Expression Analysis of Hippo Signaling Pathway Related-genes in Yak (Bos grunniens) Ovaries During Different Reproductive Cycles
LI Yi-Juan1, FAN Jiang-Feng1,2,*, YU Si-Jiu1,2, ZHOU Ying-Cong1, QIAN Wen-Jie1, YAO Ying1
1 College of Veterinary Medicine, Gansu Agricultural University, Lanzhou 730070, China; 2 College of Veterinary Medicine, Gansu Agricultural University/Research Center of Bovine and Sheep Embryo Engineering Technology, Lanzhou 730070, China
Abstract Hippo signaling pathway is a key pathway regulating the growth and development of animal tissues and organs, and plays an important role in mammalian ovarian development. To investigate the Hippo signaling pathway related genes mammalian ste20-like protein kinase 1 (MST1), large tumor suppressor 1 (LATS1) and Yes-associated protein 1 (YAP1) in yak (Bos grunniens) ovaries during different reproductive cycles, in this study, the ovarian tissues of healthy female yaks (Bos grunniens) from different reproductive cycles (follicular, luteal, gestation) were selected as the test samples. qRT-PCR and Western blot were used to detect the expression differences of the MST1, LATS1 and YAP1 and their protein in yak ovaries of different reproductive cycles. Immunohistochemistry (IHC) was used to detect the distribution of genes related to Hippo signaling pathway in yak ovaries at 3 cycles. The results of qRT-PCR showed that there were significant differences in the expression of MST1 gene in ovarian tissues at different stages (P<0.05) with the highest expression in follicular stage, the second in gestation stage and the lowest in luteal stage. The expression levels of LATS1 and YAP1 genes in pregnancy were significantly higher than those in follicular and luteal phases (P<0.05). Western blot results showed that the protein expressions of MST1, LATS1 and YAP1 increased gradually, and their expression reached the peak in pregnancy phase, then decreased and maintained at a low level, which was significantly different from other stages of pregnancy phase (P<0.05). IHC results showed that MST1 was mainly distributed in granulosa cells, membrane cells, granular luteal cells, membranous luteal cells and ovarian germ epithelium of ovarian follicles. The distribution positions of LATS1 and YAP1 in different ovarian cycles were consistent with those of MST1. MST1, LATS1 and YAP1 were expressed in yak ovaries in different reproductive cycles, and the expression levels were significantly different, suggesting that MST1, LATS1 and YAP1 may be involved in the regulation of reproductive physiology related processes in yak. This study provides insights into the relationship between the expression of MST1, LATS1 and YAP1 in follicular granulosa cells of yaks at different developmental stages, which will provide a theoretical basis for further research on the reproductive performance of yaks by exploring Hippo signaling pathway.
LI Yi-Juan,FAN Jiang-Feng,YU Si-Jiu, et al. Expression Analysis of Hippo Signaling Pathway Related-genes in Yak (Bos grunniens) Ovaries During Different Reproductive Cycles[J]. 农业生物技术学报, 2023, 31(10): 2106-2114.
[1] 鲍建军, 苏锐, 王庆增, 等. 2016. Smads与Hippo通道中YAP1基因在湖羊肌肉组织中时空表达研究及关联分析[J]. 中国农业科学, 49(11): 2203-2213. (Bao J J, Su R, Wang Q Z, et al.2016. Temporal and spatial expression and correlation analysis of YAP1 gene in Smads and Hippo channels in Hu sheep muscle tissue[J]. Chinese Agricultural Sciences, 49(11): 2203-2213.) [2] 谷博. 2019. 绵羊的卵巢不同发育时期miRNA-mRNA表达谱整合分析研究[D]. 博士学位论文, 吉林农业大学, 导师: 姜怀志, pp. 84-87. (Gu B.2019. Integrated analysis of miRNA-mRNA expression profile in sheep ovaries at different developmental stages[D]. Jilin Agricultural University, Supervisor: Jiang H Z, pp. 84-87.) [3] 郭立文. 2015. Hippo信号通路下游转录效应因子TAZ/YAP在乳腺癌中作用的初步研究[D]. 博士学位论文, 浙江大学, 导师: 滕理送, pp. 50-52. (Guo L W.2015. A preliminary study on the role of TAZ/YAP transcription factor downstream of Hippo signaling pathway in breast cancer[D]. Zhejiang University, Supervisor: Teng L S, pp. 50-52.) [4] 李佳, 袁树晟, 曹秀萍, 等. 2020. Hippo信号通路调节卵巢物质代谢对卵巢功能影响的研究进展[J]. 上海交通大学学报(医学版), 40(11): 1536-1539. (Li J, Yuan S S, Cao X P, et al.2020. Research progress of Hippo signaling pathway in regulating ovarian metabolism and ovarian function[J]. Journal of Shanghai Jiao Tong University (med), 40(11): 1536-1539.) [5] 闾少冬, 魏勇鹏, 王卓, 等. 2021. Hippo信号通路中MST1和MST2的表达在肝癌干细胞干性维持和自我更新中的意义[J]. 肝脏, 26(08): 866-870. (Lu S D, Wei Y P, Wang Z, et al.2021. Expression of MST1 and MST2 in Hippo signaling pathway and its significance in dry maintenance and self-renewal of hepatocarcinoma stem cells[J]. The liver, 26(08) : 866-870.) [6] 李晓宇, 田字彬, 单信芝, 等. 2013. Hippo通路中Yes相关蛋白在胰腺癌中的表达及意义[J]. 世界华人消化杂志, 21(28): 3013-3018. (Li X Y, Tian Z B, Shan X Z, et al.2013. Expression and significance of Yes-related proteins in Hippo pathway in pancreatic cancer[J]. World Chinese Journal of Digestion, 21(28): 3013-3018.) [7] 潘阳阳, 王萌, 余四九, 等. 2017. EGF对牦牛冻精运动性能、细胞凋亡和IVF胚胎发育潜力的影响[J]. 畜牧兽医学报, 48(05): 854-862. (Pan Y Y, Wang M, Yu S J, et al.2017. Effects of EGF on motor performance, cell apoptosis and embryo development potential of yak IVF[J].Chinese Journal of Animal Science and Veterinary Medicine, 48(05): 854-862.) [8] 任洁, 林文洋, 阮灵伟. 2023. Hippo信号通路与海洋无脊椎动物天然免疫[J]. 中国生物化学与分子生物学报, 39(01): 33-41. (Ren J, Lin W Y, Ruan L W.2023. Hippo signaling pathway and natural immunity in Marine invertebrates[J]. Chinese Journal of Biochemistry and Molecular Biology, 39(01): 33-41.) [9] 徐瑞, 李丽娜, 王文娟. 2020. Hippo信号通路介导PTEN调节PI3K/AKT/mTOR信号通路影响肝癌细胞增殖能力[J]. 山西医科大学学报, 51(12): 1277-1283. (Xu R, LI L N, Wang W J.2020. Hippo signaling pathway mediated by PTEN regulates PI3K/AKT/mTOR signaling pathway and affects the proliferation of hepatocarcinoma cells[J]. Journal of Shanxi Medical University, 51(12): 1277-1283.) [10] 阎萍, 潘和平. 2004. 环境因素对牦牛繁殖性能的影响[J]. 畜牧与兽医, (05): 15-16. (Yan P, Pan H P. 2004. Effects of environmental factors on reproductive performance of Yak[J]. Animal Husbandry and Veterinary Medicine, (05): 15-16.) [11] 杨珊珊, 何翃闳, 潘阳阳, 等. 2020. 整合素αv在牦牛繁殖周期输卵管不同部位中的表达与定位[J]. 畜牧兽学报, 51(11): 2710-2719. (Yang S S, He H H, Pan Y Y, et al.2020. Expression and localization of integrin αv in different parts of oviduct during reproductive cycle of yak[J]. Chinese Journal of Animal Science, 51(11): 2710-2719.) [12] 尧阳扬. 2020. E3泛素连接酶FBXW5失活Hippo通路促进胃癌细胞转移的分子机制[D]. 博士学位论文, 南昌大学, 导师: 熊建萍, pp. 21-23. (Yao Y Y.2020. Molecular mechanism of Hippo pathway inactivation of E3 ubiquitin ligase FBXW5 promoting gastric cancer cell metastasis[D]. Nanchang University, Supervisor: Xiong J P, pp. 21-23.) [13] 张凡, 王星星, 朱新景, 等. 2021. 紫草素通过调控Hippo-YAP信号通路对咪喹莫特诱导银屑病小鼠保护作用的研究[J]. 现代中西医结合杂志, 30(06): 578-583. (Zhang F, Wang X X, Zhu X J, et al.2021. Study on the protective effect of Shikonin on imiquimote-induced psoriasis in mice by regulating Hippo-YAP signaling pathway[J]. Journal of Modern Integrated Chinese and Western Medicine, 30(06): 578-583.) [14] 张慧玲, 郑华川, 赵爽, 等. 2019. 过表达LATS1抑制头颈部鳞状细胞癌B88细胞增殖和侵袭及其临床意义[J]. 肿瘤, 39(1): 1-9. (Zhang H L, Zheng H C, Zhao S, et al.2019. Overexpression of LATS1 inhibits proliferation and invasion of head and neck squamous cell carcinoma B88 cells and its clinical significance[J]. Cancer, 39(1): 1-9.) [15] 周梅, 徐成良, 王洁茹, 等. 2022. Hippo信号通路在大白猪不同直径卵泡中的表达特征研究[J]. 上海农业学报, 38(01): 73-76. (Zhou M, Xu C L, Wang J R, et al.2022. Characterization of Hippo signaling pathway in follicles of large white pigs[J]. Shanghai Journal of Agricultural Sciences, 38(01): 73-76.) [16] Abreu R, Penalva L O, Marcotte E M, et al.2009. Global signatures of protein and mRNA expression levels[J]. Molecular BioSystems, 5(12): 1512-1526. [17] Anorga, Sandybell, Overstreet, et al.2018. Deregulation of Hippo-TAZ pathway during renal injury confers a fibrotic maladaptive phenotype[J]. FASEB Journal, 32(5): 2644-2657. [18] Barry E R, Morikawa T, Butler B L, et al.2013. Restriction of intestinalstem cell expansion and the regenerative response by YAP[J]. Nature, 493(7430): 106-110. [19] Bodwell J E, Kingsley L A, Hamilton J W.2004. Arsenic at very low concentrations alters glucocorticoid receptor (GR)-mediated gene activation but not GR-mediated gene repression: Complex dose-response effects are closely correlated with levels of activated GR and require a functional GR DNA binding domain[J]. Chemical Research in Toxicology, 17(8): 1064-1076. [20] Calses P C, Crawford J J, Lill J R, et al.2019. Hippo pathway in cancer: Aberrant regulation and therapeutic opportunities[J]. Trends Cancer, 5(5): 297-307. [21] Cordenonsi M, Zanconato F, Azzolin L, et al.2011. The Hippo transducer TAZ confers cancer stem cell-related traits on breast cancer cells[J]. Cell, 147(4): 759-772. [22] Jin H, Zhang Y, Jing D, et al.2014. Role of Hippo signaling in cancer stem cells[J]. Journal of Cellular Physiology, 229(3): 266-270. [23] Koontz L M, Liu-Chittenden Y, Yin F, et al.2013. The Hippo effector Yorkie controls normal tissue growth by antagonizing scalloped-mediated default repression[J]. Developmental Cell, 25(4): 388-401. [24] Mo J S, Park H W, Guan K L.2014. The Hippo signaling pathway in stem cell biology and cancer[J]. EMBO Reports, 15(6): 642-656. [25] Pate J L, Johnson-Larson C J, Ottobre J S.2012. Life or death decisions in the corpus luteum[J]. Reproduction in Domestic Animals, 47(Supplement s4): 297-303. [26] Staley BK, Irvine KD.2012. Hippo signaling in drosophila: Recent advances and insights[J]. Developmental Dynamics, 241(1): 3-15. [27] Totaro A, Panciera T, Piccolo S.2018. YAP/TAZ upstream signals and downstream responses[J], Nature Cell Biology, 20(8): 888-899. [28] Wang C, Zhu Z M, Liu C J, et al.2015. Knockdown of Yes-associated protein inhibits proliferation and downregulates large tumor suppressor1 expression in MHCC97H human hepatocellular carcinoma cells[J]. Molecular Medicine Reports, 11(6): 4101-4108. [29] Wu C, Xu B, Yuan P, et al.2010. Genome-wide interrogation identifies YAP1 variants associated with survival of small-cell lung cancer patients[J]. Cancer Research, 70(23): 9721-9729. [30] Xu C M, Liu W W, Liu C J, et al.2013. Mst1 overexpression inhibited the growth of human non-small cell lung cancer in vitro and in vivo[J]. Cancer Gene Therapy, 20(8): 453-460. [31] Xiang C, Li J, Hu L, et al.2015. Hippo signaling pathway reveals a spatio-temporal correlation with the size of primordial follicle pool in mice[J]. Cellular Physiology and Biochemistry, 35(3): 957-968. [32] Yu S J, Yong Y H, Cui Y.2010. Oocyte morphology from primordial to early tertiary follicles of yak[J]. Reproduction in Domestic Animals, 45(5): 779-785. [33] Zeng Q, Hong W.2008. The emerging role of the hippo pathway in cell contact inhibition,organ size control,and cancer development in mammals[J]. Cancer Cell, 13(3): 188-192. [34] Zhao B, Li L, Tumaneng K, et al.2010a. A coordinated phosphorylation by LATS and CK1 regulates YAP stability through SCF (beta-TRCP)[J]. Genes & Development , 24(1): 72-85 [35] Zhao B, Li L, Lei Q, et al.2010b. The Hippo-YAP pathway in organ size control and tumorigenesis: An updated version[J]. Genes & Development, 24(9): 862-874. [36] Zhao B, Tumaneng K, Guan KL.2011. The Hippo pathway in organ size control, tissue regeneration and stem cell self -renewal[J]. Nature Cell Biology, 13(8): 877-883.
