Expression of enzymes involved in polyunsaturated fatty acid synthesis in the stallion testis and epididymis
Camille Gautier
A , Dragos Scarlet B , Reinhard Ertl C , Ingrid Walter D , Manuela Wulf E , Christina Nagel E , Jörg Aurich B and Christine Aurich A F
+ Author Affiliations
- Author Affiliations
A Artificial Insemination and Embryo Transfer, Department for Small Animals and Horses, Vetmeduni Vienna, Veterinärplatz 1, 1210 Vienna, Austria.
B Gynecology, Obstetrics and Andrology, Department for Small Animals and Horses, Vetmeduni Vienna, Veterinärplatz 1, 1210 Vienna, Austria.
C Vetcore Facility for Research, Vetmeduni Vienna, Veterinärplatz 1, 1210 Vienna, Austria.
D Institute of Pathology, Department of Pathobiology, Vetmeduni Vienna, Veterinärplatz 1, 1210 Vienna, Austria.
E Graf Lehndorff Institute, Vetmeduni Vienna, Hauptgestüt 10, 16845 Neustadt (Dosse), Germany.
F Corresponding author. Email: camillemarie.gautier@vetmeduni.ac.at
Reproduction, Fertility and Development 32(9) 851-861 https://doi.org/10.1071/RD19342
Submitted: 2 September 2019 Accepted: 17 March 2020 Published: 2 June 2020
Abstract
The aim of the present study was to characterise key enzymes involved in polyunsaturated fatty acid (PUFA) synthesis in the testis and epididymis collected from 2-year-old healthy warmblood stallions (n = 10). The mRNA expression of fatty acid synthase, the Δ9-, Δ6-, Δ5- and Δ4-desaturases and elongases 6, 5 and 2 (encoded by the fatty acid synthase (FASN), the stearoyl-CoA desaturase (SCD), the fatty acid desaturase 2 (FADS2), the fatty acid desaturase 1 (FADS1), the delta 4-desaturase, sphingolipid 1 (DEGS1), ELOVL fatty acid elongase 6 (ELOVL6), ELOVL fatty acid elongase 5 (ELOVL5), ELOVL fatty acid elongase 2 (ELOVL2) genes respectively) was determined in equine testis and epididymis. All enzymes were present in testicular tissue and along the epididymis, but mRNA expression differed among localisations. The protein localisation of FADS1, FADS2 and ELOVL5 was determined by immunohistochemistry. In the testes, FADS1 was expressed in the germinal cells and ELOVL5 was expressed in germinal and Leydig cells; FADS2 was not detected. In the epididymis, FADS1 and FADS2 were expressed in the principal and basal cells, whereas ELOVL5 was found only in the principal cells of the caput. All three enzymes were present in epididymal vesicles secreted by an apocrine mechanism. These results suggest active PUFA metabolism during spermatogenesis and epididymal sperm maturation in stallions.
Additional keyword: horse.
References
Argov, N., Sklan, D., Zeron, Y., and Roth, Z. (2007). Association between seasonal changes in fatty-acid composition, expression of VLDL receptor and bovine sperm quality. Theriogenology 67, 878–885.| Association between seasonal changes in fatty-acid composition, expression of VLDL receptor and bovine sperm quality.Crossref | GoogleScholarGoogle Scholar | 17157373PubMed |
Argov-Argaman, N., Mahgrefthe, K., Zeron, Y., and Roth, Z. (2013). Season-induced variation in lipid composition is associated with semen quality in Holstein bulls. Reproduction 145, 479–489.
| Season-induced variation in lipid composition is associated with semen quality in Holstein bulls.Crossref | GoogleScholarGoogle Scholar | 23630332PubMed |
Aumüller, G., and Seitz, J. (1990). Protein secretion and secretory processes in male accessory sex glands. Int. Rev. Cytol. 121, 127–231.
| Protein secretion and secretory processes in male accessory sex glands.Crossref | GoogleScholarGoogle Scholar | 2190945PubMed |
Aurich, C. (2016). Seasonal influences on cooled–shipped and frozen–thawed stallion semen. J. Equine Vet. Sci. 43, 1–5.
| Seasonal influences on cooled–shipped and frozen–thawed stallion semen.Crossref | GoogleScholarGoogle Scholar |
Aurich, C., Ortega Ferrusola, C., Peña Vega, F. J., Schrammel, N., Morcuende, D., and Aurich, J. (2018). Seasonal changes in the sperm fatty acid composition of Shetland pony stallions. Theriogenology 107, 149–153.
| Seasonal changes in the sperm fatty acid composition of Shetland pony stallions.Crossref | GoogleScholarGoogle Scholar | 29154176PubMed |
Bradford, M. M. (1976). A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72, 248–254.
| A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding.Crossref | GoogleScholarGoogle Scholar | 942051PubMed |
Gervasi, M. G., and Visconti, P. E. (2017). Molecular changes and signaling events occurring in sperm during epididymal maturation. Andrology 5, 204–218.
| Molecular changes and signaling events occurring in sperm during epididymal maturation.Crossref | GoogleScholarGoogle Scholar | 28297559PubMed |
Gottschalk, M., Sieme, H., Martinsson, G., and Distl, O. (2016). Analysis of breed effects on semen traits in light horse, warmblood, and draught horse breeds. Theriogenology 85, 1375–1381.
| Analysis of breed effects on semen traits in light horse, warmblood, and draught horse breeds.Crossref | GoogleScholarGoogle Scholar | 26893165PubMed |
Guillou, H., Zadravec, D., Martin, P. G. P., and Jacobsson, A. (2010). The key roles of elongases and desaturases in mammalian fatty acid metabolism: insights from transgenic mice. Prog. Lipid Res. 49, 186–199.
| The key roles of elongases and desaturases in mammalian fatty acid metabolism: insights from transgenic mice.Crossref | GoogleScholarGoogle Scholar | 20018209PubMed |
Hall, J. C., Hadley, J., and Doman, T. (1991). Correlation between changes in rat sperm membrane lipids, protein, and the membrane physical state during epididymal maturation. J. Androl. 12, 76–87.
| Correlation between changes in rat sperm membrane lipids, protein, and the membrane physical state during epididymal maturation.Crossref | GoogleScholarGoogle Scholar | 1849130PubMed |
Hughes, J., and Berger, T. (2015). Development of apical blebbing in the boar epididymis. PLoS One 10, e0126848.
| Development of apical blebbing in the boar epididymis.Crossref | GoogleScholarGoogle Scholar | 26555134PubMed |
Janett, F., Thun, R., Bettschen, S., Burger, D., and Hassig, M. (2003a). Seasonal changes of semen quality and freezability in Franches–Montagnes stallions. Anim. Reprod. Sci. 77, 213–221.
| Seasonal changes of semen quality and freezability in Franches–Montagnes stallions.Crossref | GoogleScholarGoogle Scholar | 12695055PubMed |
Janett, F., Thun, R., Niederer, K., Burger, D., and Hässig, M. (2003b). Seasonal changes in semen quality and freezability in the warmblood stallion. Theriogenology 60, 453–461.
| Seasonal changes in semen quality and freezability in the warmblood stallion.Crossref | GoogleScholarGoogle Scholar | 12763159PubMed |
Kenkel, C. D., Aglyamova, G., Alamaru, A., Bhagooli, R., Capper, R., Cunning, R., deVillers, A., Haslun, J. A., Hédouin, L., Keshavmurthy, S., Kuehl, K. A., Mahmoud, H., McGinty, E. S., Montoya-Maya, P. H., Palmer, C. V., Pantile, R., Sánchez, J. A., Schils, T., Silverstein, R. N., Squiers, L. B., Tang, P.-C., Goulet, T. L., and Matz, M. V. (2011). Development of gene expression markers of acute heat-light stress in reef-building corals of the genus Porites. PLoS One 6, e26914.
| Development of gene expression markers of acute heat-light stress in reef-building corals of the genus Porites.Crossref | GoogleScholarGoogle Scholar | 22046408PubMed |
Lenzi, A., Gandini, L., Maresca, V., Rago, R., Sgrò, P., Dondero, F., and Picardo, M. (2000). Fatty acid composition of spermatozoa and immature germ cells. Mol. Hum. Reprod. 6, 226–231.
| Fatty acid composition of spermatozoa and immature germ cells.Crossref | GoogleScholarGoogle Scholar | 10694269PubMed |
Leonard, A. E., Kelder, B., Bobik, E. G., Chuang, L.-T., Lewis, C. J., Kopchick, J. J., Mukerji, P., and Huang, Y.-S. (2002). Identification and expression of mammalian long-chain PUFA elongation enzymes. Lipids 37, 733–740.
| Identification and expression of mammalian long-chain PUFA elongation enzymes.Crossref | GoogleScholarGoogle Scholar | 12371743PubMed |
Livak, K. J., and Schmittgen, T. D. (2001). Analysis of relative gene expression data using real-time quantitative PCR and the 2–ΔΔCT method. Methods 25, 402–408.
| Analysis of relative gene expression data using real-time quantitative PCR and the 2–ΔΔCT method.Crossref | GoogleScholarGoogle Scholar | 11846609PubMed |
Loomis, P. R., and Graham, J. K. (2008). Commercial semen freezing: individual male variation in cryosurvival and the response of stallion sperm to customized freezing protocols. Anim. Reprod. Sci. 105, 119–128.
| Commercial semen freezing: individual male variation in cryosurvival and the response of stallion sperm to customized freezing protocols.Crossref | GoogleScholarGoogle Scholar | 18178040PubMed |
Macías García, B., González Fernández, L., Ortega Ferrusola, C., Morillo Rodríguez, A., Gallardo Bolaños, J. M., Rodríguez Martinez, H., Tapia, J. A., Morcuende, D., and Peña, F. J. (2011). Fatty acids and plasmalogens of the phospholipids of the sperm membranes and their relation with the post-thaw quality of stallion spermatozoa. Theriogenology 75, 811–818.
| Fatty acids and plasmalogens of the phospholipids of the sperm membranes and their relation with the post-thaw quality of stallion spermatozoa.Crossref | GoogleScholarGoogle Scholar | 21144567PubMed |
Macías-García, B., Gonzalez-Fernandez, L., Loux, S. C., Rocha, A. M., Guimaraes, T., Pena, F. J., Varner, D. D., and Hinrichs, K. (2015). Effect of calcium, bicarbonate, and albumin on capacitation-related events in equine sperm. Reproduction 149, 87–99.
| Effect of calcium, bicarbonate, and albumin on capacitation-related events in equine sperm.Crossref | GoogleScholarGoogle Scholar | 25349439PubMed |
Matsuzaka, T., Shimano, H., Yahagi, N., Amemiya-Kudo, M., Yoshikawa, T., Hasty, A. H., Tamura, Y., Osuga, J., Okazaki, H., Iizuka, Y., Takahashi, A., Sone, H., Gotoda, T., Ishibashi, S., and Yamada, N. (2002). Dual regulation of mouse Δ5- and Δ6-desaturase gene expression by SREBP-1 and PPARα. J. Lipid Res. 43, 107–114.
| 11792729PubMed |
Mohammed, B. S., Sankarappa, S., Geiger, M., and Sprecher, H. (1995). Reevaluation of the pathway for the metabolism of 7,10,13,16-docosatetraenoic acid to 4,7,10,13,16-docosapentaenoic acid in rat liver. Arch. Biochem. Biophys. 317, 179–184.
| Reevaluation of the pathway for the metabolism of 7,10,13,16-docosatetraenoic acid to 4,7,10,13,16-docosapentaenoic acid in rat liver.Crossref | GoogleScholarGoogle Scholar | 7872781PubMed |
Oresti, G. M., Reyes, J. G., Luquez, J. M., Osses, N., Furland, N. E., and Aveldaño, M. I. (2010). Differentiation-related changes in lipid classes with long-chain and very long-chain polyenoic fatty acids in rat spermatogenic cells. J. Lipid Res. 51, 2909–2921.
| Differentiation-related changes in lipid classes with long-chain and very long-chain polyenoic fatty acids in rat spermatogenic cells.Crossref | GoogleScholarGoogle Scholar | 20610732PubMed |
Parks, J. E., and Lynch, D. V. (1992). Lipid composition and thermotropic phase behavior of boar, bull, stallion, and rooster sperm membranes. Cryobiology 29, 255–266.
| Lipid composition and thermotropic phase behavior of boar, bull, stallion, and rooster sperm membranes.Crossref | GoogleScholarGoogle Scholar | 1582232PubMed |
Rejraji, H., Sion, B., Prensier, G., Carreras, M., Motta, C., Frenoux, J.-M., Vericel, E., Grizard, G., Vernet, P., and Drevet, J. R. (2006). Lipid remodeling of murine epididymosomes and spermatozoa during epididymal maturation. Biol. Reprod. 74, 1104–1113.
| Lipid remodeling of murine epididymosomes and spermatozoa during epididymal maturation.Crossref | GoogleScholarGoogle Scholar | 16510839PubMed |
Ronquist, G., Brody, I., Gottfries, A., and Stegmayr, B. (1978a). An Mg2+ and Ca2+-stimulated adenosine triphosphatase in human prostatic fluid: part I. Andrologia 10, 261–272.
| An Mg2+ and Ca2+-stimulated adenosine triphosphatase in human prostatic fluid: part I.Crossref | GoogleScholarGoogle Scholar | 152589PubMed |
Ronquist, G., Brody, I., Gottfries, A., and Stegmayr, B. (1978b). An Mg2+ and Ca2+-stimulated adenosine triphosphatase in human prostatic fluid – part II. Andrologia 10, 427–433.
| An Mg2+ and Ca2+-stimulated adenosine triphosphatase in human prostatic fluid – part II.Crossref | GoogleScholarGoogle Scholar | 153718PubMed |
Sæther, T., Tran, T. N., Rootwelt, H., Christophersen, B. O., and Haugen, T. B. (2003). Expression and regulation of Δ5-desaturase, Δ6-desaturase, stearoyl-coenzyme A (CoA) desaturase 1, and stearoyl-CoA desaturase 2 in rat testis. Biol. Reprod. 69, 117–124.
| Expression and regulation of Δ5-desaturase, Δ6-desaturase, stearoyl-coenzyme A (CoA) desaturase 1, and stearoyl-CoA desaturase 2 in rat testis.Crossref | GoogleScholarGoogle Scholar | 12606372PubMed |
Santiago Valtierra, F. X., Peñalva, D. A., Luquez, J. M., Furland, N. E., Vásquez, C., Reyes, J. G., Aveldaño, M. I., and Oresti, G. M. (2018). Elovl4 and Fa2h expression during rat spermatogenesis: a link to the very-long-chain PUFAs typical of germ cell sphingolipids. J. Lipid Res. 59, 1175–1189.
| Elovl4 and Fa2h expression during rat spermatogenesis: a link to the very-long-chain PUFAs typical of germ cell sphingolipids.Crossref | GoogleScholarGoogle Scholar | 29724783PubMed |
Scarlet, D., Ertl, R., Aurich, C., and Steinborn, R. (2015). The orthology clause in the next generation sequencing era: novel reference genes identified by RNA-seq in humans improve normalization of neonatal equine ovary RT-qPCR data. PLoS One 10, e0142122.
| The orthology clause in the next generation sequencing era: novel reference genes identified by RNA-seq in humans improve normalization of neonatal equine ovary RT-qPCR data.Crossref | GoogleScholarGoogle Scholar | 26536597PubMed |
Voss, A., Reinhart, M., Sankarappa, S., and Sprecher, H. (1991). The metabolism of 7,10,13,16,19-docosapentaenoic acid to 4,7,10,13,16,19-docosahexaenoic acid in rat liver is independent of a 4-desaturase. J. Biol. Chem. 266, 19995–20000.
| 1834642PubMed |
Wach-Gygax, L., Burger, D., Malama, E., Bollwein, H., Fleisch, A., Jeannerat, E., Thomas, S., Schuler, G., and Janett, F. (2017). Seasonal changes of DNA fragmentation and quality of raw and cold-stored stallion spermatozoa. Theriogenology 99, 98–104.
| Seasonal changes of DNA fragmentation and quality of raw and cold-stored stallion spermatozoa.Crossref | GoogleScholarGoogle Scholar | 28708505PubMed |
Wrench, N., Pinto, C. R., Klinefelter, G. R., Dix, D. J., Flowers, W. L., and Farin, C. E. (2010). Effect of season on fresh and cryopreserved stallion semen. Anim. Reprod. Sci. 119, 219–227.
| Effect of season on fresh and cryopreserved stallion semen.Crossref | GoogleScholarGoogle Scholar | 20227205PubMed |
Xie, F., Xiao, P., Chen, D., Xu, L., and Zhang, B. (2012). miRDeepFinder: a miRNA analysis tool for deep sequencing of plant small RNAs. Plant Mol. Biol. 80, 75–84.
| miRDeepFinder: a miRNA analysis tool for deep sequencing of plant small RNAs.Crossref | GoogleScholarGoogle Scholar |
