-X-C Motif Chemokine Ligand 12 Improves the Developmental Potential of Bovine Oocytes by Activating SH2 Domain-Containing Tyrosine Phosphatase 2 During Maturation,” Biology of Reproduction 109, no. 3 (2023): 282–298, https://doi.org/10.1093/biolre/ioad079.

[4] Y. Luo, J. Li, L. Zheng, et al., “Procyanidin B2 Improves Developmental Capacity of Bovine Oocytes via Promoting PPARγ/UCP1-Mediated Uncoupling Lipid Catabolism During In Vitro Maturation,” Cell Proliferation 57, no. 11 (2024): e13687, https://doi.org/10.1111/cpr.13687.

[5] B. Pan, J. Qin, K. Du, et al., “Integrated Ultrasensitive Metabolomics and Single-Cell Transcriptomics Identify Crucial Regulators of Sheep Oocyte Maturation and Early Embryo Development In Vitro,” Journal of Advanced Research S2090-1232, no. 24 (2024): 381–383, https://doi.org/10.1016/j.jare.2024.08.040.

[6] C. C. L. Fernandes, L. H. Aguiar, C. E. M. Calderón, et al., “Nutritional Impact on Gene Expression and Competence of Oocytes Used to Support Embryo Development and Livebirth by Cloning Procedures in Goats,” Animal Reproduction Science 188 (2018): 1–12, https://doi.org/10.1016/j.anireprosci.2017.10.012.

[7] J. M. Kelly, D. O. Kleemann, and S. K. Walker, “Enhanced Efficiency in the Production of Offspring From 4- to 8-Week-Old Lambs,” Theriogenology 63, no. 7 (2005): 1876–1890, https://doi.org/10.1016/j.theriogenology.2004.09.010.

[8] K. M. Gou, H. Guan, J. H. Bai, et al., “Field Evaluation of Juvenile In Vitro Embryo Transfer (JIVET) in Sheep,” Animal Reproduction Science 112, no. 3-4 (2009): 316–324, https://doi.org/10.1016/j.anireprosci.2008.05.008.

[9] J. R. Gruhn, A. P. Zielinska, V. Shukla, et al., “Chromosome Errors in Human Eggs Shape Natural Fertility Over Reproductive Life Span,” Science 365, no. 6460 (2019): 1466–1469, https://doi.org/10.1126/science.aav7321.

[10] P. Patrizio and D. Albertini, “Old Is Bad, Young Is Good, but What About Very Young? Oocytes Obtained From Pre-Pubertal Ovarian Tissue of Very Young Patients Are Incapable of In Vitro Maturation,” Fertility and Sterility 112, no. 2 (2019): 239–240, https://doi.org/10.1016/j.fertnstert.2019.05.002.

[11] M. G. Palmerini, S. A. Nottola, G. G. Leoni, et al., “In Vitro Maturation Is Slowed in Prepubertal Lamb Oocytes: Ultrastructural Evidences,” Reproductive Biology and Endocrinology 12 (2014): 115, https://doi.org/10.1186/1477-7827-12-115.

[12] S. Silber, “Ovarian Tissue Cryopreservation and Transplantation: Scientific Implications,” Journal of Assisted Reproduction and Genetics 33, no. 12 (2016): 1595–1603, https://doi.org/10.1007/s10815-016-0814-1.

[13] S. Wang, Y. Jiang, R. Zheng, Y. Yu, W. Wei, and N. Li, “Mobilizing China and the Global Community to Confront the Treatment Desert for Pediatric Solid Tumors,” Cancer Discovery 14, no. 1 (2024): 26–29, https://doi.org/10.1158/2159-8290.Cd-23-0451.

[14] Y. Guo, L. Xue, W. Tang, et al., “Ovarian Microenvironment: Challenges and Opportunities in Protecting Against Chemotherapy-Associated Ovarian Damage,” Human Reproduction Update 30, no. 5 (2024): 614–647, https://doi.org/10.1093/humupd/dmae020.

[15] E. L. Tsui, H. B. Mcdowell, and M. M. Laronda, “Restoring Ovarian Fertility and Hormone Function: Recent Advancements, Ongoing Efforts and Future Applications,” Journal of the Endocrine Society 8, no. 6 (2024): bvae073, https://doi.org/10.1210/jendso/bvae073.

[16] L. A. Jaffe and J. R. Egbert, “Regulation of Mammalian Oocyte Meiosis by Intercellular Communication Within the Ovarian Follicle,” Annual Review of Physiology 79 (2017): 237–260, https://doi.org/10.1146/annurev-physiol-022516-034102.

[17] Z. Pei, K. Deng, C. Xu, and S. Zhang, “The Molecular Regulatory Mechanisms of Meiotic Arrest and Resumption in Oocyte Development and Maturation,” Reproductive Biology and Endocrinology 21, no. 1 (2023): 90, https://doi.org/10.1186/s12958-023-01143-0.

[18] R. Pasquariello, L. Bogliolo, F. Di Filippo, et al., “Use of Assisted Reproductive Technologies (ARTs) to Shorten the Generational Interval in Ruminants: Current Status and Perspectives,” Theriogenology 225 (2024): 16–32, https://doi.org/10.1016/j.theriogenology.2024.05.026.

[19] G. Karavani, N. Schachter-Safrai, A. Revel, T. Mordechai-Daniel, D. Bauman, and T. Imbar, “In Vitro Maturation Rates in Young Premenarche Patients,” Fertility and Sterility 112, no. 2 (2019): 315–322, https://doi.org/10.1016/j.fertnstert.2019.03.026.

[20] P. Ntostis, D. Iles, G. Kokkali, et al., “The Impact of Maternal Age on Gene Expression During the GV to MII Transition in Euploid Human Oocytes,” Human Reproduction 37, no. 1 (2021): 80–92, https://doi.org/10.1093/humrep/deab226.

[21] X. Yuan, N. Chen, Y. Feng, et al., “Single-Cell Multi-Omics Profiling Reveals Key Regulatory Mechanisms That Poise Germinal Vesicle Oocytes for Maturation in Pigs,” Cellular and Molecular Life Sciences 80, no. 8 (2023): 222, https://doi.org/10.1007/s00018-023-04873-x.

[22] Z. Liu, G. Zhang, M. Deng, et al., “Inhibition of Lysine-Specific Histone Demethylase 1A Results in Meiotic Aberration During Oocyte Maturation In Vitro in Goats,” Theriogenology 143 (2020): 168–178, https://doi.org/10.1016/j.theriogenology.2019.12.011.

[23] K. Becker, A. Bluhm, N. Casas-Vila, et al., “Quantifying Post-Transcriptional Regulation in the Development of Drosophila Melanogaster,” Nature Communications 9, no. 1 (2018): 4970, https://doi.org/10.1038/s41467-018-07455-9.

[24] W. Hu, H. Zeng, Y. Shi, et al., “Single-Cell Transcriptome and Translatome Dual-Omics Reveals Potential Mechanisms of Human Oocyte Maturation,” Nature Communications 13, no. 1 (2022): 5114, https://doi.org/10.1038/s41467-022-32791-2.

[25] J. Huang, P. Chen, L. Jia, et al., “Multi-Omics Analysis Reveals Translational Landscapes and Regulations in Mouse and Human Oocyte Aging,” Advanced Science 10, no. 26 (2023): e2301538, https://doi.org/10.1002/advs.202301538.

[26] Y. Yuan, R. Liu, X. Zhang, et al., “Effects of Recipient Oocyte Source, Number of Transferred Embryos and Season on Somatic Cell Nuclear Transfer Efficiency in Sheep,” Reproduction in Domestic Animals 54, no. 11 (2019): 1443–1448, https://doi.org/10.1111/rda.13546.

[27] K. Liu, L. Zhang, Q. Qi, J. Li, F. Yan, and J. Hou, “Growth Hormone Treatment Improves the Development of Follicles and Oocytes in Prepubertal Lambs,” Journal of Ovarian Research 16, no. 1 (2023): 132, https://doi.org/10.1186/s13048-023-01209-y.

[28] Y. Li, J. Sun, Y. Ling, et al., “Transcription Profiles of Oocytes During Maturation and Embryos During Preimplantation Development In Vivo in the Goat,” Reproduction, Fertility, and Development 32, no. 7 (2020): 714–725, https://doi.org/10.1071/rd19391.

[29] H. Mu, T. Zhang, Y. Yang, et al., “METTL3-Mediated MRNA N(6)-Methyladenosine Is Required for Oocyte and Follicle Development in Mice,” Cell Death & Disease 12, no. 11 (2021): 989, https://doi.org/10.1038/s41419-021-04272-9.

[30] Y. He, H. Huang, L. Jin, et al., “CircZNF609 Enhances Hepatocellular Carcinoma Cell Proliferation, Metastasis, and Stemness by Activating the Hedgehog Pathway Through the Regulation of miR-15a-5p/15b-5p and GLI2 Expressions,” Cell Death & Disease 11, no. 5 (2020): 358, https://doi.org/10.1038/s41419-020-2441-0.

[31] I. Menéndez-Blanco, S. Soto-Heras, M. G. Catalá, M. Roura, D. Izquierdo, and M. T. Paramio, “Effect of Crocetin Added to IVM Medium for Prepubertal Goat Oocytes on Blastocyst Outcomes After IVF, Intracytoplasmic Sperm Injection and Parthenogenetic Activation,” Theriogenology 155 (2020): 70–76, https://doi.org/10.1016/j.theriogenology.2020.06.008.

[32] G. Yang, Q. Xin, and J. Dean, “Degradation and Translation of Maternal MRNA for Embryogenesis,” Trends in Genetics 40, no. 3 (2024): 238–249, https://doi.org/10.1016/j.tig.2023.12.008.

[33] M. Caudron-Herger, R. E. Jansen, E. Wassmer, and S. Diederichs, “RBP2GO: A Comprehensive Pan-Species Database on RNA-Binding Proteins, Their Interactions and Functions,” Nucleic Acids Research 49, no. D1 (2021): D425–D436, https://doi.org/10.1093/nar/gkaa1040.

[34] Q. Zhou, Y. Yin, M. Yu, et al., “GTPBP4 Promotes Hepatocellular Carcinoma Progression and Metastasis via the PKM2 Dependent Glucose Metabolism,” Redox Biology 56 (2022): 102458, https://doi.org/10.1016/j.redox.2022.102458.

[35] R. Su, L. H. Fan, C. Cao, et al., “Global Profiling of RNA-Binding Protein Target Sites by LACE-Seq,” Nature Cell Biology 23, no. 6 (2021): 664–675, https://doi.org/10.1038/s41556-021-00696-9.

[36] T. Mino, Y. Murakawa, A. Fukao, et al., “Regnase-1 and Roquin Regulate a Common Element in Inflammatory mRNAs by Spatiotemporally Distinct Mechanisms,” Cell 161, no. 5 (2015): 1058–1073, https://doi.org/10.1016/j.cell.2015.04.029.

[37] Z. Zou, C. Zhang, Q. Wang, et al., “Translatome and Transcriptome Co-Profiling Reveals a Role of TPRXs in Human Zygotic Genome Activation,” Science 378, no. 6615 (2022): abo7923, https://doi.org/10.1126/science.abo7923.

[38] Y. Liu, W. Tao, S. Wu, et al., “Maternal MRNA Deadenylation Is Defective in In Vitro Matured Mouse and Human Oocytes,” Nature Communications 15, no. 1 (2024): 5550, https://doi.org/10.1038/s41467-024-49695-y.

[39] R. Iyyappan, D. Aleshkina, H. Ming, et al., “The Translational Oscillation in Oocyte and Early Embryo Development,” Nucleic Acids Research 51, no. 22 (2023): 12076–12091, https://doi.org/10.1093/nar/gkad996.

[40] M. Li, S. Yin, J. Yuan, et al., “Cdc25A Promotes G2/M Transition in Oocytes,” Cell Cycle 7, no. 9 (2008): 1301–1302, https://doi.org/10.4161/cc.7.9.5958.

[41] O. Timofeev, O. Cizmecioglu, F. Settele, T. Kempf, and I. Hoffmann, “Cdc25 Phosphatases Are Required for Timely Assembly of CDK1-Cyclin B at the G2/M Transition,” Journal of Biological Chemistry 285, no. 22 (2010): 16978–16990, https://doi.org/10.1074/jbc.M109.096552.

[42] K. Song, X. Jiang, X. Xu, et al., “Ste20-Like Kinase Activity Promotes Meiotic Resumption and Spindle Microtubule Stability in Mouse Oocytes,” Cell Proliferation 56, no. 4 (2023): e13391, https://doi.org/10.1111/cpr.13391.

[43] C. R. Yang, G. Rajkovic, E. M. Daldello, X. G. Luong, J. Chen, and M. Conti, “The RNA-Binding Protein DAZL Functions as Repressor and Activator of MRNA Translation During Oocyte Maturation,” Nature Communications 11, no. 1 (2020): 1399, https://doi.org/10.1038/s41467-020-15209-9.

[44] K. Lee, K. Cho, R. Morey, and H. Cook-Andersen, “An Extended Wave of Global MRNA Deadenylation Sets Up a Switch in Translation Regulation Across the Mammalian Oocyte-To-Embryo Transition,” Cell Reports 43, no. 2 (2024): 113710, https://doi.org/10.1016/j.celrep.2024.113710.

[45] Y. F. Chen, S. H. Wang, and J. S. Jan, “Peptide-Based Nanoparticles Suppress Hepatic Inflammation via Blockage of Human Antigen R,” Small 30 (2024): e2406963, https://doi.org/10.1002/smll.202406963.

[46] H. Hu, K. Zhao, D. Fang, et al., “The RNA Binding Protein RALY Suppresses p53 Activity and Promotes Lung Tumorigenesis,” Cell Reports 42, no. 4 (2023): 112288, https://doi.org/10.1016/j.celrep.2023.112288.

[47] T. G. Meng, W. L. Lei, J. Li, et al., “Degradation of Ccnb3 Is Essential for Maintenance of MII Arrest in Oocyte,” Biochemical and Biophysical Research Communications 521, no. 1 (2020): 265–269, https://doi.org/10.1016/j.bbrc.2019.10.124.

[48] J. S. Oh, A. Susor, K. Schindler, et al., “Cdc25A Activity Is Required for the Metaphase II Arrest in Mouse Oocytes,” Journal of Cell Science 126, no. Pt 5 (2013): 1081–1085, https://doi.org/10.1242/jcs.115592.

[49] Z. Xiong, K. Xu, Z. Lin, et al., “Ultrasensitive Ribo-Seq Reveals Translational Landscapes During Mammalian Oocyte-To-Embryo Transition and Pre-Implantation Development,” Nature Cell Biology 24, no. 6 (2022): 968–980, https://doi.org/10.1038/s41556-022-00928-6.

[50] Y. Lu, S. Li, Z. Cui, et al., “The Cohesion Establishment Factor Esco1 Acetylates α-Tubulin to Ensure Proper Spindle Assembly in Oocyte Meiosis,” Nucleic Acids Research 46, no. 5 (2018): 2335–2346, https://doi.org/10.1093/nar/gky001.

[51] Y. Lu, X. Dai, M. Zhang, et al., “Cohesin Acetyltransferase Esco2 Regulates SAC and Kinetochore Functions via Maintaining H4K16 Acetylation During Mouse Oocyte Meiosis,” Nucleic Acids Research 45, no. 16 (2017): 9388–9397, https://doi.org/10.1093/nar/gkx563.

[52] C. H. Spruck, M. P. De Miguel, A. P. Smith, et al., “Requirement of Cks2 for the First Metaphase/Anaphase Transition of Mammalian Meiosis,” Science 300, no. 5619 (2003): 647–650, https://doi.org/10.1126/science.1084149.

[53] B. P. Mihalas, G. H. Pieper, M. Aboelenain, et al., “Age-Dependent Loss of Cohesion Protection in Human Oocytes,” Current Biology 34, no. 1 (2024): 117–131, https://doi.org/10.1016/j.cub.2023.11.061.

[54] W. El Yakoubi, E. Buffin, D. Cladière, et al., “Mps1 Kinase-Dependent Sgo2 Centromere Localisation Mediates Cohesin Protection in Mouse Oocyte Meiosis I,” Nature Communications 8, no. 1 (2017): 694, https://doi.org/10.1038/s41467-017-00774-3.

[55] T. Kumon, J. Ma, R. B. Akins, et al., “Parallel Pathways for Recruiting Effector Proteins Determine Centromere Drive and Suppression,” Cell 184, no. 19 (2021): 4904–4918, https://doi.org/10.1016/j.cell.2021.07.037.

[56] T. Wu, J. Dong, J. Fu, et al., “The Mechanism of Acentrosomal Spindle Assembly in Human Oocytes,” Science 378, no. 6621 (2022): eabq7361, https://doi.org/10.1126/science.abq7361.

[57] Y. Wang, L. H. Fan, W. Yue, et al., “CENP-W Regulates Kinetochore-Microtubule Attachment and Meiotic Progression of Mouse Oocytes,” Biochemical and Biophysical Research Communications 527, no. 1 (2020): 8–14, https://doi.org/10.1016/j.bbrc.2020.04.078.

[58] C. J. Zhou, X. Y. Wang, Y. H. Dong, et al., “CENP-F-Dependent DRP1 Function Regulates APC/C Activity During Oocyte Meiosis I,” Nature Communications 13, no. 1 (2022): 7732, https://doi.org/10.1038/s41467-022-35461-5.

[59] R. B. Lakshmi, P. Nayak, L. Raz, et al., “CKAP5 Stabilizes CENP-E at Kinetochores by Regulating Microtubule-Chromosome Attachments,” EMBO Reports 25, no. 4 (2024): 1909–1935, https://doi.org/10.1038/s44319-024-00106-9.

[60] M. E. Pesenti, T. Raisch, D. Conti, et al., “Structure of the Human Inner Kinetochore CCAN Complex and Its Significance for Human Centromere Organization,” Molecular Cell 82, no. 11 (2022): 2113–2131, https://doi.org/10.1016/j.molcel.2022.04.027.

[61] L. Li, S. Zhu, W. Shu, et al., “Characterization of Metabolic Patterns in Mouse Oocytes During Meiotic Maturation,” Molecular Cell 80, no. 3 (2020): 525–540, https://doi.org/10.1016/j.molcel.2020.09.022.

[62] A. Yesilaltay, G. A. Dokshin, D. Busso, et al., “Excess Cholesterol Induces Mouse Egg Activation and May Cause Female Infertility,” Proceedings of the National Academy of Sciences of the United States of America 111, no. 46 (2014): E4972–E4980, https://doi.org/10.1073/pnas.1418954111.

[63] F. Xu and L. Wang, “Deciphering ER Stress-Unfolded Protein Response Relationship by Visualizing Unfolded Proteins in the ER,” Cell Reports 43, no. 6 (2024): 114358, https://doi.org/10.1016/j.celrep.2024.114358.

[64] Y. Wang, S. H. Li, S. J. Yang, et al., “Exposure to Phenanthrene Affects Oocyte Meiosis by Inducing Mitochondrial Dysfunction and Endoplasmic Reticulum Stress,” Cell Proliferation 56, no. 1 (2023): e13335, https://doi.org/10.1111/cpr.13335.

[65] S. A. Wu, C. Shen, X. Wei, et al., “The Mechanisms to Dispose of Misfolded Proteins in the Endoplasmic Reticulum of Adipocytes,” Nature Communications 14, no. 1 (2023): 3132, https://doi.org/10.1038/s41467-023-38690-4.

[66] A. B. Mehrtash and M. Hochstrasser, “Ectopic RING Activity at the ER Membrane Differentially Impacts Erad Protein Quality Control Pathways,” Journal of Biological Chemistry 299, no. 3 (2023): 102927, https://doi.org/10.1016/j.jbc.2023.102927.

[67] K. Yamano, S. Tanaka-Yamano, and T. Endo, “Tom7 Regulates Mdm10-Mediated Assembly of the Mitochondrial Import Channel Protein Tom40,” Journal of Biological Chemistry 285, no. 53 (2010): 41222–41231, https://doi.org/10.1074/jbc.M110.163238.

[68] Y. Araiso, A. Tsutsumi, J. Qiu, et al., “Structure of the Mitochondrial Import Gate Reveals Distinct Preprotein Paths,” Nature 575, no. 7782 (2019): 395–401, https://doi.org/10.1038/s41586-019-1680-7.

[69] C. Kunitomi, M. Romero, E. M. Daldello, et al., “Multiple Intersecting Pathways Are Involved in CPEB1 Phosphorylation and Regulation of Translation During Mouse Oocyte Meiosis,” Development 151, no. 11 (2024): dev202712, https://doi.org/10.1242/dev.202712.

[70] H. F. Noller, “The Ribosome Comes to Life,” Cell 187, no. 23 (2024): 6486–6500, https://doi.org/10.1016/j.cell.2024.10.035.

[71] X. Liang, M. Q. Zuo, Y. Zhang, et al., “Structural Snapshots of Human Pre-60S Ribosomal Particles Before and After Nuclear Export,” Nature Communications 11, no. 1 (2020): 3542, https://doi.org/10.1038/s41467-020-17237-x.

[72] A. Lunardi, G. Di Minin, P. Provero, et al., “A Genome-Scale Protein Interaction Profile of Drosophila p53 Uncovers Additional Nodes of the Human p53 Network,” Proceedings of the National Academy of Sciences of the United States of America 107, no. 14 (2010): 6322–6327, https://doi.org/10.1073/pnas.1002447107.

[73] N. J. Laping, B. A. Olson, and Y. Zhu, “Identification of a Novel Nuclear Guanosine Triphosphate-Binding Protein Differentially Expressed in Renal Disease,” Journal of the American Society of Nephrology 12, no. 5 (2001): 883–890, https://doi.org/10.1681/asn.V125883.

[74] H. Liu, Q. Xue, F. Yang, et al., “Foot-and-Mouth Disease Virus VP1 Degrades YTHDF2 Through Autophagy to Regulate IRF3 Activity for Viral Replication,” Autophagy 20, no. 7 (2024): 1597–1615, https://doi.org/10.1080/15548627.2024.2330105.

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