Integrated analysis and systematic characterization of the regulatory network for human germline development

Yashi Gu; Jiayao Chen; Ziqi Wang; Qizhe Shao; Zhekai Li; Yaxuan Ye; Xia Xiao; Yitian Xiao; Wenyang Liu; Sisi Xie; Lingling Tong; Jin Jiang; Xiaoying Xiao; Ya Yu; Min Jin; Yanxing Wei; Robert S. Young; Lei Hou; Di Chen

doi:10.1016/j.jgg.2024.11.005

Volume 52 Issue 2

Feb. 2025

Turn off MathJax

Article Contents

Article Navigation > Journal of Genetics and Genomics > 2025 > 52(2): 204-219

PDF( 0 KB)

Integrated analysis and systematic characterization of the regulatory network for human germline development

doi: 10.1016/j.jgg.2024.11.005

a. Center for Reproductive Medicine of the Second Affiliated Hospital, Center for Regeneration and Cell Therapy of Zhejiang University-University of Edinburgh Institute (ZJU-UoE Institute), Zhejiang University School of Medicine, Zhejiang University, Hangzhou, Zhejiang 310003, China;
b. Edinburgh Medical School:Biomedical Sciences, College of Medicine and Veterinary Medicine, The University of Edinburgh, Edinburgh, UK;
c. Center for Reproductive Medicine of the Second Affiliated Hospital, Zhejiang University School of Medicine, Zhejiang University, Hangzhou, Zhejiang 310003, China;
d. Department of Obstetrics and Gynecology, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong 510515, China;
e. Key Laboratory of Functional Proteomics of Guangdong Province, School of Basic Medical Sciences, Southern Medical University, Guangzhou, Guangdong 510515, China;
f. Research Centre for Women's and Infants' Health, Lunenfeld-Tanenbaum Research Institute, Sinai Health System, Toronto, M5T3H7, Canada;
g. Center for Global Health Research, Usher Institute, University of Edinburgh, Teviot Place, 5-7 Little France Road, Edinburgh BioQuarter-Gate 3, Edinburgh, EH16 4UX, UK;
h. Zhejiang University-University of Edinburgh Institute, Zhejiang University, Haining, Zhejiang 314400, China;
i. Section of Biomedical Genetics, Department of Medicine, Boston University School of Medicine, Boston, MA 02118, United States;
j. State Key Laboratory of Biobased Transportation Fuel Technology, Haining, Zhejiang 314400, China

Received Date: 2024-07-13
Accepted Date: 2024-11-11
Rev Recd Date: 2024-11-10

Available Online: 2025-07-11

Publish Date: 2024-11-19

Abstract

Abstract

Primordial germ cells (PGCs) are the precursors of germline that are specified at the embryonic stage. Recent studies reveal that humans employ different mechanisms for PGC specification compared with model organisms such as mice. Moreover, the specific regulatory machinery remains largely unexplored, mainly due to the inaccessible nature of this complex biological process in humans. Here, we curate and integrate multi-omics data, including 581 RNA-seq, 54 ATAC-seq, 45 ChIP-seq, and 69 single-cell RNA-seq samples from different stages of human PGC development to recapitulate the precisely controlled and stepwise process, presenting an atlas in the human PGC database (hPGCdb). With these uniformly processed data and integrated analyses, we characterize the potential key transcription factors and regulatory networks governing human germ cell fate. We validate the important roles of some of the key factors in germ cell development by CRISPRi knockdown. We also identify the soma-germline interaction network and discover the involvement of SDC2 and LAMA4 for PGC development, as well as soma-derived NOTCH2 signaling for germ cell differentiation. Taken together, we have built a database for human PGCs (http://43.131.248.15:6882) and demonstrate that hPGCdb enables the identification of the missing pieces of mechanisms governing germline development, including both intrinsic and extrinsic regulatory programs.
- Primordial germ cells,
- Regulatory network,
- Soma-germ cell interaction,
- Database

FullText(HTML)

References(72)

References

Becht, E., McInnes, L., Healy, J., Dutertre, C.-A., Kwok, I.W.H., Ng, L.G., Ginhoux, F., Newell, E.W., 2019. Dimensionality reduction for visualizing single-cell data using UMAP. Nat. Biotechnol. 37, 38-44.

Campolo, F., Gori, M., Favaro, R., Nicolis, S., Pellegrini, M., Botti, F., Rossi, P., Jannini, E.A., Dolci, S., 2013. Essential Role of Sox2 for the Establishment and Maintenance of the Germ Cell Line. STEM CELLS 31, 1408-1421.

Chen, D., Gell, J.J., Tao, Y., Sosa, E., Clark, A.T., 2017a. Modeling human infertility with pluripotent stem cells. Stem Cell Res. 21, 187-192.

Chen, D., Liu, W., Lukianchikov, A., Hancock, G.V., Zimmerman, J., Lowe, M.G., Kim, R., Galic, Z., Irie, N., Surani, M.A., Jacobsen, S.E., Clark, A.T., 2017b. Germline competency of human embryonic stem cells depends on EOMESodermin. Biol. Reprod. 97, 850-861.

Chen, D., Liu, W., Zimmerman, J., Pastor, W.A., Kim, R., Hosohama, L., Ho, J., Aslanyan, M., Gell, J.J., Jacobsen, S.E., Clark, A.T., 2018a. The TFAP2C-Regulated OCT4 Naive Enhancer Is Involved in Human Germline Formation. Cell Rep. 25, 3591-3602.e5.

Chen, D., Liu, W., Zimmerman, J., Pastor, W.A., Kim, R., Hosohama, L., Ho, J., Aslanyan, M., Gell, J.J., Jacobsen, S.E., Clark, A.T., 2018b. The TFAP2C-Regulated OCT4 Naive Enhancer Is Involved in Human Germline Formation. Cell Rep. 25, 3591-3602.e5.

Chen, D., Sun, N., Hou, L., Kim, R., Faith, J., Aslanyan, M., Tao, Y., Zheng, Y., Fu, J., Liu, W., Kellis, M., Clark, A., 2019. Human Primordial Germ Cells Are Specified from Lineage-Primed Progenitors. Cell Rep. 29, 4568-4582.e5.

Chen, L., Wang, D., Wu, Z., Ma, L., Daley, G.Q., 2010. Molecular basis of the first cell fate determination in mouse embryogenesis. Cell Res. 20, 982-993.

Chen, M., Long, X., Chen, M., Hao, F., Kang, J., Wang, N., Wang, Y., Wang, M., Gao, Y., Zhou, M., et al., 2022. Integration of single-cell transcriptome and chromatin accessibility of early gonads development among goats, pigs, macaques, and humans. Cell Rep. 41, 111587.

Chitiashvili, T., Dror, I., Kim, R., Hsu, F.-M., Chaudhari, R., Pandolfi, E., Chen, D., Liebscher, S., Schenke-Layland, K., Plath, K., et al., 2020. Female human primordial germ cells display X-chromosome dosage compensation despite the absence of X-inactivation. Nat. Cell Biol. 22, 1436-1446.

Dai, M., Pei, X., Wang, X.-J., 2022. Accurate and fast cell marker gene identification with COSG. Brief. Bioinform. 23, bbab579.

Fang, F., Angulo, B., Xia, N., Sukhwani, M., Wang, Z., Carey, C.C., Mazurie, A., Cui, J., Wilkinson, R., Wiedenheft, B., et al., 2018. A PAX5-OCT4-PRDM1 developmental switch specifies human primordial germ cells. Nat. Cell Biol. 20, 655-665.

Farrell, J.A., Wang, Y., Riesenfeld, S.J., Shekhar, K., Regev, A., Schier, A.F., 2018. Single-cell reconstruction of developmental trajectories during zebrafish embryogenesis. Science 360.

Feregrino, C., Tschopp, P., 2022. Assessing evolutionary and developmental transcriptome dynamics in homologous cell types. Dev. Dyn. 251, 1472-1489.

Garcia-Castro, M.I., Anderson, R., Heasman, J., Wylie, C., 1997. Interactions between Germ Cells and Extracellular Matrix Glycoproteins during Migration and Gonad Assembly in the Mouse Embryo. J. Cell Biol. 138, 471-480.

Gell, J.J., Liu, W., Sosa, E., Chialastri, A., Hancock, G., Tao, Y., Wamaitha, S.E., Bower, G., Dey, S.S., Clark, A.T., 2020. An Extended Culture System that Supports Human Primordial Germ Cell-like Cell Survival and Initiation of DNA Methylation Erasure. Stem Cell Rep. 14, 433-446.

Gkountela, S., Zhang, K.X., Shafiq, T.A., Liao, W.-W., Hargan-Calvopina, J., Chen, P.-Y., Clark, A.T., 2015. DNA Demethylation Dynamics in the Human Prenatal Germline. Cell 161, 1425-1436.

Gopal, S., Amran, A., Elton, A., Ng, L., Pocock, R., 2021. A somatic proteoglycan controls Notch-directed germ cell fate. Nat. Commun. 12, 6708.

Guo, J., Grow, E.J., Mlcochova, H., Maher, G.J., Lindskog, C., Nie, X., Guo, Y., Takei, Y., Yun, J., Cai, L., et al., 2018. The adult human testis transcriptional cell atlas. Cell Res. 28, 1141-1157.

Haghverdi, L., Lun, A.T.L., Morgan, M.D., Marioni, J.C., 2018. Batch effects in single-cell RNA-sequencing data are corrected by matching mutual nearest neighbors. Nat. Biotechnol. 36, 421-427.

Hancock, G.V., Wamaitha, S.E., Peretz, L., Clark, A.T., 2021. Mammalian primordial germ cell specification. Development 148, dev189217.

Hao, Y., Hao, S., Andersen-Nissen, E., Mauck, W.M., Zheng, S., Butler, A., Lee, M.J., Wilk, A.J., Darby, C., Zager, M., et al., 2021. Integrated analysis of multimodal single-cell data. Cell 184, 3573-3587.e29.

Hara, K., Kanai-Azuma, M., Uemura, M., Shitara, H., Taya, C., Yonekawa, H., Kawakami, H., Tsunekawa, N., Kurohmaru, M., Kanai, Y., 2009. Evidence for crucial role of hindgut expansion in directing proper migration of primordial germ cells in mouse early embryogenesis. Dev. Biol. 330, 427-439.

Hwang, Y.S., Suzuki, S., Seita, Y., Ito, J., Sakata, Y., Aso, H., Sato, K., Hermann, B.P., Sasaki, K., 2020. Reconstitution of prospermatogonial specification in vitro from human induced pluripotent stem cells. Nat. Commun. 11, 5656.

Inhorn, M.C., Patrizio, P., 2015. Infertility around the globe: new thinking on gender, reproductive technologies and global movements in the 21st century. Hum. Reprod. Updat. 21, 411-426.

Irie, N., Lee, S.-M., Lorenzi, V., Xu, H., Chen, J., Inoue, M., Kobayashi, T., Sancho-Serra, C., Drousioti, E., Dietmann, S., et al., 2023. DMRT1 regulates human germline commitment. Nat. Cell Biol. 25, 1439-1452.

Irie, N., Weinberger, L., Tang, W.W.C., Kobayashi, T., Viukov, S., Manor, Y.S., Dietmann, S., Hanna, J.H., Surani, M.A., 2015. SOX17 Is a Critical Specifier of Human Primordial Germ Cell Fate. Cell 160, 253-268.

Jin, S., Guerrero-Juarez, C.F., Zhang, L., Chang, I., Ramos, R., Kuan, C.-H., Myung, P., Plikus, M.V., Nie, Q., 2021. Inference and analysis of cell-cell communication using CellChat. Nat. Commun. 12, 1088.

Kobayashi, T., Zhang, H., Tang, W.W.C., Irie, N., Withey, S., Klisch, D., Sybirna, A., Dietmann, S., Contreras, D.A., Webb, R., et al., 2017. Principles of early human development and germ cell program from conserved model systems. Nature 546, 416-420.

Kojima, Y., Sasaki, K., Yokobayashi, S., Sakai, Y., Nakamura, T., Yabuta, Y., Nakaki, F., Nagaoka, S., Woltjen, K., Hotta, A., et al., 2017. Evolutionarily Distinctive Transcriptional and Signaling Programs Drive Human Germ Cell Lineage Specification from Pluripotent Stem Cells. Cell Stem Cell 21, 517-532.e5.

Kojima, Y., Yamashiro, C., Murase, Y., Yabuta, Y., Okamoto, I., Iwatani, C., Tsuchiya, H., Nakaya, M., Tsukiyama, T., Nakamura, T., et al., 2021. GATA transcription factors, SOX17 and TFAP2C, drive the human germ-cell specification program. Life Sci. Alliance 4, e202000974.

Langfelder, P., Horvath, S., 2008a. WGCNA: an R package for weighted correlation network analysis. BMC Bioinform. 9, 559.

Langmead, B., Salzberg, S.L., 2012. Fast gapped-read alignment with Bowtie 2. Nat. Methods 9, 357-359.

Langmead, B., Trapnell, C., Pop, M., Salzberg, S.L., 2009. Ultrafast and memory-efficient alignment of short DNA sequences to the human genome. Genome Biol. 10, R25.

Larose, H., Shami, A.N., Abbott, H., Manske, G., Lei, L., Hammoud, S.S., 2019. Gametogenesis: A journey from inception to conception. Curr. Top. Dev. Biol. 132, 257-310.

Li, L., Dong, J., Yan, L., Yong, J., Liu, X., Hu, Y., Fan, X., Wu, X., Guo, H., Wang, X., et al., 2017. Single-Cell RNA-Seq Analysis Maps Development of Human Germline Cells and Gonadal Niche Interactions. Cell Stem Cell 20, 858-873.e4.

Liao, Y., Smyth, G.K., Shi, W., 2014. featureCounts: an efficient general purpose program for assigning sequence reads to genomic features. Bioinformatics 30, 923-930.

Love, M.I., Huber, W., Anders, S., 2014. Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biol. 15, 550.

Magnusdottir, E., Dietmann, S., Murakami, K., Gunesdogan, U., Tang, F., Bao, S., Diamanti, E., Lao, K., Gottgens, B., Surani, M.A., 2013. A tripartite transcription factor network regulates primordial germ cell specification in mice. Nat. Cell Biol. 15, 905-915.

Miao, Q., Hill, M.C., Chen, F., Mo, Q., Ku, A.T., Ramos, C., Sock, E., Lefebvre, V., Nguyen, H., 2019. SOX11 and SOX4 drive the reactivation of an embryonic gene program during murine wound repair. Nat. Commun. 10, 4042.

Moreno, C.S., 2020. SOX4: The unappreciated oncogene. Semin. Cancer Biol. 67, 57-64.

Murase, Y., Yabuta, Y., Ohta, H., Yamashiro, C., Nakamura, T., Yamamoto, T., Saitou, M., 2020. Long-term expansion with germline potential of human primordial germ cell-like cells in vitro. EMBO J. 39, e104929.

Nakaki, F., Hayashi, K., Ohta, H., Kurimoto, K., Yabuta, Y., Saitou, M., 2013. Induction of mouse germ-cell fate by transcription factors in vitro. Nature 501, 222-226.

Onyeisi, J.O.S., Lopes, C.C., Gotte, M., 2021. Syndecan-4 as a Pathogenesis Factor and Therapeutic Target in Cancer. Biomolecules 11, 503.

Pastor, W.A., Chen, D., Liu, W., Kim, R., Sahakyan, A., Lukianchikov, A., Plath, K., Jacobsen, S.E., Clark, A.T., 2016. Naive Human Pluripotent Cells Feature a Methylation Landscape Devoid of Blastocyst or Germline Memory. Cell Stem Cell 18, 323-329.

Ramirez, F., Dundar, F., Diehl, S., Gruning, B.A., Manke, T., 2014. deepTools: a flexible platform for exploring deep-sequencing data. Nucleic Acids Res. 42, W187-W191.

Richardson, B.E., Lehmann, R., 2010. Mechanisms guiding primordial germ cell migration: strategies from different organisms. Nat. Rev. Mol. Cell Biol. 11, 37-49.

Ritchie, M.E., Phipson, B., Wu, D., Hu, Y., Law, C.W., Shi, W., Smyth, G.K., 2015. limma powers differential expression analyses for RNA-sequencing and microarray studies. Nucleic Acids Res. 43, e47-e47.

Saitou, M., Hayashi, K., 2021. Mammalian in vitro gametogenesis. Science 374, eaaz6830.

Saitou, M., Miyauchi, H., 2016. Gametogenesis from Pluripotent Stem Cells. Cell Stem Cell 18, 721-735.

Sasaki, K., Nakamura, T., Okamoto, I., Yabuta, Y., Iwatani, C., Tsuchiya, H., Seita, Y., Nakamura, S., Shiraki, N., Takakuwa, T., et al., 2016. The Germ Cell Fate of Cynomolgus Monkeys Is Specified in the Nascent Amnion. Dev. Cell 39, 169-185.

Sasaki, K., Yokobayashi, S., Nakamura, T., Okamoto, I., Yabuta, Y., Kurimoto, K., Ohta, H., Moritoki, Y., Iwatani, C., Tsuchiya, H., et al., 2015. Robust In Vitro Induction of Human Germ Cell Fate from Pluripotent Stem Cells. Cell Stem Cell 17, 178-194.

Sosa, E., Chen, D., Rojas, E.J., Hennebold, J.D., Peters, K.A., Wu, Z., Lam, T.N., Mitchell, J.M., Sukhwani, M., Tailor, R.C., et al., 2018. Differentiation of primate primordial germ cell-like cells following transplantation into the adult gonadal niche. Nat. Commun. 9, 5339.

Sybirna, A., Tang, W.W.C., Smela, M.P., Dietmann, S., Gruhn, W.H., Brosh, R., Surani, M.A., 2020. A critical role of PRDM14 in human primordial germ cell fate revealed by inducible degrons. Nat. Commun. 11, 1282.

Tang, W.W.C., Dietmann, S., Irie, N., Leitch, H.G., Floros, V.I., Bradshaw, C.R., Hackett, J.A., Chinnery, P.F., Surani, M.A., 2015. A Unique Gene Regulatory Network Resets the Human Germline Epigenome for Development. Cell 161, 1453-1467.

Tang, W.W.C., Kobayashi, T., Irie, N., Dietmann, S., Surani, M.A., 2016. Specification and epigenetic programming of the human germ line. Nat. Rev. Genet. 17, 585-600.

Tarasov, A., Vilella, A.J., Cuppen, E., Nijman, I.J., Prins, P., 2015. Sambamba: fast processing of NGS alignment formats. Bioinformatics 31, 2032-2034.

Trapnell, C., Cacchiarelli, D., Grimsby, J., Pokharel, P., Li, S., Morse, M., Lennon, N.J., Livak, K.J., Mikkelsen, T.S., Rinn, J.L., 2014. The dynamics and regulators of cell fate decisions are revealed by pseudotemporal ordering of single cells. Nat. Biotechnol. 32, 381-386.

Wang, L., Trasanidis, N., Wu, T., Dong, G., Hu, M., Bauer, D.E., Pinello, L., 2023. Dictys: dynamic gene regulatory network dissects developmental continuum with single-cell multiomics. Nat. Methods 20, 1368-1378.

Wang, Xiaoman, Veerapandian, V., Yang, X., Song, K., Xu, X., Cui, M., Yuan, W., Huang, Y., Xia, X., Yao, Z., Wan, C., et al., 2021. The chromatin accessibility landscape reveals distinct transcriptional regulation in the induction of human primordial germ cell-like cells from pluripotent stem cells. Stem Cell Rep. 16, 1245-1261.

Wilkinson, A.L., Zorzan, I., Rugg-Gunn, P.J., 2023. Epigenetic regulation of early human embryo development. Cell Stem Cell 30, 1569-1584.

Wolock, S.L., Lopez, R., Klein, A.M., 2019. Scrublet: Computational Identification of Cell Doublets in Single-Cell Transcriptomic Data. Cell Syst. 8, 281-291.e9.

Wu, T., Hu, E., Xu, S., Chen, M., Guo, P., Dai, Z., Feng, T., Zhou, L., Tang, W., Zhan, L., et al., 2021. clusterProfiler 4.0: A universal enrichment tool for interpreting omics data. Innov. 2, 100141.

Xiao, L., Ma, L., Wang, Z., Yu, Y., Lye, S.J., Shan, Y., Wei, Y., 2020. Deciphering a distinct regulatory network of TEAD4, CDX2 and GATA3 in humans for trophoblast transition from embryonic stem cells. Biochim. Biophys. Acta (BBA) - Mol. Cell Res. 1867, 118736.

Yagi, R., Kohn, M.J., Karavanova, I., Kaneko, K.J., Vullhorst, D., DePamphilis, M.L., Buonanno, A., 2007. Transcription factor TEAD4 specifies the trophectoderm lineage at the beginning of mammalian development. Development 134, 3827-3836.

Yamashiro, C., Sasaki, K., Yabuta, Y., Kojima, Y., Nakamura, T., Okamoto, I., Yokobayashi, S., Murase, Y., Ishikura, Y., Shirane, K., et al., 2018. Generation of human oogonia from induced pluripotent stem cells in vitro. Science 362, 356-360.

Yokobayashi, S., Okita, K., Nakagawa, M., Nakamura, T., Yabuta, Y., Yamamoto, T., Saitou, M., 2017. Clonal variation of human induced pluripotent stem cells for induction into the germ cell fate. Biol. Reprod. 96, 1154-1166.

Yu, L., Wei, Y., Sun, H.-X., Mahdi, A.K., Arteaga, C.A.P., Sakurai, M., Schmitz, D.A., Zheng, C., Ballard, E.D., Li, J., et al., 2021. Derivation of Intermediate Pluripotent Stem Cells Amenable to Primordial Germ Cell Specification. Cell Stem Cell 28, 550-567.e12.

Zhang, Y., Liu, T., Meyer, C.A., Eeckhoute, J., Johnson, D.S., Bernstein, B.E., Nusbaum, C., Myers, R.M., Brown, M., Li, W., Liu, X.S., 2008. Model-based Analysis of ChIP-Seq (MACS). Genome Biol. 9, R137.

Zhao, L., Arsenault, M., Ng, E.T., Longmuss, E., Chau, T.C.-Y., Hartwig, S., Koopman, P., 2017. SOX4 regulates gonad morphogenesis and promotes male germ cell differentiation in mice. Dev. Biol. 423, 46-56.

Zhou, Y., Huang, T., Cheng, A.S.L., Yu, J., Kang, W., To, K.F., 2016. The TEAD Family and Its Oncogenic Role in Promoting Tumorigenesis. Int. J. Mol. Sci. 17, 138.

Zhu, T., Zhou, X., You, Y., Wang, L., He, Z., Chen, D., 2023. cisDynet: An integrated platform for modeling gene-regulatory dynamics and networks. iMeta 2, e152.

Relative Articles

Supplements(0)

Cited By

Proportional views