c-Myc基因在栉孔扇贝年周期发育性腺中的表达特征
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摘要
c-MYC在高等脊椎动物的细胞增殖、细胞分化和癌症发生中具有重要的调控作用。本研究克隆了栉孔扇贝cMyc基因的全长cDNA序列,分析了其在性腺年周期发育和配子发生过程中的表达特征。栉孔扇贝c-Myc cDNA全长2 943bp,可编码397个氨基酸,推导的氨基酸序列中除了包含MYC家族的保守结构域外,还具有c-MYC蛋白所特有的Myc-N转录激活区。qRT-PCR结果显示,栉孔扇贝卵巢中的c-Myc mRNA水平随着卵巢的发育和成熟显著升高;精巢中的表达水平在成熟期之前(休止期至生长期)逐渐升高,但在成熟期精巢中的表达水平显著低于增殖期和生长期。原位杂交和免疫组织化学结果显示,c-Myc mRNA及c-MYC蛋白的阳性信号均可在栉孔扇贝精巢的精原细胞和精母细胞以及卵巢的所有生殖细胞中检测到,推测其可能与栉孔扇贝的性腺发育和配子发生过程有关。
c-MYC plays important roles in regulating the cell proliferation,cell differentiation,and tumorigenesis in vertebrates.In the present study,we identified a sequence of c-Myc cDNA in the scallop Chlamys farreri and analyzed the expression characteristics of target gene in gonads of annual cycle and gametogenesis.The full-length of C.farreri c-Myc was 2 943 bp with an open reading frame of 1 194 bp which encoded a protein of 397 amino acids.The deduced amino acid sequence contained the conserved domains of the MYC family proteins and the conserved Myc-N transcriptional activation region of the c-MYC protein.The qRT-PCR showed that the levels of c Myc mRNA in C.farreri ovaries increased significantly with the development and maturation of ovary(P<0.05).In the testes,the levels of c-Myc mRNA also increased gradually before the mature stage,while it decreased significantly in mature testis compared with proliferative and growing testes(P<0.05).Results of in situ hybridization and immunohistochemistry revealed that positive signals of the c-Myc mRNA and c-MYC protein were visible in spermatogonia and spermatocytes of the testes,and all the oogonia and oocytes of the ovaries in C.farreri.We speculated preliminarily that c-Myc might be related to the gonad development and gametogenesis in C.farreri.
c-MYC plays important roles in regulating the cell proliferation,cell differentiation,and tumorigenesis in vertebrates.In the present study,we identified a sequence of c-Myc cDNA in the scallop Chlamys farreri and analyzed the expression characteristics of target gene in gonads of annual cycle and gametogenesis.The full-length of C.farreri c-Myc was 2 943 bp with an open reading frame of 1 194 bp which encoded a protein of 397 amino acids.The deduced amino acid sequence contained the conserved domains of the MYC family proteins and the conserved Myc-N transcriptional activation region of the c-MYC protein.The qRT-PCR showed that the levels of c Myc mRNA in C.farreri ovaries increased significantly with the development and maturation of ovary(P<0.05).In the testes,the levels of c-Myc mRNA also increased gradually before the mature stage,while it decreased significantly in mature testis compared with proliferative and growing testes(P<0.05).Results of in situ hybridization and immunohistochemistry revealed that positive signals of the c-Myc mRNA and c-MYC protein were visible in spermatogonia and spermatocytes of the testes,and all the oogonia and oocytes of the ovaries in C.farreri.We speculated preliminarily that c-Myc might be related to the gonad development and gametogenesis in C.farreri.
引文
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[23] Li Y,Zhang L,Sun Y,et al.Transcriptome sequencing and comparative analysis of ovary and testis identifies potential key sex-related genes and pathways in scallop Patinopecten yessoensis[J].Marine Biotechnology,2016,18(4):453-465.
[24] Packham G,Cleveland J L.c-Myc and apoptosis[J].Biochimica Et Biophysica Acta,1995,1242(1):11-28.
[25] Wang Y,Liu S,Zhang G,et al.Knockdown of c-Myc expression by RNAi inhibits MCF-7breast tumor cells growthin vitro andin vivo[J].Breast Cancer Research,2004,7(2):R220.
[2]潘晶晶.c-Myc蛋白去泛素化机制的研究[D].上海:华东师范大学,2015.Pan J.Study on c-Myc Deubiquitination[D].Shanghai:East China Normal University,2015.
[3] Dang C V,O’Donnell K A,Zeller K I,et al.The c-Myc target gene network[J].Seminars in Cancer biology,2006,16(4):253-264.
[4] Bretones G,Delgado M,Leon J.Myc and cell cycle control[J].Biochimica et Biophysica Acta,2015,1849(5):506-516.
[5] Dwivedi S,Mcmeekin S,Slaughter K,et al.Role of TGF-βsignaling in uterine carcinosarcoma[J].Oncotarget,2015,6(16):14646-14655.
[6] Chen B,Wu Y,Tanaka Y,et al.Small molecules targeting c-Myc oncogene:promising anti-cancer therapeutics[J].International Journal of Biological Sciences,2014,10(10):1084-1096.
[7]邹治铭,郭华雄.c-Myc在肿瘤中作用的研究进展[J].现代医学,2016,44(7):1024-2026.Zou Z,Guo H.Related research in cancer of c-Myc[J].Modern Medical Journal,2016,44(7):1024-2026.
[8] Takahashi K,Yamanaka S.Induction of pluripotent stem cells from mouse embryonic and adult fibro-blast cultures by defined factors[J].Cell,2006,126(4):663-676.
[9] Meyer N,Penn L.Reflecting on 25years with MYC[J].Nature Reviews Cancer,2008,8(12):976-990.
[10]魏小元,徐雍羽,张儒,等.c-Myc对诱导性多能干细胞的诱导效率及全能性影响的研究[J].中国细胞生物学学报,2014,36(4):430-437.Wei X,Xu Y,Zhang R,et al.Influence of c-Myc on induction Efficiency and pluripotency of induced pluripotent stem cells[J].Chinese Journal of Cell Biology,2014,36(4):430-437.
[11]韩威,苏一军,朱云芬,等.文昌鸡性成熟启动前后下丘脑cMyc/LIN28B/let-7a通路表达规律分析[J].畜牧兽医学报,2016,47(4):709-715.Han W,Su Y,Zhu Y,et al.Analysis on changes of Hypothalamic c-Myc/LIN28B/let-7apathway transcription during puberty onset in Wenchang chicken[J].Acta Veterinaria et Zootechnica Sinica,2016,47(4):709-715.
[12] Zhang G,Fang X,Guo X,et al.The oyster genome reveals stress adaptation and complexity of shell formation[J].Nature,2012,490(7418):49-54.
[13]张若云,徐广锐,潘聪,等.合浦珠母贝iPS细胞相关的转录因子的克隆及分析[J].海洋科学,2014,38(4):7-14.Zhang R,Xu G,Pan C,et al.Cloning and sequence analysis of iPS related transcription factors fromPinctada fucata[J].Marine Sciences,2014,38(4):7-14.
[14] Wang S,Hou R,Bao Z,et al.Transcriptome sequencing of Zhikong scallop(Chlamys farreri)and comparative transcriptomic analysis with Yesso scallop(Patinopecten yessoensis)[J].PloS One,2013,8(5):e63927.
[15] Bao Y,Xu F,Shimeld S M.Phylogenetics of lophotrochozoan bHLH genes and the evolution of lineage-specific gene duplicates[J].Genome Biology and Evolution,2017,9(4):869-886.
[16] Wang S,Zhang J,Jiao W,et al.Scallop genome provides insights into evolution of bilaterian karyotype and development[J].Nature Ecology and Evolution,2017,1:120.
[17]廖承义,徐应馥,王远隆.栉孔扇贝生殖周期[J].水产学报,1983,7:1-13.Liao C,Xu Y,Wang Y.Reproductive cycle of the scallop Chlamys farreri(Jones and Preston)at Qingdao[J].Journal of Fisheries of China/Shuichan Xuebao,1983,7(1):1-13.
[18] Liu J,Zhang Z,Zhang L,et al.Variations of estradiol-17βand testosterone levels correlated with gametogenesis in the gonad of Zhikong scallop(Chlamys farreri)during annual reproductive cycle[J].Canadian Journal of Zoology,2014,92(3):195-204.
[19] Bustin A,Benes V,Garson A,et al.The MIQE guidelines:Minimum information for publication of quantitative real-time PCR experiments[J].Clinical Chemistry,2009,55(4):611-622.
[20] Zhou Z,Wang L,Shi X,et al.The expression of dopa decarboxylase and dopamine beta hydroxylase and their responding to bacterial challenge during the ontogenesis of scallop Chlamys farreri[J].Fish and Shellfish Immunology,2012,33(1):67-74.
[21]刘晓玲.栉孔扇贝Cf-Foxl2基因的克隆、表达模式及功能初探[D].青岛:中国海洋大学,2012.Liu X.Molecular Cloning,Expression Pattern and Function Analysis of Cf-foxl2in the Scallop Chlamys farreri[D].Qingdao:Ocean University of China,2012.
[22] Ma Y,Zhang Z,Shao M,et al.Sulfide:Quinoneoxidoreductase from echiuran wormUrechis unicinctus[J].Marine Biotechnology,2011,13(1):93-107.
[23] Li Y,Zhang L,Sun Y,et al.Transcriptome sequencing and comparative analysis of ovary and testis identifies potential key sex-related genes and pathways in scallop Patinopecten yessoensis[J].Marine Biotechnology,2016,18(4):453-465.
[24] Packham G,Cleveland J L.c-Myc and apoptosis[J].Biochimica Et Biophysica Acta,1995,1242(1):11-28.
[25] Wang Y,Liu S,Zhang G,et al.Knockdown of c-Myc expression by RNAi inhibits MCF-7breast tumor cells growthin vitro andin vivo[J].Breast Cancer Research,2004,7(2):R220.