Transcriptome Analysis Reveals Differences in Skeletal Muscle Gene Expression in Sheep (Ovis aries) of Different Genders
LOU Meng-Yu1,2, LU Jia-Ni1,2, DUAN Qin1,2, YANG Wang-Xin1,2, ZHENG Qi1,2, ZHU Cui-Yun1,2, ZHANG Si-Huan1,2, LI Shuang1,2, LING Ying-Hui1,2,*
1 College of Animal Science and Technology, Anhui Agricultural University, Hefei 230036, China; 2 Anhui Provincial Laboratory of Animal Genetic Resources Protection and Biological Breeding, Hefei 230036, China
Abstract:Sex is one of the main factors affecting skeletal muscle development and meat production performance in sheep (Ovis aries). This study investigated the differences in gene expression in sheep longest dorsal muscle of different sexes using transcriptome sequencing technology, aiming to reveal the molecular mechanism of the effect of sex on skeletal muscle development in sheep. A total of 14 587 genes were identified by transcriptome sequencing and analysis, among which 29 genes were significantly differentially expressed. 23 genes were significantly up-regulated and 6 were down-regulated in the ram group compared with the ewe group. The differential genes included xin actin binding repeat containing 1 (XIRP1), glutathione peroxidase 2 (GPX2), acyl-CoA synthetase medium chain family member 1 (ACSM1), fatty acid binding protein 4 (FABP4), testis specific serine kinase 6 (TSSK6), etc. The GO analysis showed that the differentially expressed genes were mainly enriched in ribonucleoside, actin-binding and cytoskeleton-binding categories, and the KEGG pathway analysis revealed that the differentially expressed genes were significantly enriched in 3 signaling pathways, namely PPAR signaling pathway, arachidonic acid metabolism and glutathione metabolism signaling pathway. Thus, differences in skeletal muscle development in sheep between the sexes may arise through the above pathways. Eight differentially expressed genes were randomly selected and their expression was verified by qPCR, and the results were generally consistent with the transcriptome sequencing results, which proved that the sequencing results were reliable. In conclusion, this study revealed the differences in skeletal muscle gene expression and related signaling pathways in sheep of different sexes, which provides reference for further understanding the mechanisms by which sex affects muscle development in sheep.
[1] 曹婷, 施力光, 荀文娟, 等. 2020. 影响猪骨骼肌发育的主要分子因素及研究进展[J]. 家畜生态学报, 43(03): 7-12. (Cao T, Shi L G, Xun W J, et al.2022. Main molecular factors affecting porcine skeletal muscle development and research rogress[J]. Journal of Livestock Ecology, 43(03): 7-12.) [2] 郭丽荣. 2020. 基于转录组测序研究绵羊骨骼肌在营养应激下的代谢调节机制[D]. 硕士学位论文, 山西农业大学, 导师: 杜荣, pp. 1-84. (GUO L R.2020. The metabolic regulation mechanism of sheep skeletal muscle under nutritional stress was studied based on transcription sequencing[D]. Thesis for M.S., Shanxi Agricultural University,Supervisor: Du R, pp. 1-84.) [3] 石田培, 王欣悦, 侯浩宾, 等. 2020. 基于全转录组测序的绵羊胚胎不同发育阶段骨骼肌circRNA的分析与鉴定[J]. 中国农业科学, 53(03): 642-657. (Shi T P, Wang X Y, Hou H B, et al.2020. Analysis and identification of skeletal muscle circRNA at different developmental stages of sheep embryos based on whole transcriptome sequencing[J]. Scientia Agricultura Sinica, 53(03): 642-657.) [4] 孙燕勇, 付绍印, 何小龙, 等. 2019. 肉用畜禽肌纤维发育特性[J]. 中国农业大学学报, 24(12): 78-85. (Sun Y Y, Fu S Y, He X L, et al.2019. Development characteristics of muscle fiber in meat livestock[J]. Journal of China Agricultural University, 24(12): 78-85.) [5] 吴祖纯, 刘新光, 陈维春. 2022. 丝状肌动蛋白的组装及其生物学功能[J]. 生命的化学, 42(05): 984-992. (Wu Z C, Liu X G, Chen W C.2022. Assembly of filamentous actin and its biological function[J]. The Chemistry of Life, 42(05): 984-992.) [6] 张安宁, 罗雪林, 黄思琴, 等. 2018. PI3K/Akt/mTOR信号通路在大鼠急性骨骼肌钝挫伤修复中的作用[J]. 中国运动医学杂志, 37(7): 594-600. (Zhang A N, Luo X L, Huang S Q, et al.2018. Role of PI3K/Akt/mTOR signaling pathway in repair of acute skeletal muscle contusion in rats[J]. Chinese Journal of Sports Medicine, 37(7): 594-600.) [7] 郑琪. 2020. 胎儿到羔羊七个阶段的山羊背最长肌组织学特性及mRNA和lncRNA调控网络分析[D]. 硕士学位论文, 安徽农业大学, 导师: 凌英会, pp. 1-60 (Zheng Q.2020. Histological characteristics and mRNA and lncRNA regulatory network analysis of goat longissimus dorsi muscle at seven stages from fetus to lamb[D]. Thesis for M.S., Anhui Agricultural University, Supervisor: Ling Y H, pp. 1-60.) [8] Anderson L J, Liu H, Garcia J M.2017. Sex differences in muscle wasting[J]. Advances in Experimental Medicine and Biology, 1043: 153-197. [9] Clarke S D, Clarke I J, Rao A, et al.2012. Sex differences in the metabolic effects of testosterone in sheep[J]. Endocrinology, 153(1): 123-131. [10] Devries M C.2016. Sex-based differences in endurance exercise muscle metabolism: Impact on exercise and nutritional strategies to optimize health and performance in women[J]. Experimental Physiology, 101(2): 243-249. [11] Duszka K, Wahli W.2018. Enteric microbiota-gut-brain axis from the perspective of nuclear receptors[J]. International Journal of Molecular Science, 19(8): 2210. [12] Freking B A, King D A, Shackelford S D, et al.2018. Effects and interactions of myostatin and callipyge mutations: I. Growth and carcass traits[J]. Journal of Animal Science,96(2): 454-461. [13] Fu M H, Maher A C, Hamadeh M J, et al.2009. Exercise, sex, menstrual cycle phase, and 17b-estradiolinfluence metabolism-related genes in human skeletal muscle[J]. Physiological Genomics, 40(1): 34-47. [14] Gao C Q, Shi H Q, Xie W Y, et al.2021. Dietary supplementation with acidifiers improves the growth performance, meat quality and intestinal health of broiler chickens[J]. Animal Nutrition, 7: 762-769. [15] Garin-Shkolnik T, Rudich A, Hotamisligil G S, et al.2014. FABP4 attenuates PPARγ and adipogenesis and is inversely correlated with PPARγ in adipose tissues[J]. Diabetes, 63(3): 900-911. [16] Guo Y, Wang M, Ge J, et al.2020. Bioactive biodegradable polycitrate nanoclusters enhances the myoblast differentiation and in vivo skeletal muscle regeneration via p38 MAPK signaling pathway[J]. Bioactive Materials, 5(3): 486-495. [17] Hawke T J, Atkinson D J, Kanatous S B, et al.2007. Xin, an actin binding protein, is expressed within muscle satellite cells and newly regenerated skeletal muscle fibers[J]. American Journal of Physiology-Cell Physiology, 293(5): C1636-44. [18] Huang J, Jia Y, Li Q, et al.2018. Glutathione content and expression of proteins involved with glutathione metabolism differs in longissimus dorsi, subcutaneous adipose, and liver tissues of finished vs. growing beef steers[J]. Journal of Animal Science, 96(12): 5152-5165. [19] He N, Lang X, Wang C, et al.2023. Expression of MSTN/Smad signaling pathway genes and its association with meat quality in Tibetan sheep (Ovis aries)[J]. Nutrition & Food Science, 11(4): 1836-1845. [20] Huang C, Blecker C, Chen L, et al.2023. Integrating identification and targeted proteomics to discover the potential indicators of postmortem lamb meat quality[J]. Meat Science, 199: 109126. [21] Liu X, Wang Y, Zhao S, et al.2017. Fibroblast growth factor 21 promotes C2C12 cells myogenic differentiation by enhancing cell cycle exit[J]. Biomed Research International, 2017: 1648715. [22] Littlefield R S, Fowler V M.2008. Thin filament length regulation in striated muscle sarcomeres: Pointed-end dynamics go beyond a nebulin ruler[J]. Seminars in Cell & Developmental Biology, 19(6): 511-519. [23] Mal A, Chattopadhyay D, Ghosh M K, et al.2000. p21 and retinoblastoma protein control the absence of DNA replication in terminally differentiated muscle cells[J]. Journal of Cell Biology, 149(2): 281-292. [24] Manickam R, Duszka K, Wahli W.2020. PPARs and microbiota in skeletal muscle health and wasting[J]. International Journal of Molecular Science, 21(21): 8056. [25] Markworthy J F, Cameron-Smith D.2013. Arachidonic acid supplementation enhances in vitro skeletal muscle cell growth via a COX-2-dependent pathway[J]. American Journal of Physiology-Cell Physiology, 304(1): C56-67. [26] Miotto P M, McGlory C, Holloway T M, et al.2018. Sex differences in mitochondrial respiratory function in human skeletal muscle[J]. American Journal of Physiology-Regulatory Integrative and Comparative Physiology, 314(6): R909-R915. [27] Needham T, Laubser J G, Kotrba R, et al.2019. Sensory characteristics of the longissimus thoracis et lumborum and biceps femoris muscles from male and female common eland (Taurotragus oryx)[J]. Meat Sciencce, 158: 107918. [28] Ono S.2010. Dynamic regulation of sarcomeric actin filaments in striated muscle[J]. Cytoskeleton (Hoboken), 67(11): 677-692. [29] Ono-Moore K D, Olfert I M, Rutkowsky J M.2021. Metabolic physiology, and skeletal muscle phenotypes in male and female myoglobin knockout mice[J]. American Journal of Physiology-Endocrinology And Metabolism, 321(1): E63-E79. [30] Oosthuyse T, Bosch A N.2012. Oestrogen's regulation of fat metabolism during exercise and gender specific effects[J]. Current Opinion Pharmacology, 12(3): 363-371. [31] O'Reilly J, Ono-Moore K D, Chintapalli S V, et al.2021. Sex differences in skeletal muscle revealed through fiber type, capillarity, and transcriptomics profiling in mice[J]. Physiological Reports, 9(18): e15031. [32] Pacholsky D, Vakeel P, Himmel M, et al.2004. Xin repeats define a novel actin-binding motif[J]. Journal of Cell Science, 117(Pt 22): 5257-5268. [33] Rudolph T E, Mayorga E J, Roths M, et al.2021. The effect of mitoquinol (MitoQ) on heat stressed skeletal muscle from pigs, and a potential confounding effect of biological sex[J]. Journal of Thermal Biology, 97: 102900. [34] Sinn H W, Balsamo J, Lilien J, et al.2002. Localization of the novel Xin protein to the adherents junction complex in cardiac and skeletal muscle during development[J]. Developmental Dynamics, 225(1): 1-13. [35] Smith A G, Muscat G E O.2005. Skeletal muscle and nuclear hormone receptors: Implications for cardiovascular and metabolic disease[J]. International Journal of Biochemistry & Cell Biology, 37(10): 2047-2063. [36] Tontonoz P, Hu E, Spiegelman B M.1994. Stimulation of adipogenesis in fibroblasts by PPAR gamma 2, a lipid-activated transcription factor[J]. Cell, 79(7): 1147-1156. [37] Wang X W, Sun Y J, Chen X, et al.2022. Interleukin-4-induced FABP4 promotes lipogenesis in human skeletal muscle cells by activating the PPAR γ signaling pathway[J]. Cell Biochemistry and Biophysics, 80(2): 355-366. [38] Yoo Y M, Jung E M, Jeung E B.2019. Rapamycin-induced autophagy decreases Myf5 and MyoD proteins in C2C12 myoblast cells[J]. Toxicol In Vitro, 58: 132-141. [39] Zhu Y, Qi C, Korenberg J R, et al.1995. Structural organization of mouse peroxisome proliferator-activated receptor gamma (mPPAR gamma) gene: Alternative promoter use and different splicing yield two mPPAR gamma isoforms[J]. Proceedings of the National Academy of Sciences of the USA, 92(17): 7921-7925. [40] Zhu Y, Alvares K, Huang Q, et al.1993. Cloning of a new member of the peroxisome proliferator-activated receptor gene family from mouse liver[J]. Journal of Biological Chemistry, 268(36): 26817-26820.