基于高通量测序的梅花鹿卵巢衰老的转录组分析

doi:10.16431/j.cnki.1671-7236.2023.08.013

Abstract

Abstract: 【Objective】 This study was aimed to obtain the transcriptome information of ovary tissues in young and old sika deer,and provide a basis for further study the potential mechanism of ovarian aging in sika deer.【Method】 The ovaries of 4 years young sika deer and 10 years old sika deer were used as test samples.The samples were processed with paraffin sectioning,hematoxylin-eosin staining and follicle counting.RNA was extracted from the ovarian samples using the Trizol method,and a transcriptome library was constructed.The Illumina HiSeq^TM 2000 sequencing platform was used to sequence,the sequencing results were analyzed for differentially expressed genes,GO function,KEGG pathway and protein interaction network analysis.Selected differentially expressed genes were further analyzed using Real-time quantitative PCR.The expression of trimethylation of lysine 4 on histone H3 protein subunit (H3K4me3) protein in ovaries of young and old sika deer were detected by Western blotting analysis using RIPA lysis solution.【Result】 The histological analysis results showed that the number of ovarian follicles in old sika deer was significantly lower than that in young sika deer (P＜0.05),and the number of atresia follicles in old sika deer was extremely significantly higher than that of young sika deer (P＜0.01).Transcriptome analysis showed that there were 1 477 differentially expressed genes of ovaries between old and young sika deer,of which 503 genes were upregulated and 974 genes were downregulated.GO function and KEGG pathway enrichment analysis showed that the immune system process,transcription,DNA damage repair,inflammatory response,apoptosis and other biological events were related to ovarian aging.Real-time quantitative PCR results showed that there were 17 genes with significant or extremely significant expression of ovaries between old and young sika deer (P＜0.05 or P＜0.01),indicating that RNA-Seq results were reliable.Western blotting analysis results showed that the expression of H3K4me3 protein in ovaries was significantly different between old and young sika deer (P＜0.05).【Conclusion】 The number of follicles in ovaries of young and old sika deer showed a significant difference.The PI3K-Akt signaling pathway had been identified to affect the efficiency of DNA damage repair and participate in ovarian aging of sika deer via RNA-Seq.The high expression of H3K4me3 in ovaries of old sika deer was related to transcription activation.

Key words: sika deer; ovarian aging; RNA-Seq; H3K4me3

CLC Number:

S825

LIN Lefeng, DIAO Yunfei, FAN Bingfeng, LIU Lixiang, SHAO Jing, ZHAO Xiangyuan, XU Baozeng. Transcriptome Analysis of Aging Sika Deer (Cervus nippon) Ovaries Based on High-throughput Sequencing[J]. China Animal Husbandry & Veterinary Medicine, 2023, 50(8): 3157-3170.

References

[1] WANG S,ZHENG Y,LI J,et al.Single-cell transcriptomic atlas of primate ovarian aging[J].Cell,2020,180(3):585-600.
[2] RUSS J E,HAYWOOD M E,LANE S L,et al.Spatially resolved transcriptomic profiling of ovarian aging in mice[J].iScience,2022,25(8):104819.
[3] TE VELDE E R,PEARSON P L.The variability of female reproductive ageing[J].Human Reproduction Update,2002,8(2):141-154.
[4] MAY-PANLOUP P,BOUCRET L,CHAO DE LA BARCA J M,et al.Ovarian ageing:The role of mitochondria in oocytes and follicles[J].Human Reproduction Update,2016,22(6):725-743.
[5] ZHANG J,CHEN Q,DU D,et al.Can ovarian aging be delayed by pharmacological strategies[J].Aging,2019,11(2):817-832.
[6] LIN W,TITUS S,MOY F,et al.Ovarian aging in women with BRCA germline mutations[J].The Journal of Clinical Endocrinology&Metabolism,2017,102(10):3839-3847.
[7] TITUS S,LI F,STOBEZKI R,et al.Impairment of brca1-related DNA double-strand break repair leads to ovarian aging in mice and humans[J].Science Translational Medicine,2013,5(172):172ra121.
[8] WU Y W,LI S,ZHENG W,et al.Dynamic mRNA degradome analyses indicate a role of histone H3K4 trimethylation in association with meiosis-coupled mRNA decay in oocyte aging[J].Nature Communications,2022,13(1):3191.
[9] SMITH J T,CLARKE I J.Seasonal breeding as a neuroendocrine model for puberty in sheep[J].Molecular and Cellular Endocrinology,2010,324(1):102-109.
[10] ALIMADADI A,MUNROE P B,JOE B,et al.Meta-analysis of dilated cardiomyopathy using cardiac RNA-Seq transcriptomic datasets[J].Genes,2020,11(1):60.
[11] GRABHERR M G,HAAS B J,YASSOUR M,et al.Full-length transcriptome assembly from RNA-Seq data without a reference genome[J].Nature Biotechnology,2011,29(7):644-652.
[12] LAN D,XIONG X,WEI Y,et al.RNA-Seq analysis of yak ovary:Improving yak gene structure information and mining reproduction-related genes[J].Science China Life Sciences,2014,57(9):925-935.
[13] WESTERMANN A J,VOGEL J.Cross-species RNA-Seq for deciphering host-microbe interactions[J].Nature Reviews Genetics,2021,22(6):361-378.
[14] HORTA F,RAVICHANDRAN A,CATT S,et al.Ageing and ovarian stimulation modulate the relative levels of transcript abundance of oocyte DNA repair genes during the germinal vesicle-metaphase Ⅱ transition in mice[J].Journal of Assisted Reproduction and Genetics,2021,38(1):55-69.
[15] TIAN C,HE J,AN Y,et al.Bone marrow mesenchymal stem cells derived from juvenile macaques reversed ovarian ageing in elderly macaques[J].Stem Cell Research&Therapy,2021,12(1):460.
[16] JIA B,LIU Y,LI Q,et al.Altered miRNA and mRNA expression in sika deer skeletal muscle with age[J].Genes,2020,11(2):172.
[17] WALKER M L,ANDERSON D C,HERNDON J G,et al.Ovarian aging in squirrel monkeys (Saimiri sciureus)[J].Reproduction,2009,138(5):793-799.
[18] JOHNSON J,CANNING J,KANEKO T,et al.Germline stem cells and follicular renewal in the postnatal mammalian ovary[J].Nature,2004,428(6979):145-150.
[19] GAO Z,MOORJANI P,SASANI T A,et al.Overlooked roles of DNA damage and maternal age in generating human germline mutations[J].Proceedings of the National Academy of Sciences of the United States of America,2019,116(19):9491-9500.
[20] HAN R,HAN L,WANG S,et al.Whole transcriptome analysis of mesenchyme tissue in sika deer antler revealed the ceRNAs regulatory network associated with antler development[J].Frontiers in Genetics,2020,10(1):1403.
[21] LIU Y,WANG R,SU L,et al.Integrative proteomic and phosphoproteomic analyses revealed complex mechanisms underlying reproductive diapause in bombus terrestris queens[J].Insects,2022,13(10):862.
[22] CHIN C H,CHEN S H,WU H H,et al.Cytohubba:Identifying hub objects and sub-networks from complex interactome[J].BMC Systems Biology,2014,8(Suppl 4):S11.
[23] LIU X C,REN Q,GUO J,et al.First detection and molecular identification of entamoeba bovis in farm-raised sika deer from Anhui province,China[J].Acta Parasitologica,2022,67(4):1782-1787.
[24] LIVAK K J,SCHMITTGEN T D.Analysis of relative gene expression data using Real-time quantitative PCR and the 2^-ΔΔCt method[J].Methods,2001,25(4):402-408.
[25] ZHU J,YANG Q,LI H,et al.Sirt3 deficiency accelerates ovarian senescence without affecting spermatogenesis in aging mice[J].Free Radical Biology and Medicine,2022,193(1):511-525.
[26] LI Q,GENG X,ZHENG W,et al.Current understanding of ovarian aging[J].Science China Life Sciences,2012,55(8):659-669.
[27] JIN Y,YANG M,GAO C,et al.Fbxo30 regulates chromosome segregation of oocyte meiosis[J].Cellular and Molecular Life Sciences,2019,76(11):2217-2229.
[28] ADHIKARI D,LIU K.Molecular mechanisms underlying the activation of mammalian primordial follicles[J].Endocrine Reviews,2009,30(5):438-464.
[29] REDDY P,LIU L,ADHIKARI D,et al.Oocyte-specific deletion of PTEN causes premature activation of the primordial follicle pool[J]. Science,2008,319(5863):611-613.
[30] MAIDARTI M,ANDERSON R A,TELFER E E.Crosstalk between PTEN/PI3K/Akt signalling and DNA damage in the oocyte:Implications for primordial follicle activation,oocyte quality and ageing[J].Cells,2020,9(1):200.
[31] MA Y,SHI Y,WANG Y,et al.Role of endoplasmic reticulum stress in nano-selenium alleviating prehierarchical follicular atresia induced by mercury in laying hens[J].Biological Trace Element Research,2022,200(12):5205-5217.
[32] MARTIN N,BERNARD D.Calcium signaling and cellular senescence[J].Cell Calcium,2018,70(1):16-23.
[33] MOHAMMED R N,KHOSRAVI M,RAHMAN H S,et al.Anastasis:Cell recovery mechanisms and potential role in cancer[J].Cell Communication and Signaling,2022,20(1):81.
[34] STAMPERNA K,GIANNOULIS T,NANAS I,et al.Short term temperature elevation during IVM affects embryo yield and alters gene expression pattern in oocytes,cumulus cells and blastocysts in cattle[J].Theriogenology,2020,156(1):36-45.
[35] SINGH R,KOLVRAA S,RATTAN S I S.Genetics of human longevity with emphasis on the relevance of HSP70 as candidate genes[J].Frontiers in Bioscience-Landmark,2007,12(7):4504-4513.
[36] HAN Y,KANG Y,YU J,et al.Increase of HSPA1A and HSPA1B genes in the resting B cells of Sirt1 knockout mice[J].Molecular Biology Reports,2019,46(4):4225-4234.
[37] LIU J,LI L,CHEN X,et al.Effects of heat stress on body temperature,milk production,and reproduction in dairy cows:A novel idea for monitoring and evaluation of heat stress-A review[J].Asian-Australasian Journal of Animal Sciences,2019,32(9):1332-1339.
[38] SOTTILE M L,NADIN S B.Heat shock proteins and DNA repair mechanisms:An updated overview[J].Cell Stress and Chaperones,2018,23(3):303-315.
[39] CARUSO BAVISOTTO C,ALBERTI G,VITALE A M,et al.Hsp60 post-translational modifications:Functional and pathological consequences[J].Frontiers in Molecular Biosciences,2020,7(1):95.
[40] ZININGA T,RAMATSUI L,SHONHAI A.Heat shock proteins as immunomodulants[J].Molecules,2018,23(11):2846.
[41] TANAKA T,NARAZAKI M,MASUDA K,et al.Regulation of IL-6 in immunity and diseases[J].Regulation of Cytokine Gene Expression in Immunity and Diseases,2016,941(1):79-88.
[42] BABAYEV E,DUNCAN F E.Age-associated changes in cumulus cells and follicular fluid:The local oocyte microenvironment as a determinant of gamete quality[J].Biology of Reproduction,2022,106(2):351-365.
[43] LLIBEROS C,LIEW S H,ZAREIE P,et al.Evaluation of inflammation and follicle depletion during ovarian ageing in mice[J].Scientific Reports,2021,11(1):278.
[44] ITSUMI M,INOUE S,ELIA A J,et al.Idh1 protects murine hepatocytes from endotoxin-induced oxidative stress by regulating the intracellular NADP⁺/NADPH ratio[J].Cell Death&Differentiation,2015,22(11):1837-1845.
[45] KAWAI T,AKIRA S.TLR signaling[J].Seminars in Immunology,2007,19(1):24-32.
[46] TURNER M L,CRONIN J G,HEALEY G D,et al.Epithelial and stromal cells of bovine endometrium have roles in innate immunity and initiate inflammatory responses to bacterial lipopeptides in vitro via Toll-like receptors TLR2,TLR1,and TLR6[J].Endocrinology,2014,155(4):1453-1465.
[47] KRAWCZYK C M,HOLOWKA T,SUN J,et al.Toll-like receptor-induced changes in glycolytic metabolism regulate dendritic cell activation[J].Blood,Journal of the American Society of Hematology,2010,115(23):4742-4749.
[48] HOOD V L,BERGER R,FREEDMAN R,et al.Transcription of PIK3CD in human brain and schizophrenia:Regulation by proinflammatory cytokines[J].Human Molecular Genetics,2019,28(19):3188-3198.
[49] TANGYE S G,BIER J,LAU A,et al.Immune dysregulation and disease pathogenesis due to activating mutations in PIK3CD-The goldilocks'effect[J].Journal of Clinical Immunology,2019,39(2):148-158.
[50] TURAN V,OKTAY K.BRCA-related ATM-mediated DNA double-strand break repair and ovarian aging[J].Human Reproduction Update,2019,26(1):43-57.
[51] PORTER JR G A,BEUTNER G.Cyclophilin D,somehow a master regulator of mitochondrial function[J].Biomolecules,2018,8(4):176.
[52] HALESTRAP A P.What is the mitochondrial permeability transition pore?[J].Journal of Molecular and Cellular Cardiology,2009,46(6):821-831.
[53] YE F,LI X,LIU Y,et al.CypD deficiency confers neuroprotection against mitochondrial abnormality caused by lead in SH-SY5Y cell[J].Toxicology Letters,2020,323(1):25-34.
[54] AMANAKIS G,MURPHY E.Cyclophilin D:An integrator of mitochondrial function[J].Frontiers in Physiology,2020,11(1):595.
[55] DEMAIN L A,CONWAY G S,NEWMAN W G.Genetics of mitochondrial dysfunction and infertility[J].Clinical Genetics,2017,91(2):199-207.
[56] DE RUIJTER A J,VAN GENNIP A H,CARON H N,et al.Histone deacetylases (HDACs):Characterization of the classical HDAC family[J].Biochemical Journal,2003,370(3):737-749.
[57] ECKSCHLAGER T,PLCH J,STIBOROVA M,et al.Histone deacetylase inhibitors as anticancer drugs[J].International Journal of Molecular Sciences,2017,18(7):1414.
[58] MA P,SCHULTZ R M.Histone deacetylase 2(HDAC2) regulates chromosome segregation and kinetochore function via H4K16 deacetylation during oocyte maturation in mouse[J].PLoS Genetics,2013,9(3):e1003377.
[59] EICHENLAUB-RITTER U.Oocyte ageing and its cellular basis[J].International Journal of Developmental Biology,2013,56(10-12):841-852.
[60] LASKOWSKI D,BÅGE R,HUMBLOT P,et al.Insulin during in vitro oocyte maturation has an impact on development,mitochondria,and cytoskeleton in bovine day 8 blastocysts[J].Theriogenology,2017,101(1):15-25.
[61] HUA K,WANG L,SUN J,et al.Impairment of Polβ-related DNA base-excision repair leads to ovarian aging in mice[J].Aging (Albany NY),2020,12(24):25207-25228.
[62] GAO Y,LAN Y,OVITT C E,et al.Functional equivalence of the zinc finger transcription factors Osr1 and Osr2 in mouse development[J].Developmental Biology,2009,328(2):200-209.
[63] CHANG H,ZHANG X,LI B,et al.MAGI2-AS3 suppresses MYC signaling to inhibit cell proliferation and migration in ovarian cancer through targeting miR-525-5p/MXD1 axis[J].Cancer Medicine,2020,9(17):6377-6386.
[64] SEN P,SHAH P P,NATIVIO R,et al.Epigenetic mechanisms of longevity and aging[J].Cell,2016,166(4):822-839.
[65] PEÑA P V,HOM R A,HUNG T,et al.Histone H3K4me3 binding is required for the DNA repair and apoptotic activities of ING1 tumor suppressor[J].Journal of Molecular Biology,2008,380(2):303-312.
[66] RIZZARDI L F,DORN E S,STRAHL B D,et al.DNA replication origin function is promoted by H3K4 di-methylation in Saccharomyces cerevisiae[J].Genetics,2012,192(2):371-384.
[67] SOMMERMEYER V,BÉNEUT C,CHAPLAIS E,et al.Spp1,a member of the Set1 complex,promotes meiotic DSB formation in promoters by tethering histone H3K4 methylation sites to chromosome axes[J].Molecular Cell,2013,49(1):43-54.
[68] ACQUAVIVA L,DROGAT J,DEHÉ P M,et al.Spp1 at the crossroads of H3K4me3 regulation and meiotic recombination[J].Epigenetics,2013,8(4):355-360.

Transcriptome Analysis of Aging Sika Deer (Cervus nippon) Ovaries Based on High-throughput Sequencing

PDF (PC)

Knowledge

Abstract

Cite this article

share this article

References

Related Articles 15

Recommended Articles

Metrics

Comments

[1]	JIA Chunyan, SUN Yanyong, BAO Yonghong, ZHANG Wenguang, DU Chenguang. Alternative Splicing Analysis of mRNA and lncRNA Induced by Exogenous Melatonin for Regulating Cashmere Growth Based on Transcriptomics [J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(7): 3165-3177.
[2]	REN Hao, ZHU Yixuan, CHAO Tingting, WANG Xiaoyi, LU Shaoxiong, YANG Yongli, CHEN Qiang, LI Mingli. Identification and Functional Prediction of lncRNA in Longissimus Dorsi Muscle of Saba Pigs with Different Growth Rates [J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(6): 2494-2505.
[3]	LI Zhiyi, LI Jie, CHEN Chuwen, NONG Yi, WANG Jiayan, WANG Zi, WU Jinbo, LI Zhixiong. Analysis and Identification of miRNA in Leg Muscle Tissue of Tibetan Chicken Embryos at Different Developmental Stages [J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(4): 1681-1693.
[4]	SHEN Yangyang, AN Zhenjiang, GAO Linna, XIA Shuwen, CHEN Kunlin, DING Qiang, GAO Yundong, XUE Jinguo, ZHONG Jifeng, WANG Huili. Comparative Analysis of Muscle Transcriptome of Xuzhou Cattle with Different Tenderness [J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(2): 522-533.
[5]	WANG Qing, MA Yukai, HAN Ruobing, XING Haihua, SUN Zihui, LI Heping. Cloning，Bioinformatics Analysis and Expression Analysis of CSRP3 Gene in Muscle Tissues of Sika Deer [J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(2): 740-748.
[6]	HUANG Yao, YAO Naiquan, YUAN Weitao, MAO Aipeng, ZHANG Ting, XU Chao. Isolation and Identification of Candida rugosa DZ018 from Deer and Its Probiotic Properties in vitro [J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(1): 389-399.
[7]	XIA Minglong, XIAO Yintao, ZHENG Saizhen, TAN Bie, YIN Yulong, CHEN Jiashun, YIN Jie. Analysis of Differentially Expressed Genes and Regulatory Pathways in Intramuscular Fat Deposition of Ningxiang Pigs at Different Developmental Stages [J]. China Animal Husbandry & Veterinary Medicine, 2024, 51(9): 3703-3714.
[8]	MA Sijia, ZHAO Zhengwei, WANG Jin, MA Lina. Discovering Key Genes in Hair Follicle Development of Tan Sheep at Different Days of Age Based on Transcriptome Data [J]. China Animal Husbandry & Veterinary Medicine, 2024, 51(9): 3726-3738.
[9]	SUN Huimin, FAN Bingfeng, ZHANG Xulin, SHAO Jing, LIU Lixiang, XU Baozeng. Transcriptomic Analysis of Blood in Early Pregnancy at Different Time Points in Sika Deer [J]. China Animal Husbandry & Veterinary Medicine, 2024, 51(9): 3892-3908.
[10]	JI Hongjin, YE Qiannan, LI Wenlong, LI Zhipeng, FENG Xia, MI Siyuan, SHI Yuanjun, ZHANG Jinning, GUO Jian, YU Ying. Screening of Key Homologous Genes for Bovine Fertility Based on High Folic Acid Mouse Testicular Transcriptome [J]. China Animal Husbandry & Veterinary Medicine, 2024, 51(7): 2751-2765.
[11]	MENG Chen, ZENG Yaqi, WANG Jianwen, YAO Xinkui, LUO Penghui, XIE Xiaoyu, LI Pengcheng, LIU Xiaoxiao, WANG Chuankun, MENG Jun. Screening of Genes Related to Lactation in Kazakh Horses Based on Transcriptomic [J]. China Animal Husbandry & Veterinary Medicine, 2024, 51(12): 5392-5404.
[12]	MA Sufang, YANG Wenqing, ZHANG Bingbing, GUO Ruonan, XU Yingxin, ZHANG Linlin, GUO Yiwen, HU Debao, GUO Hong, DING Xiangbin, LI Xin. Joint Analysis of Transcriptome and Translatome During the Development of Bovine Skeletal Muscle Satellite Cells [J]. China Animal Husbandry & Veterinary Medicine, 2024, 51(11): 4665-4677.
[13]	ZHANG Ting, HUANG Yao, YUAN Weitao, GUO Xiaolan, WANG Kaiying, XU Chao. Effects of Vitamin D₃ on Growth Performance，Apparent Digestibility of Nutrients and Serum Biochemical Indexes of 10-12 Months Old Sika Deer (Cervus nippon) [J]. China Animal Husbandry & Veterinary Medicine, 2024, 51(10): 4336-4342.
[14]	CHEN Xu, DONG Yimeng, XING Xiumei, YANG Sukun, LIU Xin. Study on the Role of SCF Gene in Pigment Synthesis in Sika Deer [J]. China Animal Husbandry & Veterinary Medicine, 2023, 50(9): 3663-3670.
[15]	SUN Yanyong, MA Fengying, GUO Lili, LIU Zaixia, LIU Bin, ZHANG Wenguang, LIU Yongbin. Advances on the Application of Animal Blood Transcriptome [J]. China Animal Husbandry & Veterinary Medicine, 2023, 50(2): 460-468.