{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,25]],"date-time":"2025-10-25T04:31:54Z","timestamp":1761366714377,"version":"build-2065373602"},"reference-count":40,"publisher":"MDPI AG","issue":"21","license":[{"start":{"date-parts":[[2025,10,23]],"date-time":"2025-10-23T00:00:00Z","timestamp":1761177600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100012166","name":"National Key Research and Development Program of China","doi-asserted-by":"publisher","award":["2021YFF1000702"],"award-info":[{"award-number":["2021YFF1000702"]}],"id":[{"id":"10.13039\/501100012166","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Animals"],"abstract":"<jats:p>(1) Background: This study aims to systematically identify key candidate genes and the regulatory networks governing ovarian development in sheep breeds with divergent fecundity. Focusing on elucidating the central regulatory roles of these factors during distinct ovarian developmental stages in highly prolific breeds, the research seeks to reveal the mechanism by which multilevel regulatory networks synergistically determine ewe reproductive capacity. (2) Methods: This study utilized the ovaries from the low-fecundity sheep breed Ujumqin sheep, the high-fecundity breed small-tailed Han sheep, and various developmental stages of small-tailed Han sheep as research subjects. Through whole-transcriptome sequencing analysis, differentially expressed mRNAs(DEGs) and non-coding RNAs (ncRNAs) were screened, and a ceRNA regulatory network was constructed and subjected to bioinformatic analysis. The dual-luciferase reporter gene detection system was employed to validate the targeting relationships within the obtained key circRNA-miRNA-mRNA networks. Finally, qRT-PCR was used to verify the accuracy of the sequencing results. (3) Results: Our analysis constructed two distinct ceRNA networks: one from different fecundity groups (116 DECs, 46 DEMs, 82 DEGs) and another from different ovarian stages (186 DECs, 143 DEMs, 338 DEGs). Functional enrichment revealed key reproduction-related pathways, including Mitogen-Activated Protein Kinase(MAPK), Janus Kinase-Signal Transducer and Activator of Transcription(JAK-STAT), and WNT signaling in the fecundity comparison, and MAPK, Ras, WNT, Hippo signaling in the developmental stage comparison. Integrated analysis identified a core circRNA-miRNA-mRNA network, pinpointing circRNA2058-miR-9226-5p-MET as a central regulatory axis. The dual-luciferase assay confirmed that circRNA2058 acts as a sponge for miR-9226-5p, thereby mediating MET expression. qRT-PCR validation of randomly selected RNAs confirmed the sequencing reliability. (4) Conclusions: this study deciphers a synergistic regulatory network and identifies, for the first time, the pivotal circRNA2058-miR-9226-5p-MET ceRNA axis as an potential critical molecular switch driving follicular dominance in sheep. This discovery provides a molecular foundation for targeting core regulators of ovine reproductive efficiency and offers significant insights for innovative strategies in enhancing sheep reproduction.<\/jats:p>","DOI":"10.3390\/ani15213077","type":"journal-article","created":{"date-parts":[[2025,10,24]],"date-time":"2025-10-24T01:35:11Z","timestamp":1761269711000},"page":"3077","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Integrated Whole-Transcriptome Analysis to Elucidate the Core Regulatory Network of circRNA Involved in Ovarian Development and Reproductive Capacity Differences in Sheep: circRNA2058-miR-9226-5p-MET Axis"],"prefix":"10.3390","volume":"15","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-4281-1621","authenticated-orcid":false,"given":"Bo","family":"Gu","sequence":"first","affiliation":[{"name":"College of Life Science, Jilin Normal University, Siping 136000, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Anqi","family":"Wang","sequence":"additional","affiliation":[{"name":"College of Animal Science and Technology, Jilin Agricultural University, Changchun 130118, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Xinmiao","family":"Yu","sequence":"additional","affiliation":[{"name":"College of Life Science, Jilin Normal University, Siping 136000, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Ying","family":"Li","sequence":"additional","affiliation":[{"name":"College of Life Science, Jilin Normal University, Siping 136000, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yao","family":"Cong","sequence":"additional","affiliation":[{"name":"College of Life Science, Jilin Normal University, Siping 136000, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Huaizhi","family":"Jiang","sequence":"additional","affiliation":[{"name":"College of Animal Science and Technology, Jilin Agricultural University, Changchun 130118, China"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2025,10,23]]},"reference":[{"key":"ref_1","first-page":"30","article-title":"An Introduction to High-Fertility Sheep Breeds","volume":"30","author":"Yang","year":"2003","journal-title":"China Anim. Husb. Vet. Med."},{"key":"ref_2","first-page":"3","article-title":"Research Progress on Genetic Diversity of Xilingol Sheep Breeds","volume":"16","author":"Zhang","year":"2020","journal-title":"Chin. Livest. Poult. Breed."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"257","DOI":"10.1016\/j.ygeno.2017.10.005","article-title":"Circular RNA profiling reveals chi_circ_0008219 function as microRNA sponges in pre-ovulatory ovarian follicles of goats (Capra hircus)","volume":"110","author":"Tao","year":"2017","journal-title":"Genomics"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"29","DOI":"10.1186\/s40813-022-00270-1","article-title":"CircRNAs in Xiang pig ovaries among diestrus and estrus stages","volume":"8","author":"Niu","year":"2022","journal-title":"Porc. Health Manag."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"62","DOI":"10.1016\/j.theriogenology.2022.10.034","article-title":"CiRS-187 regulates BMPR2 expression by targeting miR-187 in bovine cumulus cells treated with BMP15 and GDF9","volume":"197","author":"Fu","year":"2023","journal-title":"Theriogenology"},{"key":"ref_6","doi-asserted-by":"crossref","unstructured":"Wang, H., Zhang, Y., Zhang, J., Du, X., Li, Q., and Pan, Z. (2022). circSLC41A1 Resists Porcine Granulosa Cell Apoptosis and Follicular Atresia by Promoting SRSF1 through miR-9820-5p Sponging. Int. J. Mol. Sci., 23.","DOI":"10.3390\/ijms23031509"},{"key":"ref_7","doi-asserted-by":"crossref","unstructured":"Liu, Y., Wang, P., Zhou, Z., He, X., Tao, L., Jiang, Y., Lan, R., Hong, Q., and Chu, M. (2022). Expression Profile Analysis to Identify Circular RNA Expression Signatures in the Prolificacy Trait of Yunshang Black Goat Pituitary in the Estrus Cycle. Front. Genet., 12.","DOI":"10.3389\/fgene.2021.801357"},{"key":"ref_8","doi-asserted-by":"crossref","unstructured":"Wang, J., Chen, H., Zhang, Y., Jiang, S., Zeng, X., and Shen, H. (2023). Comprehensive Analysis of Differentially Expressed CircRNAs in the Ovaries of Low- and High-Fertility Sheep. Animals, 13.","DOI":"10.3390\/ani13020236"},{"key":"ref_9","doi-asserted-by":"crossref","unstructured":"Lv, W., An, R., Li, X., Zhang, Z., Geri, W., Xiong, X., Yin, S., Fu, W., Liu, W., and Lin, Y. (2024). Multi-Omics Approaches Uncovered Critical mRNA-miRNA-lncRNA Networks Regulating Multiple Birth Traits in Goat Ovaries. Int. J. Mol. Sci., 25.","DOI":"10.3390\/ijms252212466"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"1138","DOI":"10.1089\/cmb.2017.0096","article-title":"cutPrimers: A New Tool for Accurate Cutting of Primers from Reads of Targeted Next Generation Sequencing","volume":"24","author":"Kechin","year":"2017","journal-title":"J. Comput. Biol."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"907","DOI":"10.1038\/s41587-019-0201-4","article-title":"Graph-based genome alignment and genotyping with HISAT2 and HISAT-genotype","volume":"37","author":"Kim","year":"2019","journal-title":"Nat. Biotechnol."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"357","DOI":"10.1038\/nmeth.3317","article-title":"HISAT: A fast spliced aligner with low memory requirements","volume":"12","author":"Kim","year":"2015","journal-title":"Nat. Methods"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"1650","DOI":"10.1038\/nprot.2016.095","article-title":"Transcript-level expression analysis of RNA-seq experiments with HISAT, StringTie and Ballgown","volume":"11","author":"Pertea","year":"2016","journal-title":"Nat. Protoc."},{"key":"ref_14","doi-asserted-by":"crossref","unstructured":"Kovaka, S., Zimin, A.V., Pertea, G.M., Razaghi, R., Salzberg, S.L., and Pertea, M. (2019). Transcriptome assembly from long-read RNA-seq alignments with StringTie2. Genome Biol., 20.","DOI":"10.1186\/s13059-019-1910-1"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"290","DOI":"10.1038\/nbt.3122","article-title":"StringTie enables improved reconstruction of a transcriptome from RNA-seq reads","volume":"33","author":"Pertea","year":"2015","journal-title":"Nat. Biotechnol."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"1277","DOI":"10.1101\/gr.202895.115","article-title":"Diverse alternative back-splicing and alternative splicing landscape of circular RNAs","volume":"26","author":"Zhang","year":"2016","journal-title":"Genome Res."},{"key":"ref_17","doi-asserted-by":"crossref","unstructured":"Gao, Y., Wang, J., and Zhao, F. (2015). CIRI: An efficient and unbiased algorithm for de novo circular RNA identification. Genome Biol., 16.","DOI":"10.1186\/s13059-014-0571-3"},{"key":"ref_18","doi-asserted-by":"crossref","unstructured":"Ye, J., Yao, Z., Si, W., Gao, X., Yang, C., Liu, Y., Ding, J., Huang, W., Fang, F., and Zhou, J. (2018). Identification and characterization of microRNAs in the pituitary of pubescent goats. Reprod. Biol. Endocrinol., 16.","DOI":"10.1186\/s12958-018-0370-x"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"8015","DOI":"10.18632\/aging.102315","article-title":"Circular RNA profiling in the oocyte and cumulus cells reveals that circARMC4 is essential for porcine oocyte maturation","volume":"11","author":"Cao","year":"2019","journal-title":"Aging"},{"key":"ref_20","doi-asserted-by":"crossref","unstructured":"Liu, J., Feng, G., Guo, C., Li, Z., Liu, D., Liu, G., Zou, X., Sun, B., Guo, Y., and Deng, M. (2024). Identification of functional circRNAs regulating ovarian follicle development in goats. BMC Genom., 25.","DOI":"10.1186\/s12864-024-10834-w"},{"key":"ref_21","doi-asserted-by":"crossref","unstructured":"Huang, C., Feng, F., Dai, R., Ren, W., Li, X., Zhaxi, T., Ma, X., Wu, X., Chu, M., and La, Y. (2024). Whole-transcriptome analysis of longissimus dorsi muscle in cattle-yaks reveals the regulatory functions of ADAMTS6 gene in myoblasts. Int. J. Biol. Macromol., 262.","DOI":"10.1016\/j.ijbiomac.2024.129985"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"33","DOI":"10.1186\/s12935-020-01730-w","article-title":"Linc00887 suppresses tumorigenesis of cervical cancer through regulating the miR-454-3p\/FRMD6-Hippo axis","volume":"21","author":"Li","year":"2021","journal-title":"Cancer Cell Int."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"1189","DOI":"10.1038\/onc.2011.318","article-title":"Human homolog of Drosophila expanded, hEx, functions as a putative tumor suppressor in human cancer cell lines independently of the Hippo pathway","volume":"31","author":"Hao","year":"2012","journal-title":"Oncogene"},{"key":"ref_24","first-page":"1321","article-title":"Research Progress on Cyclins and Cyclin-dependent Kinases Related with Primordial Follicle Activation","volume":"43","author":"Yang","year":"2024","journal-title":"Genom. Appl. Biol."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"bqab238","DOI":"10.1210\/endocr\/bqab238","article-title":"The Hippo Pathway Effectors YAP and TAZ Regulate LH Release by Pituitary Gonadotrope Cells in Mice","volume":"163","author":"Boyer","year":"2022","journal-title":"Endocrinology"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"1001","DOI":"10.1093\/biolre\/ioz139","article-title":"Yes-associated protein 1 is required for proliferation and function of bovine granulosa cells in vitro","volume":"101","author":"Plewes","year":"2019","journal-title":"Biol. Reprod."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"71","DOI":"10.1186\/s13048-016-0280-5","article-title":"Differential regulation of Janus kinase 3 (JAK3) in bovine preovulatory follicles and identification of JAK3 interacting proteins in granulosa cells","volume":"9","author":"Ndiaye","year":"2016","journal-title":"J. Ovarian Res."},{"key":"ref_28","doi-asserted-by":"crossref","unstructured":"Yue, J., and L\u00f3pez, J.M. (2020). Understanding MAPK Signaling Pathways in Apoptosis. Int. J. Mol. Sci., 21.","DOI":"10.3390\/ijms21072346"},{"key":"ref_29","doi-asserted-by":"crossref","unstructured":"Xu, H., Cai, Y., Yang, H., Zhang, C., Liu, W., Zhao, B., Wang, F., and Zhang, Y. (2025). AMH regulates granulosa cell function via ESR2\/p38-MAPK signaling pathway in sheep. Commun. Biol., 8.","DOI":"10.1038\/s42003-025-08259-6"},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"137","DOI":"10.1016\/j.theriogenology.2021.12.024","article-title":"Effect of RFRP-3, the mammalian ortholog of GnIH, on apoptosis and autophagy in porcine ovarian granulosa cells via the p38MAPK pathway","volume":"180","author":"Zhang","year":"2022","journal-title":"Theriogenology"},{"key":"ref_31","doi-asserted-by":"crossref","unstructured":"Wang, J., Bai, J., Liu, Y., Zhang, Z., Lv, Z., Hai, G., Li, H., Liu, W., Tang, L., and Hua, Y. (2025). Transcriptomic characterization of lamb ovaries and oocytes reveals key biomarkers after superovulation. J. Anim. Sci., skaf193.","DOI":"10.1093\/jas\/skaf193"},{"key":"ref_32","doi-asserted-by":"crossref","unstructured":"Li, G., Yan, L., Wang, L., Ma, W., Wu, H., Guan, S., Yao, Y., Deng, S., Yang, H., and Zhang, J. (2024). Ovarian overexpression of ASMT gene increases follicle numbers in transgenic sheep: Association with lipid metabolism. Int. J. Biol. Macromol., 269.","DOI":"10.1016\/j.ijbiomac.2024.131803"},{"key":"ref_33","unstructured":"Song, Z. (2020). Association Analysis of SNPs in CTNNB1 and pri-miR-458b with Laying Traits in Layers. [Master\u2019s Thesis, Shandong Agricultural University]."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"e13686","DOI":"10.1111\/cpr.13686","article-title":"MicroRNA-3061 downregulates the expression of PAX7\/Wnt\/Ca2+ signalling axis genes to induce premature ovarian failure in mice","volume":"57","author":"Liu","year":"2024","journal-title":"Cell Prolif."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"373","DOI":"10.1038\/s41419-020-2578-x","article-title":"SMAD4 activates Wnt signaling pathway to inhibit granulosa cell apoptosis","volume":"11","author":"Du","year":"2020","journal-title":"Cell Death Dis."},{"key":"ref_36","first-page":"3413","article-title":"The research progress of HGF\/c-MET signaling pathway regulating reproductive cytogenesis","volume":"35","author":"Li","year":"2019","journal-title":"J. Pract. Med."},{"key":"ref_37","first-page":"3769","article-title":"Association Analysis of MER Gene Single Nucleotide Polymorphism with Reproduction and Milk Production on Traits in Chinese Holstein Cattle","volume":"53","author":"Xu","year":"2022","journal-title":"Acta Vet. Et Zootech. Sin."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"46","DOI":"10.1186\/s13287-019-1136-x","article-title":"Human amnion-derived mesenchymal stem cell (hAD-MSC) transplantation improves ovarian function in rats with premature ovarian insufficiency (POI) at least partly through a paracrine mechanism","volume":"10","author":"Ling","year":"2019","journal-title":"Stem Cell Res. Ther."},{"key":"ref_39","first-page":"3891","article-title":"Regulation of miRNAs on c-met protein expression in ovarian cancer and its implication","volume":"28","author":"Liu","year":"2024","journal-title":"Eur. Rev. Med. Pharmacol. Sci."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"553","DOI":"10.1095\/biolreprod59.3.553","article-title":"Developmental and hormonal regulation of hepatocyte growth factor expression and action in the bovine ovarian follicle","volume":"59","author":"Parrott","year":"1998","journal-title":"Biol. Reprod."}],"container-title":["Animals"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2076-2615\/15\/21\/3077\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,25]],"date-time":"2025-10-25T04:26:04Z","timestamp":1761366364000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2076-2615\/15\/21\/3077"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,10,23]]},"references-count":40,"journal-issue":{"issue":"21","published-online":{"date-parts":[[2025,11]]}},"alternative-id":["ani15213077"],"URL":"https:\/\/doi.org\/10.3390\/ani15213077","relation":{},"ISSN":["2076-2615"],"issn-type":[{"type":"electronic","value":"2076-2615"}],"subject":[],"published":{"date-parts":[[2025,10,23]]}}}