DOI QR코드

DOI QR Code

Alterations of mRNA and lncRNA profiles associated with the extracellular matrix and spermatogenesis in goats

  • Chen, Haolin (College of Animal Science and Technology, Gansu Agricultural University) ;
  • Miao Xiaomeng (Institute of Animal Husbandry and Veterinary, Guizhou Academy of Agricultural Sciences) ;
  • Xu, Jinge (Institute of Animal Husbandry and Veterinary, Guizhou Academy of Agricultural Sciences) ;
  • Pu, Ling (Institute of Animal Husbandry and Veterinary, Guizhou Academy of Agricultural Sciences) ;
  • Li, Liang (Institute of Animal Husbandry and Veterinary, Guizhou Academy of Agricultural Sciences) ;
  • Han, Yong (Institute of Animal Husbandry and Veterinary, Guizhou Academy of Agricultural Sciences) ;
  • Mao, Fengxian (Guizhou Province Livestock and Poultry Genetic Resources Management Station) ;
  • Ma, Youji (College of Animal Science and Technology, Gansu Agricultural University)
  • Received : 2021.06.03
  • Accepted : 2021.08.16
  • Published : 2022.04.01

Abstract

Objective: Spermatozoa are produced within the seminiferous tubules after sexual maturity. The expression levels of mRNAs and lncRNAs in testicular tissues are different at each stage of testicular development and are closely related to formation of the extracellular matrix (ECM) and spermatogenesis. Therefore, we set out to study the expression of lncRNAs and mRNAs during the different developmental stages of the goat testis. Methods: We constructed 12 RNA libraries using testicular tissues from goats aged 3, 6, and 12 months, and studied the functions of mRNAs and lncRNAs using the gene ontogeny (GO) and Kyoto encyclopedia of genes and genomes (KEGG) databases. Relationships between differentially expressed genes (DEGs) were analyzed by lncRNA-mRNA co-expression network and protein-protein interaction network (PPI). Finally, the protein expression levels of matrix metalloproteinase 2 (MMP2), insulin-like growth factor 2 (IGF2), and insulin-like growth factor-binding protein 6 (IGFBP6) were detected by western blotting. Results: We found 23, 8, and 135 differentially expressed lncRNAs and 161, 12, and 665 differentially expressed mRNAs that were identified between 3 vs 6, 6 vs 12, and 3 vs 12 months, respectively. GO, KEGG, and PPI analyses showed that the differential genes were mainly related to the ECM. Moreover, MMP2 was a hub gene and co-expressed with the lncRNA TCONS-0002139 and TCONS-00093342. The results of quantitative reverse-transcription polymerase chain reaction verification were consistent with those of RNA-seq sequencing. The expression trends of MMP2, IGF2, and IGFBP6 protein were the same as that of mRNA, which all decreased with age. IGF2 and MMP2 were significantly different in the 3 vs 6-month-old group (p<0.05). Conclusion: These results improve our understanding of the molecular mechanisms involved in sexual maturation of the goat testis.

Keywords

Acknowledgement

This study was supported by the Science and Technology Program of Guizhou Province ([2020]4009(002)); China Agriculture Research System of MOF and MARA; Team Cultivation Project of Guizhou Animal Husbandry and Veterinary Research Institute (No. 03[2018]).

References

  1. Zhu Z, Li C, Yang S, et al. Dynamics of the transcriptome during human spermatogenesis: predicting the potential key genes regulating male gametes generation. Sci Rep 2016;6:19069. https://doi.org/10.1038/srep19069
  2. Li C, Shen C, Shang X, et al. Two novel testis-specific long noncoding RNAs produced by 1700121C10Rik are dispensable for male fertility in mice. J Reprod Dev 2020;66:57-65. https://doi.org/10.1262/jrd.2019-104
  3. Zeng F, Chen Y, Guo C, et al. Analysis of differentially abundant proteins related to boar fertility in seminal plasma using iTRAQ-based quantitative proteomics. J Proteomics 2021;236:104120. https://doi.org/10.1016/j.jprot.2021.104120
  4. Cole JB, Durr JW, Nicolazzi EL. Invited review: The future of selection decisions and breeding programs: What are we breeding for, and who decides? J Dairy Sci 2021;104:5111-24. https://doi.org/10.3168/jds.2020-19777
  5. Mayerhofer A, Walenta L, Mayer C, Eubler K, Welter H. Human testicular peritubular cells, mast cells and testicular inflammation. Andrologia 2018;50:e13055. https://doi.org/10.1111/and.13055
  6. Nosrati R, Kheirouri S, Ghodsi R, Ojaghi H. The effects of zinc treatment on matrix metalloproteinases: a systematic review. J Trace Elem Med Biol 2019;56:107-15. https://doi.org/10.1016/j.jtemb.2019.08.001
  7. Matuszczak E, Komarowska M, Sankiewicz A, et al. Plasma concentration of MMP-1 and MMP-2 in boys with cryptorchidism and its lack of correlation with INSL3 and inhibin B. Scand J Clin Lab Invest 2019;79:412-8. https://doi.org/10.1080/00365513.2019.1637534
  8. Wathes D, Cheng Z, Fenwick MA, Fitzpatrick R, Patton J. Influence of energy balance on the somatotrophic axis and matrix metalloproteinase expression in the endometrium of the postpartum dairy cow. Reproduction (Cambridge, England) 2011;141:269-81. https://doi.org/10.1530/rep-10-0177
  9. Voit-Ostricki L, Lovas S, Watts CR. Conformation and domain movement analysis of human matrix metalloproteinase-2: role of associated Zn2+ and Ca2+ ions. Int J Mol Sci 2019;20:4194. https://doi.org/10.3390/ijms20174194
  10. Clemmons DR. Role of IGF-binding proteins in regulating IGF responses to changes in metabolism. J Mol Endocrinol 2018;61:T139-T69. https://doi.org/10.1530/jme-18-0016
  11. Batista P, Chang H. Long noncoding RNAs: cellular address codes in development and disease. Cell 2013;152:1298-307. https://doi.org/10.1016/j.cell.2013.02.012
  12. Zhang C, Gao L, Xu EY. LncRNA, a new component of expanding RNA-protein regulatory network important for animal sperm development. Semin Cell Dev Biol 2016;59:110-7. https://doi.org/10.1016/j.semcdb.2016.06.013
  13. Liu K, Li T, Ton H, Mao X, Chen Y. Advances of long noncoding RNAs-mediated regulation in reproduction. Chin Med J 2018;131:226-34. https://doi.org/10.4103/0366-6999.222337
  14. Liu Y, Sun Y, Li Y, et al. Analyses of long non-coding RNA and mRNA profiling using RNA sequencing in chicken testis with extreme sperm motility. Sci Rep 2017;7:9055. https://doi.org/10.1038/s41598-017-08738-9
  15. Zhang Y, Yang H, Han L, et al. Long noncoding RNA expression profile changes associated with dietary energy in the sheep testis during sexual maturation. Sci Rep 2017;7:5180. https://doi.org/10.1038/s41598-017-05443-5
  16. Weng B, Ran M, Chen B, He C, Dong L, Peng F. Genome-wide analysis of long non-coding RNAs and their role in postnatal porcine testis development. Genomics 2017;109:446-56. https://doi.org/10.1016/j.ygeno.2017.07.001
  17. Langmead B, Salzberg SL. Fast gapped-read alignment with Bowtie 2. Nat Methods 2012;9:357-9. https://doi.org/10.1038/nmeth.1923
  18. Kim D, Pertea G, Trapnell C, Pimentel H, Kelley R, Salzberg SL. TopHat2: accurate alignment of transcriptomes in the presence of insertions, deletions and gene fusions. Genome Biol 2013;14:R36. https://doi.org/10.1186/gb-2013-14-4-r36
  19. Sun L, Luo H, Bu D, et al. Utilizing sequence intrinsic composition to classify protein-coding and long non-coding transcripts. Nucleic Acids Res 2013;41:e166. https://doi.org/10.1093/nar/gkt646
  20. Kong L, Zhang Y, Ye Z, et al. CPC: assess the protein-coding potential of transcripts using sequence features and support vector machine. Nucleic Acids Res 2007;35:W345-9. https://doi.org/10.1093/nar/gkm391
  21. El-Gebali S, Mistry J, Bateman A, et al. The Pfam protein families database in 2019. Nucleic Acids Res 2019;47:D427-32. https://doi.org/10.1093/nar/gky995
  22. Wang L, Park H, Dasari S, Wang S, Kocher JP, Li W. CPAT: Coding-potential assessment tool using an alignment-free logistic regression model. Nucleic Acids Res 2013;41:e74. https://doi.org/10.1093/nar/gkt006
  23. Trapnell C, Williams B, Pertea G, et al. Transcript assembly and quantification by RNA-Seq reveals unannotated transcripts and isoform switching during cell differentiation. Nat Biotechnol 2010;28:511-5. https://doi.org/10.1038/nbt.1621
  24. Young M, Wakefield M, Smyth G, Oshlack A. Gene ontology analysis for RNA-seq: accounting for selection bias. Genome Biol 2010;11:R14. https://doi.org/10.1186/gb-2010-11-2-r14
  25. Xie C, Mao X, Huang J, et al. KOBAS 2.0: a web server for annotation and identification of enriched pathways and diseases. Nucleic Acids Res 2011;39:W316-22. https://doi.org/10.1093/nar/gkr483
  26. Langfelder P, Horvath S. WGCNA: an R package for weighted correlation network analysis. BMC Bioinformatics 2008;9:559. https://doi.org/10.1186/1471-2105-9-559
  27. Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PCR and the 2 ΔΔ C T method. Methods 2001;25:402-8. https://doi.org/10.1006/meth.2001.1262
  28. Veneziano D, Nigita G, Ferro A. Computational approaches for the analysis of ncRNA through deep sequencing techniques. Front Bioeng Biotechnol 2015;3:77. https://doi.org/10.3389/fbioe.2015.00077
  29. Bao J, Wu J, Schuster A, Hennig G, Yan W. Expression profiling reveals developmentally regulated lncRNA repertoire in the mouse male germline. Biol Reprod 2013;89:107. https://doi.org/10.1095/biolreprod.113.113308
  30. Joshi M, Rajender S. Long non-coding RNAs (lncRNAs) in spermatogenesis and male infertility. Reprod Biol Endocrinol 2020;18:103. https://doi.org/10.1186/s12958-020-00660-6
  31. Yang H, Wang F, Li F, et al. Comprehensive analysis of long noncoding RNA and mRNA expression patterns in sheep testicular maturation. Biol Reprod 2018;99:650-61. https://doi.org/10.1093/biolre/ioy088
  32. Gao Y, Li S, Lai Z, et al. Analysis of long non-coding RNA and mRNA expression profiling in immature and mature bovine (Bos taurus) testes. Front Genet 2019;10:646. https://doi.org/10.3389/fgene.2019.00646
  33. Bo D, Jiang X, Liu G, Hu R, Chong Y. RNA-Seq implies divergent regulation patterns of lincRNA on spermatogenesis and testis growth in goats. Animals 2021;11:625. https://doi.org/10.3390/ani11030625
  34. Bai M, Sun L, Zhao J, et al. Histological analysis and identification of spermatogenesis-related genes in 2-, 6-, and 12- month-old sheep testes. Naturwissenschaften 2017;104:84. https://doi.org/10.1007/s00114-017-1505-1
  35. Mayerhofer A. Human testicular peritubular cells: more than meets the eye. Reproduction (Cambridge, England) 2013; 145:R107-16. https://doi.org/10.1530/rep-12-0497
  36. Siu MKY, Cheng CY. Extracellular matrix: recent advances on its role in junction dynamics in the seminiferous epithelium during spermatogenesis. Biol Reprod 2004;71:375-91. https://doi.org/10.1095/biolreprod.104.028225
  37. Tan KS, Kulkeaw K, Nakanishi Y, Sugiyama D. Expression of cytokine and extracellular matrix mRNAs in fetal hepatic stellate cells. Genes Cells 2017;22:836-44. https://doi.org/10.1111/gtc.12517
  38. van Dam S, Vosa U, van der Graaf A, Franke L, de Magalhaes JP. Gene co-expression analysis for functional classification and gene-disease predictions. Brief Bioinform 2018;19:575-92. https://doi.org/10.1093/bib/bbw139
  39. Huang H, Zhao W, Tang Z, et al. Characterization of porcine MMP-2 and its association with immune traits. Gene 2009;435:63-71. https://doi.org/10.1016/j.gene.2009.01.002
  40. Schrade A, Kyronlahti A, Akinrinade O, et al. GATA4 regulates blood-testis barrier function and lactate metabolism in mouse sertoli cells. Endocrinology 2016;157:2416-31. https://doi.org/10.1210/en.2015-1927
  41. McMillen P, Holley SA. Integration of cell-cell and cell-ECM adhesion in vertebrate morphogenesis. Curr Opin Cell Biol 2015;36:48-53. https://doi.org/10.1016/j.ceb.2015.07.002
  42. Safian D, Morais R, Bogerd J, Schulz R. Igf binding proteins protect undifferentiated spermatogonia in the zebrafish testis against excessive differentiation. Endocrinology 2016;157:4423-33. https://doi.org/10.1210/en.2016-1315
  43. Su YY, Nishimoto T, Hoffman S, et al. Insulin-like growth factor binding protein-4 exerts antifibrotic activity by reducing levels of connective tissue growth factor and the C-X-C chemokine receptor 4. FASEB bioAdvances 2019;1:167-79. https://doi.org/10.1096/fba.2018-00015
  44. Yao Q, Chen T. LINC01128 regulates the development of osteosarcoma by sponging miR-299-3p to mediate MMP2 expression and activating Wnt/β-catenin signalling pathway. J Cell Mol Med 2020;24:14293-305. https://doi.org/10.1111/jcmm.16046
  45. Chen H, Fok K, Yu S, et al. CD147 is required for matrix metalloproteinases-2 production and germ cell migration during spermatogenesis. Mol Hum Reprod 2011;17:405-14. https://doi.org/10.1093/molehr/gar013
  46. Finkielstain GP, Forcinito P, Lui JCK, et al. An extensive genetic program occurring during postnatal growth in multiple tissues. Endocrinology 2009;150:1791-800. https://doi.org/10.1210/en.2008-0868