绵羊不同繁殖状态下SIX1和TEF基因的表达特征分析
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摘要
试验旨在探究SIX1和TEF基因在苏尼特羊(Sunite sheep,SNT)和小尾寒羊(Small Tail Han sheep,STH)相关组织中的表达特征,有助于揭示上述2个基因在绵羊季节性发情和繁殖调控中的重要作用。选取季节性发情的SNT母羊在短光照(模拟繁殖季节)和长光照(模拟休情期)条件下及常年发情的STH母羊在不同繁殖时期(卵泡期和黄体期)的下丘脑等10种组织,利用qPCR技术分析上述不同繁殖状态下各组织中SIX1和TEF基因的相对表达量。结果表明,SIX1基因在SNT和STH的垂体组织中均高表达,其它组织中微弱表达;TEF基因在2个绵羊品种的多个组织中广泛表达;SNT垂体中TEF基因在短光照条件下其表达量显著高于长光照条件(P<0.05),STH垂体中TEF基因在卵泡期其表达量显著高于黄体期(P<0.05);SNT子宫体中TEF在长光照条件下其表达量显著高于短光照条件(P<0.05),STH子宫体中TEF在黄体期其表达量极显著高于卵泡期(P<0.01)。研究结果显示,SIX1和TEF基因表达在绵羊垂体中发挥重要作用,2个基因在SNT垂体中的表达变化趋势与已知的长光照诱导基因EYA3的表达变化不同,暗示绵羊垂体中SIX1和TEF基因不是通过转录水平变化来参与季节性发情上游基因的调控;TEF基因可能参与绵羊不同繁殖状态下子宫生理变化的调控。本研究为深入探究这2个基因在绵羊繁殖性能调控方面的作用奠定了基础。
The aim of the study is to analyze the expression characteristics of the SIX1 and TEFgenes in related tissues of Sunite sheep and Small Tail Han sheep,which might help reveal important role of two genes in sheep seasonal estrus and reproductive regulation.The qPCR method was used to detect the relative expression of SIX1 and TEF genes in ten tissues including hypothalamus of seasonal estrous sheep(Sunite sheep,SNT)at different photoperiods(short photoperiod and long photoperiod)and year-round estrous sheep(Small Tail Han sheep,STH)at different reproductive stages(follicular stage and luteal stage).The results showed that the SIX1 gene was highly expressed in pituitary gland and weakly expressed in other tissues of two breeds(SNT and STH).However,the TEFgene was widely expressed in related tissues of two breeds.Expression level of the TEFgene at short photoperiod(SP)was significantly higher than that at long photoperiod(LP)in pituitary gland of SNT(P<0.05),and its expression level was significantly higher at follicular stage than that at luteal stage in pituitary gland of STH(P<0.05).Expression level of the TEF gene at LP was significantly higher than that at SP in uterine body of SNT(P<0.05),and its expression level was extremely higher at luteal stage than that at follicular stage in uterine body of STH(P<0.01).The expression of SIX1 and TEF may play an important role in pituitary gland of sheep.The expression of two genes in pituitary gland of SNT is different from the expression of the EYA3 gene which is known induced gene at LP in pituitary gland of seasonal estrus sheep,which indicated that the two genes may not be involved in the regulation of upstream genes of sheep seasonal estrus pathway by change of transcription.Additionally,the TEFgene may be involved in uterus' physiological regulation between different reproductive stages in sheep.The results will lay a foundation for further study of two genes' function in sheep reproduction regulation.
The aim of the study is to analyze the expression characteristics of the SIX1 and TEFgenes in related tissues of Sunite sheep and Small Tail Han sheep,which might help reveal important role of two genes in sheep seasonal estrus and reproductive regulation.The qPCR method was used to detect the relative expression of SIX1 and TEF genes in ten tissues including hypothalamus of seasonal estrous sheep(Sunite sheep,SNT)at different photoperiods(short photoperiod and long photoperiod)and year-round estrous sheep(Small Tail Han sheep,STH)at different reproductive stages(follicular stage and luteal stage).The results showed that the SIX1 gene was highly expressed in pituitary gland and weakly expressed in other tissues of two breeds(SNT and STH).However,the TEFgene was widely expressed in related tissues of two breeds.Expression level of the TEFgene at short photoperiod(SP)was significantly higher than that at long photoperiod(LP)in pituitary gland of SNT(P<0.05),and its expression level was significantly higher at follicular stage than that at luteal stage in pituitary gland of STH(P<0.05).Expression level of the TEF gene at LP was significantly higher than that at SP in uterine body of SNT(P<0.05),and its expression level was extremely higher at luteal stage than that at follicular stage in uterine body of STH(P<0.01).The expression of SIX1 and TEF may play an important role in pituitary gland of sheep.The expression of two genes in pituitary gland of SNT is different from the expression of the EYA3 gene which is known induced gene at LP in pituitary gland of seasonal estrus sheep,which indicated that the two genes may not be involved in the regulation of upstream genes of sheep seasonal estrus pathway by change of transcription.Additionally,the TEFgene may be involved in uterus' physiological regulation between different reproductive stages in sheep.The results will lay a foundation for further study of two genes' function in sheep reproduction regulation.
引文
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[18]WU W,HUANG R,WU Q,et al.The role of SIX1in the genesis of muscle cell and skeletal muscle development[J].International Journal of Biological Sciences,2014,10(9):983-989.
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[21]LI W K,LI H,LU Y F,et al.Atorvastatin alters the expression of genes related to bile acid metabolism and circadian clock in livers of mice[J].Peer J,2017,5(1):e3348.
[22]NAKAO N,ONO H,Yamamura T,et al.Thyrotrophin in the pars tuberalis triggers photoperiodic response[J].Nature,2008,452(7185):317-322.
[23]HENNINGSEN J B,GAUE F,SIMONNEAUX V.RFRP neurons-the doorway to understanding seasonal reproduction in mammals[J].Frontiers in Endocrinology,2016,7(9):36-45.
[24]SURBHI,KUMAR V.Avian photoreceptors and their role in the regulation of daily and seasonal physiology[J].General&Comparative Endocrinology,2015,220:13-22.
[25]DUPRE S M,MIEDZINSKA K,DUVAL C V,et al.Identification of EYA3and TAC1as long-day signals in the sheep pituitary[J].Current Biology,2010,20(9):829-835.
[26]MASUMOTO K,UKAITADENUMA M,KASUKAWA T,et al.Acute induction of EYA3by late-night light stimulation triggers TSHβexpression in photoperiodism[J].Current Biology,2010,20(24):2 199-2 206.
[27]曹晓涵,刘秋月,王翔宇,等.小尾寒羊黄体期和卵泡期生殖轴凋亡基因的表达研究及其与发情的关系[J].畜牧兽医学报,2017,48(11):2 091-2 097.
[28]董新龙,胡文萍,贺小云,等.绵羊BMP15组织表达特征及FecXGr、FecXO和G971A突变的检测[J].农业生物技术学报,2016,24(12):1 810-1 819.
[29]汤继顺,狄冉,刘秋月,等.精卵融合相关基因CD9和CD81在绵羊组织中的表达分析[J].农业生物技术学报,2018,26(1):87-95.
[30]SCHMITTGEN T D,LIVAK K J.Analyzing real-time PCR data by the comparative C(T)method[J].Nature Protocols,2008,3(6):1 101-1 108.
[31]LEWIS P D.A review of lighting for broiler breeders[J].British Poultry Science,2006,47(4):393-404.
[32]DARDENTE H.Circannual biology:the double life of the seasonal thyrotroph[J].Current Biology,2015,25(20):988-991.
[33]TAMAI T K,YOSHIMURA T.Molecular and neuroendocrine mechanisms of avian seasonal reproduction[J].Advances in Experimental Medicine and Biology,2017,1001:125-136.
[34]DARDENTE H,WYSE C A,BIMIE M J,et al.A molecular switch for photoperiod responsiveness in mammals[J].Current Biology,2010,20(24):2 193-2 198.
[35]DARDENTE H,LOMET D,ROBERT V,et al.Seasonal breeding in mammals:from basic science to applications and back[J].Theriogenology,2016,86(1):324-332.
[36]WOOD S H,CHRISTIAN H C,MIEDZINSKA K,et al.Binary switching of calendar cells in the pituitary defines the phase of the circannual cycle in mammals[J].Current Biology,2015,25(20):2 651-2 662.
[37]夏青,张效生,刘秋月,等.EYA3和TSHβ在季节性发情和常年发情绵羊中的表达模式[J].畜牧兽医学报,2018,49(2):263-269.
[2]宋陈玲,胡永波.禽类繁殖季节性研究[J].畜牧与饲料科学,2010,31(4):32-33.
[3]黄冬维,储明星,狄冉,等.山羊繁殖季节性相关基因KiSS-1与RFRP的表达研究[J].畜牧兽医学报,2015,46(6):924-931.
[4]DALLMANN R,BROWN S A,GACHON F.Chronopharmacology:new insights and therapeutic implications[J].Annual Review of Pharmacology,2014,54(1):339-361.
[5]LIU Y,HAN N,ZHOU S,et al.The DACH/EYA/SIX gene network and its role in tumor initiation and progression[J].International Journal of Cancer,2015,138(5):1 067-1 075.
[6]STIERWALD M,YANZE N,BAMERT R P,et al.The sine oculis/SIX class family of homeobox genes in jellyfish with and without eyes:development and eye regeneration[J].Developmental Biology,2004,274(1):70-81.
[7]ANDERSON A M,WEASNER B M,WEASNER B P,et al.Dual transcriptional activities of SIX proteins define their roles in normal and ectopic eye development[J].Development,2012,139(5):991-1 000.
[8]ZOU D,SILVIUS D,FRITZSCH B,et al.EYA1and SIX1are essential for early steps of sensory neurogenesis in mammalian cranial placodes[J].Development,2004,131(22):5 561-5 572.
[9]IMARAZENE B,ANDOUCHE A,BASSAGLIA Y,et al.Eye development in sepia officinalis embryo:what the uncommon gene expression profiles tell us about eye evolution[J].Frontiers in Physiology,2017,8:613-627.
[10]NIE X,XU J,ELHASHASH A,et al.SIX1regulates grem1expression in the metanephric mesenchyme to initiate branching morphogenesis[J].Developmental Biology,2011,352(1):141-151.
[11]KONISHI Y,IKEDA K,IWAKURA Y,et al.SIX1and SIX4promote survival of sensory neurons during early trigeminal gangliogenesis[J].Brain Research,2006,1116(1):93-102.
[12]FONSECA B F,COULY G,DUPIN E.Respective contribution of the cephalic neural crest and mesoderm to SIX1-expressing head territories in the avian embryo[J].BMC Developmental Biology,2017,17(1):13-27.
[13]WU W,REN Z,ZHANG L,et al.Overexpression of SIX1gene suppresses proliferation and enhances expression of fast-type muscle genes in C2C12 myoblasts[J].Molecular and Cellular Biochemistry,2013,380(1-2):23-32.
[14]WU W,REN Z,WANG Y,et al.Molecular characterization,expression patterns and polymorphism analysis of porcine SIX1gene[J].Molecular Biology Reports,2011,38(4):2619-2632.
[15]MOOTOOSAMY R C,DIETRICH S.Distinct regulatory cascades for head and trunk myogenesis[J].Development,2002,129(3):573-583.
[16]SATO S,IKEDA K,SHIOI G,et al.Regulation of SIX1expression by evolutionarily conserved enhancers in tetrapods[J].Developmental Biology,2012,368(1):95-108.
[17]KUMAR J P.The sine oculis homeobox(SIX)family of transcription factors as regulators of development and disease[J].Cellular&Molecular Life Sciences,2009,66(4):565-583.
[18]WU W,HUANG R,WU Q,et al.The role of SIX1in the genesis of muscle cell and skeletal muscle development[J].International Journal of Biological Sciences,2014,10(9):983-989.
[19]GACHON F,FONJALLAZ P,DAMIOLA F,et al.The loss of circadian PAR bZip transcription factors results in epilepsy[J].Genes&Development,2004,18(12):1 397-1 412.
[20]CHOPRA A,TYE S J,LEE K H,et al.Underlying neurobiology and clinical correlates of mania status-post subthalamic nucleus deep brain stimulation in parkinson's disease:a review of the literature[J].Journal of Neuropsychiatry&Clinical Neurosciences,2012,24(1):102-110.
[21]LI W K,LI H,LU Y F,et al.Atorvastatin alters the expression of genes related to bile acid metabolism and circadian clock in livers of mice[J].Peer J,2017,5(1):e3348.
[22]NAKAO N,ONO H,Yamamura T,et al.Thyrotrophin in the pars tuberalis triggers photoperiodic response[J].Nature,2008,452(7185):317-322.
[23]HENNINGSEN J B,GAUE F,SIMONNEAUX V.RFRP neurons-the doorway to understanding seasonal reproduction in mammals[J].Frontiers in Endocrinology,2016,7(9):36-45.
[24]SURBHI,KUMAR V.Avian photoreceptors and their role in the regulation of daily and seasonal physiology[J].General&Comparative Endocrinology,2015,220:13-22.
[25]DUPRE S M,MIEDZINSKA K,DUVAL C V,et al.Identification of EYA3and TAC1as long-day signals in the sheep pituitary[J].Current Biology,2010,20(9):829-835.
[26]MASUMOTO K,UKAITADENUMA M,KASUKAWA T,et al.Acute induction of EYA3by late-night light stimulation triggers TSHβexpression in photoperiodism[J].Current Biology,2010,20(24):2 199-2 206.
[27]曹晓涵,刘秋月,王翔宇,等.小尾寒羊黄体期和卵泡期生殖轴凋亡基因的表达研究及其与发情的关系[J].畜牧兽医学报,2017,48(11):2 091-2 097.
[28]董新龙,胡文萍,贺小云,等.绵羊BMP15组织表达特征及FecXGr、FecXO和G971A突变的检测[J].农业生物技术学报,2016,24(12):1 810-1 819.
[29]汤继顺,狄冉,刘秋月,等.精卵融合相关基因CD9和CD81在绵羊组织中的表达分析[J].农业生物技术学报,2018,26(1):87-95.
[30]SCHMITTGEN T D,LIVAK K J.Analyzing real-time PCR data by the comparative C(T)method[J].Nature Protocols,2008,3(6):1 101-1 108.
[31]LEWIS P D.A review of lighting for broiler breeders[J].British Poultry Science,2006,47(4):393-404.
[32]DARDENTE H.Circannual biology:the double life of the seasonal thyrotroph[J].Current Biology,2015,25(20):988-991.
[33]TAMAI T K,YOSHIMURA T.Molecular and neuroendocrine mechanisms of avian seasonal reproduction[J].Advances in Experimental Medicine and Biology,2017,1001:125-136.
[34]DARDENTE H,WYSE C A,BIMIE M J,et al.A molecular switch for photoperiod responsiveness in mammals[J].Current Biology,2010,20(24):2 193-2 198.
[35]DARDENTE H,LOMET D,ROBERT V,et al.Seasonal breeding in mammals:from basic science to applications and back[J].Theriogenology,2016,86(1):324-332.
[36]WOOD S H,CHRISTIAN H C,MIEDZINSKA K,et al.Binary switching of calendar cells in the pituitary defines the phase of the circannual cycle in mammals[J].Current Biology,2015,25(20):2 651-2 662.
[37]夏青,张效生,刘秋月,等.EYA3和TSHβ在季节性发情和常年发情绵羊中的表达模式[J].畜牧兽医学报,2018,49(2):263-269.