MITF基因表达与绵羊季节性发情及产羔数关系初探
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摘要
为初步探究MITF基因表达和绵羊季节性发情及产羔数的关系,本试验以季节性发情的苏尼特羊(Sunite sheep,SNT)和常年发情的小尾寒羊(Small Tail Han sheep,STH)为研究对象,利用实时荧光定量PCR(qPCR)技术分析SNT在不同光照条件下(短光照、长光照)及STH不同繁殖时期(卵泡期、黄体期)的下丘脑等组织中MITF基因的表达差异及该基因在STH单、多羔母羊卵泡期繁殖器官(卵巢、子宫体、输卵管)中的表达差异。结果表明:1)MITF基因在2个绵羊品种的不同组织中广泛表达且繁殖器官(卵巢、子宫体和输卵管)中该基因表达量较高;2)在长光照条件下SNT垂体中MITF表达量极显著高于短光照条件下表达量(P<0.01);3)STH卵巢中MITF表达量在卵泡期极显著高于黄体期(P<0.01),而子宫体、输卵管中该基因表达量在黄体期显著(P<0.05)或极显著(P<0.01)高于卵泡期;4)单羔组STH子宫体、输卵管中MITF表达水平极显著(P<0.01)或显著(P<0.05)高于多羔组。综上,MITF基因表达与绵羊季节性发情及产羔数存在一定关联,需要进一步进行生物学功能验证。
Aiming to tentatively explore the relationship between MITF gene expression and seasonal estrus and litter size in sheep,the real-time fluorescent quantitative PCR(qPCR)method was used to detect the expression difference of MITF gene in nine tissues including hypothalamus in seasonal estrous sheep(Sunite sheep,SNT)at different photoperiods(short photoperiod,long photoperiod)and year-round estrous sheep(Small Tail Han sheep,STH)at different reproduction stages(follicular stage and luteal stage),and the expression difference of MITF gene between single birth tissues and multiple birth tissues including ovary,uterine body and oviduct of STH at follicular stage.Theresults showed that:The MITF gene was widely expressed in different tissues and its' expression level in ovary,uterine body and oviduct tissues were higher than that in other tissues of two breeds.The expression level of MITF gene in pituitary gland was extremely higher at long photoperiod than that at short photoperiod in SNT(P<0.01).The expression level of MITFin ovary was extremely higher at follicular stage than that at luteal stage(P<0.01),while its expression level in uterine body was significantly higher at luteal stage than that at follicular stage(P<0.05)and its expression in oviduct was extremely higher at luteal stage than that at follicular stage(P<0.01)in STH.The expression level of MITF gene in uterine body in single birth group was extremely higher than that in multiple birth group(P<0.01),while its' expression level in oviduct in single birth group was significantly higher than that in multiple birth group(P<0.05)of STH at follicular stage.In conclusion,the research results indicated that MITF gene expression may be related to sheep seasonal estrus and litter size and further biological function verification will be required.
Aiming to tentatively explore the relationship between MITF gene expression and seasonal estrus and litter size in sheep,the real-time fluorescent quantitative PCR(qPCR)method was used to detect the expression difference of MITF gene in nine tissues including hypothalamus in seasonal estrous sheep(Sunite sheep,SNT)at different photoperiods(short photoperiod,long photoperiod)and year-round estrous sheep(Small Tail Han sheep,STH)at different reproduction stages(follicular stage and luteal stage),and the expression difference of MITF gene between single birth tissues and multiple birth tissues including ovary,uterine body and oviduct of STH at follicular stage.Theresults showed that:The MITF gene was widely expressed in different tissues and its' expression level in ovary,uterine body and oviduct tissues were higher than that in other tissues of two breeds.The expression level of MITF gene in pituitary gland was extremely higher at long photoperiod than that at short photoperiod in SNT(P<0.01).The expression level of MITFin ovary was extremely higher at follicular stage than that at luteal stage(P<0.01),while its expression level in uterine body was significantly higher at luteal stage than that at follicular stage(P<0.05)and its expression in oviduct was extremely higher at luteal stage than that at follicular stage(P<0.01)in STH.The expression level of MITF gene in uterine body in single birth group was extremely higher than that in multiple birth group(P<0.01),while its' expression level in oviduct in single birth group was significantly higher than that in multiple birth group(P<0.05)of STH at follicular stage.In conclusion,the research results indicated that MITF gene expression may be related to sheep seasonal estrus and litter size and further biological function verification will be required.
引文
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[2]黄冬维,储明星,狄冉,刘秋月,胡文萍,王翔宇,潘章源,郭晓飞.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):924-931Huang D W,Chu M X,Di R,Liu Q Y,Hu W P,Wang X Y,Pan Z Y,Guo X F.Study on expression of KiSS-1and PFRP genes related to reproductive seasonality in goats[J].Chinese Journal of Animal and Veterinary Sciences,2015,46(6):924-931(in Chinese)
[3] Dallmann R,Brown S A,Gachon F.Chronopharmacology:New insights and therapeutic implications[J].Annual Review of Pharmacology,2014,54(1):339-361
[4] Montgomery G W,Crawford A M,Penty J M,Dodds K G,Ede A J,Henry H M,Pierson C A,Lord E A,Galloway S M,Schmack A E.The ovine Booroola fecundity gene(FecB)is linked to markers from a region of human chromosome 4q[J].Nature Genetics,1993,4(4):410-414
[5] Souza C J,Campbell B K,Webb R,Baird D T.Secretion of inhibin A and follicular dynamics throughout the estrous cycle in the sheep with and without the Booroola gene(FecB)[J].Endocrinology,1997,138(12):5333-5340
[6] Galloway S M,Mcnatty K P,Cambridge L M,Laitinen M P,Juenge J L,Jokiranta T S,McLaren R J,Luiro K,Dodds K G,Montgomery G W,Beattie A E,Davis G H,Ritvos O.Mutations in an oocyte-derived growth factor gene(BMP15)cause increased ovulation rate and infertility in a dosagesensitive manner[J].Nature Genetics,2000,25(3):279-283
[7] Dong J,Albertini D F,Nishimori K,Kumar T R,Lu N,Matzuk M M.Growth differentiation factor-9is required during early ovarian folliculogenesis[J].Nature,1996,383(6600):531-535
[8] Bodensteiner K J,Clay C M,Moeller C L,Sawyer H R.Molecular cloning of the ovine growth/differentiation factor-9gene and expression of growth/differentiation factor-9in ovine and bovine ovaries[J].Biology of Reproduction,1999,60(2):381-386
[9] Xia Q Y,Wang X T,Ye S B,Wang X,Li R,Shi S S,Fang R,Zhang R S,Ma H H,Lu Z F,Shen Q,Bao W,Zhou X J,Rao Q.Novel gene fusion of PRCC-MITF defines a new member of MiT family translocation renal cell carcinoma:Clinicopathological analysis and detection of the gene fusion by RNA sequencing and fish[J].Histopathology,2018,72(5):786-794
[10] Meadows N A,Sharma S M,Faulkner G J,Ostrowski M C,Hume D A,Cassady A I.The expression of Clcn7 and Ostm1 in osteoclasts is coregulated by microphthalmia transcription factor[J].Journal of Biological Chemistry,2007,282(3):1891-1904
[11] Martina J A,Diab H I,Li H Q,Puertollano R.Novel roles for the MITF/TFE family of transcription factors in organelle biogenesis,nutrient sensing,and energy homeostasis[J].Cellular and Molecular Life Sciences,2014,71(13):2483-2497
[12] Hodgkinson C A,Moore K J,Nakayama A,Steingrímsson E,Copeland N G,Jenkins N A,Arnheiter H.Mutations at the mouse microphthalmia locus are associated with defects in agene encoding a novel basic-helix-loop-helix-zipper protein[J].Cell,1993,74(2):395-404
[13]Watanabe H,Tatsumi K,Yokoi H,Higuchi T,Iwai M,Fukuoka M,Fujiwara H,Fujita K,Nakayama H,Mori T,Fujita J.Ovulation defect and its restoration by bone marrow transplantation in osteopetrotic mutant mice of Mitf(mi)/Mitf(mi)genotype[J].Biology of Reproduction,1997,57(6):1394-1400
[14] Vachtenheim J,Borovansky J.Transcription physiology of pigment formation in melanocytes:Central role of MITF[J].Experimental Dermatology,2010,19(7):617-627
[15] Monika R, Arne L.Pleiotropic effects of coat colourassociated mutations in humans,mice and other mammals[J].Seminars in Cell&Developmental Biology,2013,24(6-7):576-586
[16] Bauer G L,Praetorius C,Bergsteinsdóttir K,Hallsson J H,Gísladóttir B K,Schepsky A,Swing D A,Osullivan T N,Arnheiter H,Bismuth K,Debbache J,Fletcher C,Warming S,Copeland N G,Jenkins N A,Steingrímsson E.The role of MITF phosphorylation sites during coat color and eye development inmice analyzed bybacterialartificial chromosome transgene rescue[J].Genetics,2009,183(2):581-594
[17] Bharti K,Liu W,Csermely T,Bertuzzi S,Arnheiter H.Alternative promoter use in eye development:The complex role and regulation of the transcription factor MITF[J].Development,2008,135(6):1169-1178
[18] Lu S Y,Wan H C,Li M,Lin Y L.Subcellular localization of MITF in monocytic cells[J].Histochemistry&Cell Biology,2010,133(6):651-658
[19]曹晓涵,刘秋月,王翔宇,狄冉,胡文萍,张效生,张金龙,曾宪垠,储明星.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):2091-2097Cao X H,Liu Q Y,Wang X Y,Di R,Hu W P,Zhang X S,Zhang J L,Zeng X Y,Chu M X.Expression levels of apoptotic gene in HPG axis during luteal and follicular periods and their association with estrus in small-tail Han sheep[J].Chinese Journal of Animal and Veterinary Sciences,2017,48(11):2091-2097(in Chinese)
[20]董新龙,胡文萍,贺小云,郭晓飞,潘章源,狄冉,刘秋月,王翔宇,曹晓涵,王金玉,储明星.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):1810-1819Dong X L,Hu W P,He X Y,Guo X F,Pan Z Y,Di R,Liu Q Y,Wang X Y,Cao X H,Wang J Y,Chu M X.Tissue expression and detection of the FecXGr,FecXO and G971A mutations of BMP15 gene in sheep[J]. Journal of Agricultural Biotechnology,2016,24(12):1810-1819(in Chinese)
[21]周梅,狄冉,胡文萍,王翔宇,张效生,张金龙,刘秋月,储明星.小尾寒羊和苏尼特羊CRH和POMC基因在繁殖相关组织中的差异性表达[J].中国草食动物科学,2017,37(04):1-4Zhou M,Di R,Hu W P,Wang X Y,Zhang X S,Zhang J L,Liu Q Y,Chu M X.Different expression of CRHand POMC genes in reproductive related tissues in small-tail Han sheep and Sunite sheep[J].China Herbivore Science,2017,37(04):1-4(in Chinese)
[22]汤继顺,狄冉,刘秋月,王翔宇,张效生,张金龙,胡文萍,储明星.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):87-95Tang J S,Di R,Liu Q Y,Wang X Y,Zhang X S,Zhang J L,Hu W P,Chu M X.Tissue expression analysis of CD9and CD81genes associated sperm-egg fusion in sheep[J].Journal of Agricultural Biotechnology,2018,26(1):87-95(in Chinese)
[23] Schmittgen T D,Livak K J.Analyzing real-time PCR data by the comparative C(T)method[J].Nature Protocols,2008,3(6):1101-1108
[24] Dardente H.Circannual biology:The double life of the seasonal thyrotroph[J].Current Biology,2015,25(20):988-991
[25] Tamai T K,Yoshimura T.Molecular and neuroendocrine mechanisms of avian seasonal reproduction[J].Advances in Experimental Medicine&Biology,2017,1001:125-136
[26] Proudman J A,Clerens S,Van D B G,Garrett W M,Verhaert P D,Vandesande F,Berghman L R.Immunohistochemical localization of chromogranin A in gonadotrophs and somatotrophs of the turkey and chicken pituitary[J].General&Comparative Endocrinology,2003,132(2):293-303
[27] Dardente H,Wyse C A,Birnie M J,DupréS M,Loudon A S,Lincoln G A,Hazlerigg D G.A molecular switch for photoperiod responsiveness in mammals[J]. Current Biology,2010,20(24):2193-2198
[28] Masumoto K,Ukaitadenuma M,Kasukawa T,Nagano M,Uno K D,Tsujino K,Horikawa K,Shigeyoshi Y,Ueda H R.Acute induction of Eya3by late-night light stimulation triggers TSHβexpression in photoperiodism[J].Current Biology,2010,20(24):2199-2206
[29] Dardente H,Lomet D,Robert V,Decourt C,Beltramo M,Pellicer-Rubio M T.Seasonal breeding in mammals:From basic science to applications and back[J].Theriogenology,2016,86(1):324-332
[30]夏青,张效生,刘秋月,王翔宇,贺小云,郭晓飞,胡文萍,张金龙,储明星,狄冉.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):263-269Xia Q,Zhang X S,Liu Q Y,Wang X Y,He X Y,Guo X F,Hu W P,Zhang J L,Chu M X,Di R.Expression patterns of Eya3and TSHβin seasonal estrus and year-round estrus sheep[J].Chinese Journal of Animal and Veterinary Sciences,2018,49(2):263-269(in Chinese)
[31] Arena G,Valente E M.PINK1in the limelight:Multiple functions of an eclectic protein in human health and disease[J].Journal of Pathology,2017,241(2):251-263
[32] Yamano K,Matsuda N,Tanaka K.The ubiquitin signal and autophagy:An orchestrated dance leading to mitochondrial degradation[J].Embo Reports,2016,17(3):300-316
[33] Ziskind G,Paltieli Y,Eibschitz I,Ohel G,Weichselbaum A.The effect of human fallopian tube epithelium on human sperm velocity motility and binding[J].Journal of Assisted Reproduction and Genetics,2000,17(3):147-150
[2]黄冬维,储明星,狄冉,刘秋月,胡文萍,王翔宇,潘章源,郭晓飞.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):924-931Huang D W,Chu M X,Di R,Liu Q Y,Hu W P,Wang X Y,Pan Z Y,Guo X F.Study on expression of KiSS-1and PFRP genes related to reproductive seasonality in goats[J].Chinese Journal of Animal and Veterinary Sciences,2015,46(6):924-931(in Chinese)
[3] Dallmann R,Brown S A,Gachon F.Chronopharmacology:New insights and therapeutic implications[J].Annual Review of Pharmacology,2014,54(1):339-361
[4] Montgomery G W,Crawford A M,Penty J M,Dodds K G,Ede A J,Henry H M,Pierson C A,Lord E A,Galloway S M,Schmack A E.The ovine Booroola fecundity gene(FecB)is linked to markers from a region of human chromosome 4q[J].Nature Genetics,1993,4(4):410-414
[5] Souza C J,Campbell B K,Webb R,Baird D T.Secretion of inhibin A and follicular dynamics throughout the estrous cycle in the sheep with and without the Booroola gene(FecB)[J].Endocrinology,1997,138(12):5333-5340
[6] Galloway S M,Mcnatty K P,Cambridge L M,Laitinen M P,Juenge J L,Jokiranta T S,McLaren R J,Luiro K,Dodds K G,Montgomery G W,Beattie A E,Davis G H,Ritvos O.Mutations in an oocyte-derived growth factor gene(BMP15)cause increased ovulation rate and infertility in a dosagesensitive manner[J].Nature Genetics,2000,25(3):279-283
[7] Dong J,Albertini D F,Nishimori K,Kumar T R,Lu N,Matzuk M M.Growth differentiation factor-9is required during early ovarian folliculogenesis[J].Nature,1996,383(6600):531-535
[8] Bodensteiner K J,Clay C M,Moeller C L,Sawyer H R.Molecular cloning of the ovine growth/differentiation factor-9gene and expression of growth/differentiation factor-9in ovine and bovine ovaries[J].Biology of Reproduction,1999,60(2):381-386
[9] Xia Q Y,Wang X T,Ye S B,Wang X,Li R,Shi S S,Fang R,Zhang R S,Ma H H,Lu Z F,Shen Q,Bao W,Zhou X J,Rao Q.Novel gene fusion of PRCC-MITF defines a new member of MiT family translocation renal cell carcinoma:Clinicopathological analysis and detection of the gene fusion by RNA sequencing and fish[J].Histopathology,2018,72(5):786-794
[10] Meadows N A,Sharma S M,Faulkner G J,Ostrowski M C,Hume D A,Cassady A I.The expression of Clcn7 and Ostm1 in osteoclasts is coregulated by microphthalmia transcription factor[J].Journal of Biological Chemistry,2007,282(3):1891-1904
[11] Martina J A,Diab H I,Li H Q,Puertollano R.Novel roles for the MITF/TFE family of transcription factors in organelle biogenesis,nutrient sensing,and energy homeostasis[J].Cellular and Molecular Life Sciences,2014,71(13):2483-2497
[12] Hodgkinson C A,Moore K J,Nakayama A,Steingrímsson E,Copeland N G,Jenkins N A,Arnheiter H.Mutations at the mouse microphthalmia locus are associated with defects in agene encoding a novel basic-helix-loop-helix-zipper protein[J].Cell,1993,74(2):395-404
[13]Watanabe H,Tatsumi K,Yokoi H,Higuchi T,Iwai M,Fukuoka M,Fujiwara H,Fujita K,Nakayama H,Mori T,Fujita J.Ovulation defect and its restoration by bone marrow transplantation in osteopetrotic mutant mice of Mitf(mi)/Mitf(mi)genotype[J].Biology of Reproduction,1997,57(6):1394-1400
[14] Vachtenheim J,Borovansky J.Transcription physiology of pigment formation in melanocytes:Central role of MITF[J].Experimental Dermatology,2010,19(7):617-627
[15] Monika R, Arne L.Pleiotropic effects of coat colourassociated mutations in humans,mice and other mammals[J].Seminars in Cell&Developmental Biology,2013,24(6-7):576-586
[16] Bauer G L,Praetorius C,Bergsteinsdóttir K,Hallsson J H,Gísladóttir B K,Schepsky A,Swing D A,Osullivan T N,Arnheiter H,Bismuth K,Debbache J,Fletcher C,Warming S,Copeland N G,Jenkins N A,Steingrímsson E.The role of MITF phosphorylation sites during coat color and eye development inmice analyzed bybacterialartificial chromosome transgene rescue[J].Genetics,2009,183(2):581-594
[17] Bharti K,Liu W,Csermely T,Bertuzzi S,Arnheiter H.Alternative promoter use in eye development:The complex role and regulation of the transcription factor MITF[J].Development,2008,135(6):1169-1178
[18] Lu S Y,Wan H C,Li M,Lin Y L.Subcellular localization of MITF in monocytic cells[J].Histochemistry&Cell Biology,2010,133(6):651-658
[19]曹晓涵,刘秋月,王翔宇,狄冉,胡文萍,张效生,张金龙,曾宪垠,储明星.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):2091-2097Cao X H,Liu Q Y,Wang X Y,Di R,Hu W P,Zhang X S,Zhang J L,Zeng X Y,Chu M X.Expression levels of apoptotic gene in HPG axis during luteal and follicular periods and their association with estrus in small-tail Han sheep[J].Chinese Journal of Animal and Veterinary Sciences,2017,48(11):2091-2097(in Chinese)
[20]董新龙,胡文萍,贺小云,郭晓飞,潘章源,狄冉,刘秋月,王翔宇,曹晓涵,王金玉,储明星.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):1810-1819Dong X L,Hu W P,He X Y,Guo X F,Pan Z Y,Di R,Liu Q Y,Wang X Y,Cao X H,Wang J Y,Chu M X.Tissue expression and detection of the FecXGr,FecXO and G971A mutations of BMP15 gene in sheep[J]. Journal of Agricultural Biotechnology,2016,24(12):1810-1819(in Chinese)
[21]周梅,狄冉,胡文萍,王翔宇,张效生,张金龙,刘秋月,储明星.小尾寒羊和苏尼特羊CRH和POMC基因在繁殖相关组织中的差异性表达[J].中国草食动物科学,2017,37(04):1-4Zhou M,Di R,Hu W P,Wang X Y,Zhang X S,Zhang J L,Liu Q Y,Chu M X.Different expression of CRHand POMC genes in reproductive related tissues in small-tail Han sheep and Sunite sheep[J].China Herbivore Science,2017,37(04):1-4(in Chinese)
[22]汤继顺,狄冉,刘秋月,王翔宇,张效生,张金龙,胡文萍,储明星.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):87-95Tang J S,Di R,Liu Q Y,Wang X Y,Zhang X S,Zhang J L,Hu W P,Chu M X.Tissue expression analysis of CD9and CD81genes associated sperm-egg fusion in sheep[J].Journal of Agricultural Biotechnology,2018,26(1):87-95(in Chinese)
[23] Schmittgen T D,Livak K J.Analyzing real-time PCR data by the comparative C(T)method[J].Nature Protocols,2008,3(6):1101-1108
[24] Dardente H.Circannual biology:The double life of the seasonal thyrotroph[J].Current Biology,2015,25(20):988-991
[25] Tamai T K,Yoshimura T.Molecular and neuroendocrine mechanisms of avian seasonal reproduction[J].Advances in Experimental Medicine&Biology,2017,1001:125-136
[26] Proudman J A,Clerens S,Van D B G,Garrett W M,Verhaert P D,Vandesande F,Berghman L R.Immunohistochemical localization of chromogranin A in gonadotrophs and somatotrophs of the turkey and chicken pituitary[J].General&Comparative Endocrinology,2003,132(2):293-303
[27] Dardente H,Wyse C A,Birnie M J,DupréS M,Loudon A S,Lincoln G A,Hazlerigg D G.A molecular switch for photoperiod responsiveness in mammals[J]. Current Biology,2010,20(24):2193-2198
[28] Masumoto K,Ukaitadenuma M,Kasukawa T,Nagano M,Uno K D,Tsujino K,Horikawa K,Shigeyoshi Y,Ueda H R.Acute induction of Eya3by late-night light stimulation triggers TSHβexpression in photoperiodism[J].Current Biology,2010,20(24):2199-2206
[29] Dardente H,Lomet D,Robert V,Decourt C,Beltramo M,Pellicer-Rubio M T.Seasonal breeding in mammals:From basic science to applications and back[J].Theriogenology,2016,86(1):324-332
[30]夏青,张效生,刘秋月,王翔宇,贺小云,郭晓飞,胡文萍,张金龙,储明星,狄冉.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):263-269Xia Q,Zhang X S,Liu Q Y,Wang X Y,He X Y,Guo X F,Hu W P,Zhang J L,Chu M X,Di R.Expression patterns of Eya3and TSHβin seasonal estrus and year-round estrus sheep[J].Chinese Journal of Animal and Veterinary Sciences,2018,49(2):263-269(in Chinese)
[31] Arena G,Valente E M.PINK1in the limelight:Multiple functions of an eclectic protein in human health and disease[J].Journal of Pathology,2017,241(2):251-263
[32] Yamano K,Matsuda N,Tanaka K.The ubiquitin signal and autophagy:An orchestrated dance leading to mitochondrial degradation[J].Embo Reports,2016,17(3):300-316
[33] Ziskind G,Paltieli Y,Eibschitz I,Ohel G,Weichselbaum A.The effect of human fallopian tube epithelium on human sperm velocity motility and binding[J].Journal of Assisted Reproduction and Genetics,2000,17(3):147-150
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