褪黑激素对内蒙古绒山羊皮肤中let-7家族及其靶基因的影响
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摘要
旨在探索褪黑激素(melatonin, MT)对let-7家族成员及其靶基因在内蒙古绒山羊皮肤周期性表达的影响。本研究选取内蒙古绒山羊罕山型个体6只,分为埋植组(皮下埋植MT,n=3)和对照组(不埋植MT,n=3),连续12个月采集绒山羊个体皮肤组织为试验材料,利用qRT-PCR技术检测皮肤组织中11个let-7家族成员的表达变化,结合RNA-Seq技术分析let-7家族靶基因的表达变化。结果表明:1)在整个皮肤毛囊周期内,MT上调let-7b、let-7e、let-7a-3p的表达,下调let-7f、let-7g、let-7c、let-7a-5p、let-7d、miR-98的表达。2)在4月份,MT上调"let-7g、let-7i、miR-98"的表达,下调PTGS2、PLCG1等靶基因的表达;下调"let-7a-5p、let-7b、let-7c"的表达,上调TGIF2、RBL1等靶基因的表达。3)在6月份,MT下调"let-7g、let-7i、miR-98"的表达,上调PTGS2、PLCG1、SRC等靶基因的表达;上调"let-7a-5p、let-7b、let-7c"的表达,下调GDF5、TGIF2、RBL1等靶基因的表达。4)let-7家族成员的靶基因主要参与VEGF、TGF-β、Oxytocin和NF-kappa B信号通路,在细胞过程、激素调节和转录因子调控方面发挥重要作用。综上表明,MT可能通过改变let-7家族重要成员的表达水平,影响相应靶基因的表达,进而在皮肤毛囊的周期性生长过程中发挥重要作用,研究结果为进一步揭示MT介导miRNAs影响羊绒生长的分子机理提供了理论依据。
The study aimed to explore the effects of melatonin on the periodic expression of let-7 family and its target genes in skin of Inner Mongolia Cashmere goat. In this study, six Inner Mongolia Hanshan Cashmere goats were selected and divided equally and randomly into 2 groups, melatonin implantation group and non-implantation group. Taking the skin tissue of the goats as experimental material in every months of the year. The expression changes of 11 members in let-7 family were quantified by reverse transcription-real time polymerase chain reaction(qRT-PCR) in the skin tissue. The expression changes of the target genes of let-7 family were analyzed using the RNA-Seq technique. The results showed that: 1) During the whole hair follicle cycle, melatonin up-regulated the expression of let-7 b, let-7 e and let-7 a-3 p, down-regulated the expression of let-7 f, let-7 g, let-7 c, let-7 a-5 p, let-7 d and miR-98. 2) In April, melatonin up-regulated the expression of "let-7 g, let-7 i and miR-98", down-regulated the expression of target genes such as PTGS2 and PLCG1. It down-regulated the expression of "let-7 a-5 p, let-7 b and let-7 c", up-regulated the expression of target genes such as TGIF2 and RBL1. 3) In June, melatonin down-regulated the expression of "let-7 g, let-7 i and miR-98", up-regulated the expression of target genes such as PTGS2, PLCG1 and SRC; It up-regulated the expression of "let-7 a-5 p, let-7 b and let-7 c", down-regulated the expression of target genes such as GDF5, TGIF2 and RBL1. 4) The target genes of let-7 family were mainly involved in VEGF, TGF-β, Oxytocin and NF-kappa B signaling pathways, and played an important role in cell processes, hormone regulation and regulation of transcription factors. In summary, MT may affect the expression of the corresponding target genes by changing the expression level of members in the let-7 family. It plays an important role in the periodic growth of skin hair follicles. These results provided a theoretical basis for further revealing the molecular mechanism of melatonin mediating miRNAs to affect cashmere growth.
The study aimed to explore the effects of melatonin on the periodic expression of let-7 family and its target genes in skin of Inner Mongolia Cashmere goat. In this study, six Inner Mongolia Hanshan Cashmere goats were selected and divided equally and randomly into 2 groups, melatonin implantation group and non-implantation group. Taking the skin tissue of the goats as experimental material in every months of the year. The expression changes of 11 members in let-7 family were quantified by reverse transcription-real time polymerase chain reaction(qRT-PCR) in the skin tissue. The expression changes of the target genes of let-7 family were analyzed using the RNA-Seq technique. The results showed that: 1) During the whole hair follicle cycle, melatonin up-regulated the expression of let-7 b, let-7 e and let-7 a-3 p, down-regulated the expression of let-7 f, let-7 g, let-7 c, let-7 a-5 p, let-7 d and miR-98. 2) In April, melatonin up-regulated the expression of "let-7 g, let-7 i and miR-98", down-regulated the expression of target genes such as PTGS2 and PLCG1. It down-regulated the expression of "let-7 a-5 p, let-7 b and let-7 c", up-regulated the expression of target genes such as TGIF2 and RBL1. 3) In June, melatonin down-regulated the expression of "let-7 g, let-7 i and miR-98", up-regulated the expression of target genes such as PTGS2, PLCG1 and SRC; It up-regulated the expression of "let-7 a-5 p, let-7 b and let-7 c", down-regulated the expression of target genes such as GDF5, TGIF2 and RBL1. 4) The target genes of let-7 family were mainly involved in VEGF, TGF-β, Oxytocin and NF-kappa B signaling pathways, and played an important role in cell processes, hormone regulation and regulation of transcription factors. In summary, MT may affect the expression of the corresponding target genes by changing the expression level of members in the let-7 family. It plays an important role in the periodic growth of skin hair follicles. These results provided a theoretical basis for further revealing the molecular mechanism of melatonin mediating miRNAs to affect cashmere growth.
引文
[1] FISCHER T W.The influence of melatonin on hair physiology[J].Hautarzt,2009,60(12):962-972.
[2] 柳建昌,尹协镇,方天祺.褪黑素对中国绒山羊在非生绒期促绒生长与绒产量的影响[J].动物学杂志,1998,33(3):8-12.LIU J C,YIN X Z,FANG T Q.Effect of melatonin on the cashmere growth and production of Chinese white goats[J].Chinese Journal of Zoology,1998,33(3):8-12.(in Chinese)
[3] FISCHER T W,SLOMINSKI A,TOBIN D J,et al.Melatonin and the hair follicle[J].J Pineal Res,2008,44(1):1-15.
[4] FISCHER T W,SWEATMAN T W,SEMAK I,et al.Constitutive and UV-induced metabolism of melatonin in keratinocytes and cell-free systems[J].FASEB J,2006,20(9):1564-1566.
[5] SLOMINSKI A,PISARCHIK A,ZBYTEK B,et al.Functional activity of serotoninergic and melatoninergic systems expressed in the skin[J].J Cell Physiol,2003,196(1):144-153.
[6] 付绍印,赵宏丽,郑竹清,等.褪黑激素对绒山羊皮肤中毛囊周期相关miRNAs表达模式的影响[J].遗传,2014,36(12):1235-1242.FU S Y,ZHAO H L,ZHENG Z Q,et al.Melatonin regulating the expression of miRNAs involved in hair follicle cycle of cashmere goats skin[J].Hereditas (Beijing),2014,36(12):1235-1242.(in Chinese)
[7] REINHART B J,SLACK F J,BASSON M,et al.The 21-nucleotide let-7 RNA regulates developmental timing in Caenorhabditis elegans[J].Nature,2000,403(6772):901-906.
[8] ROUSH S,SLACK F J.The let-7 family of microRNAs[J].Trends Cell Biol,2008,18(10):505-516.
[9] ZHANG W Q,WU J H,LI J Q,et al.A subset of skin-expressed microRNAs with possible roles in goat and sheep hair growth based on expression profiling of mammalian microRNAs[J].OMICS,2007,11(4):385-396.
[10] WANG X R,CAO L,WANG Y Y,et al.Regulation of let-7 and its target oncogenes (Review)[J].Oncol Lett,2012,3(5):955-960.
[11] POWERS J T,TSANOV K M,PEARSON D S,et al.Multiple mechanisms disrupt the let-7 microRNA family in neuroblastoma[J].Nature,2016,535(7611):246-251.
[12] 张璐,张燕军,苏蕊,等.microRNA对皮肤毛囊发育的调控机制[J].遗传,2014,36(7):655-660.ZHANG L,ZHANG Y J,SU R,et al.The regulatory mechanism of microRNAs in skin and hair follicle development[J].Hereditas(Beijing),2014,36(7):655-660.(in Chinese)
[13] 江倩,李建平,曲海娥,等.miR-let7a及其靶基因IGF-1R在绒山羊毛囊发育周期中的表达[J].中国兽医学报,2014,34(10):1622-1626.JIANG Q,LI J P,QU H E,et al.micro-RNA-let7a and targeting IGF-1R controls hair cycle-associated changes in gene expression programs of the cashmere goats[J].Chinese Journal of Veterinary Science,2014,34(10):1622-1626.(in Chinese)
[14] WU Y,ZHONG J L,HOU N,et al.microRNA Let-7b inhibits keratinocyte migration in cutaneous wound healing by targeting IGF2BP2[J].Exp Dermatol,2017,26(2):116-123.
[15] GAO X R,GE J,LI W Y,et al.NF-κB/let-7f-5p/IL-10 pathway involves in wear particle-induced osteolysis by inducing M1 macrophage polarization[J].Cell Cycle,2018,17(17):2134-2145.
[16] MA?ACHOWSKA B,WYKA K,NOWICKA Z,et al.Temporal dynamics of serum let-7g expression mirror the decline of residual beta-cell function in longitudinal observation of children with type 1 diabetes[J].Pediatr Diabetes,2018,19(8):1407-1415.
[17] ZHANG W,ZENG Q R,BAN Z Y,et al.Effects of let-7c on the proliferation of ovarian carcinoma cells by targeted regulation of CDC25a gene expression[J].Oncol Lett,2018,16(5):5543-5550.
[18] ASAMA H,SUZUKI R,HIKICHI T,et al.microRNA let-7d targets thrombospondin-1 and inhibits the activation of human pancreatic stellate cells[J].Pancreatology,2019,19(1):196-203.
[19] LI Z J,PAN W H,SHEN Y,et al.IGF1/IGF1R and microRNA let-7e down-regulate each other and modulate proliferation and migration of colorectal cancer cells[J].Cell Cycle,2018,17(10):1212-1219.
[20] NGUYEN T,SU C,SINGH M.Let-7i inhibition enhances progesterone-induced functional recovery in a mouse model of ischemia[J].Proc Natl Acad Sci U S A,2018,115(41):E9668-E9677.
[21] GENG C,DONG T Y,JIN W L,et al.microRNA-98 regulates hepatic cholesterol metabolism via targeting sterol regulatory element-binding protein 2[J].Biochem Biophys Res Commun,2018,504(2):422-426.
[22] FATIMA A,LYNN D J,O’BOYLE P,et al.The miRNAome of the postpartum dairy cow liver in negative energy balance[J].BMC Genomics,2014,15:279.
[23] 刘思齐.内蒙古绒山羊皮肤毛囊let-7家族表达研究[D].呼和浩特:内蒙古农业大学,2015.LIU S Q.Expression profile of let-7 in skin hair follicle of neimongol cashmere goat[D].Hohhot:Inner Mongolia Agricultural University,2015.(in Chinese)
[24] 刘斌,杨军,阿云嘎,等.外源褪黑激素促进绒山羊皮肤毛囊相关基因表达差异的研究[J].畜牧兽医学报,2016,47(11):2210-2217.LIU B,YANG J,AI Y G,et al.Exogenous melatonin promotes expression differences of skin hair follicles related genes in cashmere goat[J].Acta Veterinaria et Zootechnica Sinica,2016,47(11):2210-2217.(in Chinese)
[25] 赵艳红,王琳,张铁佳,等.褪黑激素促进绒山羊绒毛生长及其作用机制的研究机制[J].中国畜牧兽医,2014,41(4):215-219.ZHAO Y H,WANG L,ZHANG T J,et al.Research progress on promoting effect and mechanism of melatonin on cashmere growth in cashmere goat[J].China Animal Husbandry & Veterinary Medicine,2014,41(4):215-219.(in Chinese)
[26] WANG D,SUN X L,WEI Y,et al.Nuclear miR-122 directly regulates the biogenesis of cell survival oncomiR miR-21 at the posttranscriptional level[J].Nucleic Acids Res,2018,46(4):2012-2029.
[27] TANG R,LI L M,ZHU D H,et al.Mouse miRNA-709 directly regulates miRNA-15a/16-1 biogenesis at the posttranscriptional level in the nucleus:evidence for a microRNA hierarchy system[J].Cell Res,2012,22(3):504-515.
[28] 李晓燕,张璐,王乐乐,等.绒山羊次级毛囊生长调控基因的筛选[J].畜牧兽医学报,2015,46(10):1733-1740.LI X Y,ZHANG L,WANG L L,et al.Selection of relative gene during secondary hair follicle growth in cashmere goat[J].Acta Veterinaria et Zootechnica Sinica,2015,46(10):1733-1740.(in Chinese)
[29] 张春兰.内蒙古阿尔巴斯白绒山羊毛囊生长周期性变化与KAP6 cDNA的特征分析[D].呼和浩特:内蒙古农业大学,2003.ZHANG C L.Periodic variety of hair follicles about Inner Mongolia ARBAS white cashmere goats and characteristic of KAP6 cDNA[D].Hohhot:Inner Mongolia Agricultural University,2003.(in Chinese)
[2] 柳建昌,尹协镇,方天祺.褪黑素对中国绒山羊在非生绒期促绒生长与绒产量的影响[J].动物学杂志,1998,33(3):8-12.LIU J C,YIN X Z,FANG T Q.Effect of melatonin on the cashmere growth and production of Chinese white goats[J].Chinese Journal of Zoology,1998,33(3):8-12.(in Chinese)
[3] FISCHER T W,SLOMINSKI A,TOBIN D J,et al.Melatonin and the hair follicle[J].J Pineal Res,2008,44(1):1-15.
[4] FISCHER T W,SWEATMAN T W,SEMAK I,et al.Constitutive and UV-induced metabolism of melatonin in keratinocytes and cell-free systems[J].FASEB J,2006,20(9):1564-1566.
[5] SLOMINSKI A,PISARCHIK A,ZBYTEK B,et al.Functional activity of serotoninergic and melatoninergic systems expressed in the skin[J].J Cell Physiol,2003,196(1):144-153.
[6] 付绍印,赵宏丽,郑竹清,等.褪黑激素对绒山羊皮肤中毛囊周期相关miRNAs表达模式的影响[J].遗传,2014,36(12):1235-1242.FU S Y,ZHAO H L,ZHENG Z Q,et al.Melatonin regulating the expression of miRNAs involved in hair follicle cycle of cashmere goats skin[J].Hereditas (Beijing),2014,36(12):1235-1242.(in Chinese)
[7] REINHART B J,SLACK F J,BASSON M,et al.The 21-nucleotide let-7 RNA regulates developmental timing in Caenorhabditis elegans[J].Nature,2000,403(6772):901-906.
[8] ROUSH S,SLACK F J.The let-7 family of microRNAs[J].Trends Cell Biol,2008,18(10):505-516.
[9] ZHANG W Q,WU J H,LI J Q,et al.A subset of skin-expressed microRNAs with possible roles in goat and sheep hair growth based on expression profiling of mammalian microRNAs[J].OMICS,2007,11(4):385-396.
[10] WANG X R,CAO L,WANG Y Y,et al.Regulation of let-7 and its target oncogenes (Review)[J].Oncol Lett,2012,3(5):955-960.
[11] POWERS J T,TSANOV K M,PEARSON D S,et al.Multiple mechanisms disrupt the let-7 microRNA family in neuroblastoma[J].Nature,2016,535(7611):246-251.
[12] 张璐,张燕军,苏蕊,等.microRNA对皮肤毛囊发育的调控机制[J].遗传,2014,36(7):655-660.ZHANG L,ZHANG Y J,SU R,et al.The regulatory mechanism of microRNAs in skin and hair follicle development[J].Hereditas(Beijing),2014,36(7):655-660.(in Chinese)
[13] 江倩,李建平,曲海娥,等.miR-let7a及其靶基因IGF-1R在绒山羊毛囊发育周期中的表达[J].中国兽医学报,2014,34(10):1622-1626.JIANG Q,LI J P,QU H E,et al.micro-RNA-let7a and targeting IGF-1R controls hair cycle-associated changes in gene expression programs of the cashmere goats[J].Chinese Journal of Veterinary Science,2014,34(10):1622-1626.(in Chinese)
[14] WU Y,ZHONG J L,HOU N,et al.microRNA Let-7b inhibits keratinocyte migration in cutaneous wound healing by targeting IGF2BP2[J].Exp Dermatol,2017,26(2):116-123.
[15] GAO X R,GE J,LI W Y,et al.NF-κB/let-7f-5p/IL-10 pathway involves in wear particle-induced osteolysis by inducing M1 macrophage polarization[J].Cell Cycle,2018,17(17):2134-2145.
[16] MA?ACHOWSKA B,WYKA K,NOWICKA Z,et al.Temporal dynamics of serum let-7g expression mirror the decline of residual beta-cell function in longitudinal observation of children with type 1 diabetes[J].Pediatr Diabetes,2018,19(8):1407-1415.
[17] ZHANG W,ZENG Q R,BAN Z Y,et al.Effects of let-7c on the proliferation of ovarian carcinoma cells by targeted regulation of CDC25a gene expression[J].Oncol Lett,2018,16(5):5543-5550.
[18] ASAMA H,SUZUKI R,HIKICHI T,et al.microRNA let-7d targets thrombospondin-1 and inhibits the activation of human pancreatic stellate cells[J].Pancreatology,2019,19(1):196-203.
[19] LI Z J,PAN W H,SHEN Y,et al.IGF1/IGF1R and microRNA let-7e down-regulate each other and modulate proliferation and migration of colorectal cancer cells[J].Cell Cycle,2018,17(10):1212-1219.
[20] NGUYEN T,SU C,SINGH M.Let-7i inhibition enhances progesterone-induced functional recovery in a mouse model of ischemia[J].Proc Natl Acad Sci U S A,2018,115(41):E9668-E9677.
[21] GENG C,DONG T Y,JIN W L,et al.microRNA-98 regulates hepatic cholesterol metabolism via targeting sterol regulatory element-binding protein 2[J].Biochem Biophys Res Commun,2018,504(2):422-426.
[22] FATIMA A,LYNN D J,O’BOYLE P,et al.The miRNAome of the postpartum dairy cow liver in negative energy balance[J].BMC Genomics,2014,15:279.
[23] 刘思齐.内蒙古绒山羊皮肤毛囊let-7家族表达研究[D].呼和浩特:内蒙古农业大学,2015.LIU S Q.Expression profile of let-7 in skin hair follicle of neimongol cashmere goat[D].Hohhot:Inner Mongolia Agricultural University,2015.(in Chinese)
[24] 刘斌,杨军,阿云嘎,等.外源褪黑激素促进绒山羊皮肤毛囊相关基因表达差异的研究[J].畜牧兽医学报,2016,47(11):2210-2217.LIU B,YANG J,AI Y G,et al.Exogenous melatonin promotes expression differences of skin hair follicles related genes in cashmere goat[J].Acta Veterinaria et Zootechnica Sinica,2016,47(11):2210-2217.(in Chinese)
[25] 赵艳红,王琳,张铁佳,等.褪黑激素促进绒山羊绒毛生长及其作用机制的研究机制[J].中国畜牧兽医,2014,41(4):215-219.ZHAO Y H,WANG L,ZHANG T J,et al.Research progress on promoting effect and mechanism of melatonin on cashmere growth in cashmere goat[J].China Animal Husbandry & Veterinary Medicine,2014,41(4):215-219.(in Chinese)
[26] WANG D,SUN X L,WEI Y,et al.Nuclear miR-122 directly regulates the biogenesis of cell survival oncomiR miR-21 at the posttranscriptional level[J].Nucleic Acids Res,2018,46(4):2012-2029.
[27] TANG R,LI L M,ZHU D H,et al.Mouse miRNA-709 directly regulates miRNA-15a/16-1 biogenesis at the posttranscriptional level in the nucleus:evidence for a microRNA hierarchy system[J].Cell Res,2012,22(3):504-515.
[28] 李晓燕,张璐,王乐乐,等.绒山羊次级毛囊生长调控基因的筛选[J].畜牧兽医学报,2015,46(10):1733-1740.LI X Y,ZHANG L,WANG L L,et al.Selection of relative gene during secondary hair follicle growth in cashmere goat[J].Acta Veterinaria et Zootechnica Sinica,2015,46(10):1733-1740.(in Chinese)
[29] 张春兰.内蒙古阿尔巴斯白绒山羊毛囊生长周期性变化与KAP6 cDNA的特征分析[D].呼和浩特:内蒙古农业大学,2003.ZHANG C L.Periodic variety of hair follicles about Inner Mongolia ARBAS white cashmere goats and characteristic of KAP6 cDNA[D].Hohhot:Inner Mongolia Agricultural University,2003.(in Chinese)