文蛤SRBI基因克隆及其在不同壳色群体中的差异表达
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摘要
为了研究SRBI基因的结构、功能以及在贝类壳色形成中的作用,利用SMART RACE技术克隆得到文蛤(Meretrix meretrix)SRBI(Mm-SRBI)基因的全长序列,并对其内含子特征及不同组织、不同壳色群体外套膜中的表达差异进行了分析。结果表明:Mm-SRBI基因c DNA全长1676 bp,开放阅读框1515 bp,编码504个氨基酸,结构域预测发现有一个CD36结构域;氨基酸序列比对发现,与华贵栉孔扇贝的同源性最高(55%),与其他物种的相似性在34%—40%,表明该基因变异较大;在Mm-SRBI基因中扩增出12个内含子,均存在于开放阅读框中,且都遵循GT-AG原则;荧光定量PCR(q RT-PCR)结果表明,Mm-SRBI在闭壳肌、外套膜、斧足、鳃、内脏团和水管6个组织均有表达,其中在外套膜中表达量显著高于其他组织(P<0.01),这可能与外套膜中类胡萝卜素含量较高有关;不同壳色群体外套膜中基因表达分析表明,Mm-SRBI在黑斑和红壳文蛤中的表达量显著高于白壳文蛤(P<0.05)。实验结果为文蛤壳色形成研究奠定了基础。
To explore the molecular structure and biological function of SRBI in the process of carotenoids accumulation in Meretrix meretrix, we cloned c DNA of Mm-SRBI by SMART RACE technique and conducted the bioinformatics, expression analysis and intron amplification. The full length c DNA of Mm-SRBI is 1723 bp that contains a complete 1515 bp ORF encoding 504 amino acids with one functional domain of Mm-SRBI protein(CD36). Mm-SRBI is highly homologous with Mimachlamys nobilis and shares 55% similarity in amino acid sequence. It shares 34%—40%similarity with vertebrates. We also amplified 12 introns of Mm-SRBI, which followed the principle of GT-AG. MmSRBI gene expressed in all six tested tissues with the highest level in mantle(P<0.01), suggesting that Mm-SRBI gene maybe relate to high carotenoid in mantle. Moreover, the expression of Mm-SRBI gene in red shell strain was significantly higher than that of white shell strain(P<0.05). In conclusion, this study provides helpful foundation to explore the shell colors on this species.
To explore the molecular structure and biological function of SRBI in the process of carotenoids accumulation in Meretrix meretrix, we cloned c DNA of Mm-SRBI by SMART RACE technique and conducted the bioinformatics, expression analysis and intron amplification. The full length c DNA of Mm-SRBI is 1723 bp that contains a complete 1515 bp ORF encoding 504 amino acids with one functional domain of Mm-SRBI protein(CD36). Mm-SRBI is highly homologous with Mimachlamys nobilis and shares 55% similarity in amino acid sequence. It shares 34%—40%similarity with vertebrates. We also amplified 12 introns of Mm-SRBI, which followed the principle of GT-AG. MmSRBI gene expressed in all six tested tissues with the highest level in mantle(P<0.01), suggesting that Mm-SRBI gene maybe relate to high carotenoid in mantle. Moreover, the expression of Mm-SRBI gene in red shell strain was significantly higher than that of white shell strain(P<0.05). In conclusion, this study provides helpful foundation to explore the shell colors on this species.
引文
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[13]Acton S,Rigotti A,Landschulz K T,et al.Identification of scavenger receptor SR-BI as a high density lipoprotein receptor[J].Science,1996,271(5248):518-520
[14]Trigatti B L,Fuller M.HDL signaling and protection against coronary artery atherosclerosis in mice[J].The Journal of Biomedical Research,2016,30(2):94-100
[15]Seizer P,Schiemann S,Merz T,et al.CD36 and macrophage scavenger receptor a modulate foam cell formation via inhibitio19n of lipid-laden platelet phagocytosis[J].Semin Thromb Hemost,2010,36(2):157-162
[16]Cai L,Ji AL,de Beer FC,et al.SR-BI protects against endotoxemia in mice through its roles in glucocorticoid production and hepatic clearance[J].Journal of Clinical Investigation,2008,118(1):364-375
[17]Mouss M,Landrier J F,Reboul E,et al.Lycopene absorption in human intestinal cells and inmice involves scavenger receptor class B type I but not Niemann-Pick C1-like 1[J].The Journal of Nutrition,2008,138(8):1432-1436
[18]Georg Lietz,Anthony Oxley,Christine Boesch-Saadatmandi,et al.Importance ofβ,β-carotene 15,15′-monooxygenase 1(BCMO1)andβ,β-carotene 9′,10′-dioxygenase 2(BCDO2)in nutrition and health[J].Molecular Nutrition&Food Research,2012,56(2):241-250
[19]Kiefer C,Sumser E,Wernet M F,et al.A class B scavenger receptor mediates the cellular uptake of carotenoids in Drosophila[J].Proceedings of the National Academy of Sciences of the United States of America,2002,99(16):10581-10586
[20]Georgomanolis T,Sofiadis K,Papantonis A.Cutting a long intron short:recursive splicing and its implications[J].Frontiers in Physiology,2016,7:1-5
[21]Niemsiri V,Wang X,Pirim D,et al.Genetic contribution of SCARB1 variants to lipid traits in African Blacks:a candidate gene association study[J].BMC Medical Genetics,2015,16(1):1-30
[22]Voolstra O,Kiefer C,Martin Hoehne,et al.The Drosophila class B scavenger receptor Nina D-I is a cell surface receptor mediating carotenoid transport for visual chromophore synthesis[J].Biochemistry,2006,45:13429-13437
[23]Dong ZP,Chai C L,Dai F Y,et al.Expression pattern and tissue localization of the class B scavenger receptor Bm SCRBQ4 in Bombyx moil[J].Insect Science,2015,22(6):739-747
[24]Kaestner K H,Ntambi J M,Kelly,T J Jr,et al.Differentiation-induced gene expression in 3T3-L1 preadipocytes.Asecond differentially expressed gene encoding stearoylCo A desaturase[J].The Journal of Biological Chemistry,1989,264(25):14755-14761
[25]Li Y,Zhou M,Zhou H,et al.The last intron of the human thrombopoietin gene enhances expression in milk of transgenic mice[J].Functional&Integrative Genomics,2014,14(1):229-236
[26]Dai P,Huan P,Wang H,et al.Characterization of a longchain fatty acid-Co A ligase 1 gene and association between its SNPs and growth traits in the clam Meretrix meretrix[J].Gene,2015,566(2):194-200
[27]Dong Z P,Chai C L,Dai F Y,et al.Cloning,Expression Feature,and Tissue Location of Bm SCRBQ1,a Class BScavenger Receptor in Bombyx mori(L.)[J].Annals of The Entomological Society of America,2013,106(4):503-509
[28]Cao G,Garcia C K,Wyne K L,et al.Structure and localization of the human gene encoding SR-BI/CLA-1.Evidence for transcriptional control by steroidogenic factor 1[J].Journal of Biological Chemistry,1997,272(52):33068-33076
[2]Tsuchida K,Sakudoh T.Recent progress in molecular genetic studies on the carotenoid transport system using cocoon-color mutants of the silkworm[J].Archives of Biochemistry&Biophysics,2015,572:151-157
[3]Dong Z P,Chai C L,Dai F Y,et al.Identification,phylogentic and expressional analyses of class B scanvenger receptor genes in silkworm,Bombyx mori[J].Science of Sericulture,2010,36(5):743-753[董占鹏,柴春利,代方银,等.家蚕B型清道夫受体基因的鉴定及系统发生与表达分析.蚕业科学,2010,36(5):743-753]
[4]Akita N,Tsujita M,Yokota T,et al.High density lipoprotein turnover is dependent on peroxisome proliferator-activated receptorαin mice[J].Journal of Atherosclerosis&Thrombosis,2010,17(11):1149-1159
[5]Acton S L,Scherer P E,Lodish H F,et al.Expression cloning of SR-BI,a CD36-related class B scavenger receptor[J].Journal of Biological Chemistry,1994,269(33):21003-21009
[6]Liu H,Zheng H,Zhang H,et al.A de novo transcriptome of the noble scallop,Chlamys nobilis,focusing on mining transcripts for carotenoid-based coloration[J].BMC Genomics,2015,16(1):2-13
[7]Lei C,Bei W L,Zheng Z,et al.Molecular characterization and expression analysis of Pm SR-B gene from pearl oyster Pinctada fucata martensii[J].Journal of Guangdong Ocean University,2017,37(1):7-14[雷超,贝伟烈,郑哲,等.马氏珠母贝B型清道夫受体Pm SR-B基因克隆与表达性分析.广东海洋大学学报,2017,37(1):7-14]
[8]Qi X Y.The analysis of molecular mechanisms of shell color formation for Meretrix meretrix[D].Ningbo University.2014[齐晓艳.文蛤(Meretrix meretrix)壳色花纹性状的分子遗传基础研究.宁波大学.2014]
[9]Zhan Y L.Identification of shell color and the related gene and micro RNA research of Meretrix meretrix[D].Zhejiang Wanli University.2015[詹艳玲.文蛤(Mere-trix meretrix)壳色的鉴定及其相关基因和micro RNA研究.浙江万里学院.2015]
[10]Gao X Y,Dong Y H,Yao H H,et al.Cloning,spatiotemporal expression and SNPs identification of HDAC1 gene in hard clam Meretrix Meretrix[J].Acta Hydrobiologica Sinica,2016,40(3):601-608[高晓艳,董迎辉,姚韩韩,等.文蛤HDAC1基因克隆、时空表达及生长相关SNP位点筛查.水生生物学报,2016,40(3):601-608]
[11]Gao X Y,Dong Y H,Shi S J,et al.Cloning,spatiotemporal expression and SNPs identification of GRB2 gene in hard clam Meretrix meretrix[J].Journal of Fisheries of China,2015,39(9):1324-1332[高晓艳,董迎辉,施沈佳,等.文蛤生长因子受体结合蛋白2(GRB2)基因克隆、时空表达及SNP位点筛查.水产学报,2015,39(9):1324-1332]
[12]Li M,Zhou S M,Liu L,et al.Molecular clone and expression of C-TYPE lectin in meretrix meretrix[J].Oceanologia et Limnologia Sinica,2015,46(5):1186-1192[李猛,周素明,刘璐,等.文蛤(Meretrix meretrix)C-型凝集素基因的分子克隆及表达分析.海洋与湖沼,2015,46(5):1186-1192]
[13]Acton S,Rigotti A,Landschulz K T,et al.Identification of scavenger receptor SR-BI as a high density lipoprotein receptor[J].Science,1996,271(5248):518-520
[14]Trigatti B L,Fuller M.HDL signaling and protection against coronary artery atherosclerosis in mice[J].The Journal of Biomedical Research,2016,30(2):94-100
[15]Seizer P,Schiemann S,Merz T,et al.CD36 and macrophage scavenger receptor a modulate foam cell formation via inhibitio19n of lipid-laden platelet phagocytosis[J].Semin Thromb Hemost,2010,36(2):157-162
[16]Cai L,Ji AL,de Beer FC,et al.SR-BI protects against endotoxemia in mice through its roles in glucocorticoid production and hepatic clearance[J].Journal of Clinical Investigation,2008,118(1):364-375
[17]Mouss M,Landrier J F,Reboul E,et al.Lycopene absorption in human intestinal cells and inmice involves scavenger receptor class B type I but not Niemann-Pick C1-like 1[J].The Journal of Nutrition,2008,138(8):1432-1436
[18]Georg Lietz,Anthony Oxley,Christine Boesch-Saadatmandi,et al.Importance ofβ,β-carotene 15,15′-monooxygenase 1(BCMO1)andβ,β-carotene 9′,10′-dioxygenase 2(BCDO2)in nutrition and health[J].Molecular Nutrition&Food Research,2012,56(2):241-250
[19]Kiefer C,Sumser E,Wernet M F,et al.A class B scavenger receptor mediates the cellular uptake of carotenoids in Drosophila[J].Proceedings of the National Academy of Sciences of the United States of America,2002,99(16):10581-10586
[20]Georgomanolis T,Sofiadis K,Papantonis A.Cutting a long intron short:recursive splicing and its implications[J].Frontiers in Physiology,2016,7:1-5
[21]Niemsiri V,Wang X,Pirim D,et al.Genetic contribution of SCARB1 variants to lipid traits in African Blacks:a candidate gene association study[J].BMC Medical Genetics,2015,16(1):1-30
[22]Voolstra O,Kiefer C,Martin Hoehne,et al.The Drosophila class B scavenger receptor Nina D-I is a cell surface receptor mediating carotenoid transport for visual chromophore synthesis[J].Biochemistry,2006,45:13429-13437
[23]Dong ZP,Chai C L,Dai F Y,et al.Expression pattern and tissue localization of the class B scavenger receptor Bm SCRBQ4 in Bombyx moil[J].Insect Science,2015,22(6):739-747
[24]Kaestner K H,Ntambi J M,Kelly,T J Jr,et al.Differentiation-induced gene expression in 3T3-L1 preadipocytes.Asecond differentially expressed gene encoding stearoylCo A desaturase[J].The Journal of Biological Chemistry,1989,264(25):14755-14761
[25]Li Y,Zhou M,Zhou H,et al.The last intron of the human thrombopoietin gene enhances expression in milk of transgenic mice[J].Functional&Integrative Genomics,2014,14(1):229-236
[26]Dai P,Huan P,Wang H,et al.Characterization of a longchain fatty acid-Co A ligase 1 gene and association between its SNPs and growth traits in the clam Meretrix meretrix[J].Gene,2015,566(2):194-200
[27]Dong Z P,Chai C L,Dai F Y,et al.Cloning,Expression Feature,and Tissue Location of Bm SCRBQ1,a Class BScavenger Receptor in Bombyx mori(L.)[J].Annals of The Entomological Society of America,2013,106(4):503-509
[28]Cao G,Garcia C K,Wyne K L,et al.Structure and localization of the human gene encoding SR-BI/CLA-1.Evidence for transcriptional control by steroidogenic factor 1[J].Journal of Biological Chemistry,1997,272(52):33068-33076