|
|
Prediction and Identification of Prox1 Nuclear Localization Signal in Chicken (Gallus gallus) |
YUAN Chao, GAO Hong-Bo, ZHOU Lei, HAN Yi-Fan, DUAN Zhi-Qiang* |
Key Laboratory of Animal Genetics,Breeding and Reproduction in Plateau Mountainous Region,Ministry of Education/Key Laboratory of Animal Genetics,Breeding and Reproduction of Guizhou Province/College of Animal Sciences,Guizhou University,Guiyang 550025,China |
|
|
Abstract Prox1 (prospero-related homeobox protein 1) is a transcription factor that plays an important role in embryonic development and carcinogenesis. This study aimed to predict and identify the nuclear localization signal (NLS) in chicken (Gallus gallus) Prox1 protein. Chicken Prox1 protein was truncated to determine the NLS region by fluorescence observation and subcellular localization analysis. The online software was then used to predict the putative NLS (pNLS) in Prox1 truncates,and the NLS was confirmed by examining the subcellular localization of the pNLS deletants. In addition,the conservation and nuclear import ability of chicken Prox1 NLS were analyzed. Moreover,the key basic amino acids determining the nuclear localization of Prox1 NLS were identified by point mutation test. The results showed that the N-terminus (1~220 aa) of chicken Prox1 protein had nuclear localization characteristics and two pNLS including pNLS1 (15KRRR18) and pNLS2 (163RAKRAR168) existed in this region by NLS software prediction. The subcellular localization analysis of pNLS deletants revealed that pNLS1 and pNLS2 codetermined the nuclear localization of chicken Prox1 protein. In addition,the NLS1 and NLS2 motifs in chicken Prox1 protein were conserved among other poultry,human (Homo sapiens) and different mammals,and had the same nuclear import ability as the NLS of SV40 large T antigen. Moreover,the results of basic amino acids mutation experiments showed that K15,R16,R18 and K165,R166,R168 were key amino acids in NLS1 and NLS2,respectively,which jointly determined nuclear localization of chicken Prox1 protein. This study confirmed two highly conserved NLSs present in chicken Prox1 protein,and both NLSs determined the nuclear localization together,which provided a reference for further studying molecular mechanism and function of the nuclear localization of chicken Prox1 protein.
|
Received: 07 April 2020
|
|
Corresponding Authors:
*zqduan@gzu.edu.cn
|
|
|
|
[1] 邓珊珊,高洪波,袁超,等.2018.鸡基质蛋白3核定位信号的预测与鉴定[J].畜牧兽医学报,2018,49(11):2486-2495. (Deng S S,Gao H B,Yuan C,et al.2018.Prediction and identification of chicken matrix protein 3 nuclear localization signal[J].Acta Veterinaria et Zootechnica Sinica,49(11):2486-2495.) [2] 邓珊珊.2019.鸡基质蛋白3在新城疫病毒复制中的作用研究[D].硕士学位论文,贵州大学,导师:嵇辛勤,pp.1-15. (Deng S S.2019.Study on the role of chicken matrix protein 3 in the replication ofNewcastle disease virus[D].Thesis for M.S.,Guizhou University,Supervisor:Ji X Q,pp.1-15.) [3] 付爱玲,孙曼霁.2003.核定位信号介导的细胞运及其机制[J].中国药理学通报,6(10):1081-1083. (Fu A L,Sun M Q.Nuclear localization signal-mediated cellular transport and its mechanism[J].Chinese Pharmacological Bulletin,2003,6(10):1081-1083.) [4] 胡焱,熊建民,赵佳福,等.2017.Importin β1介导的病毒蛋白入核转运在病毒复制中作用的研究进展[J].中国细胞生物学学报,39(8):1091-1098. (Hu Y,Xiong J M,Zhao J F,et al.2017.Advances in importin β1-mediated virus protein nuclear transfer in viral replication[J].Chinese Journal of Cell Biology,39(8):1091-1098.) [5] 陶华平.2014.核定位信号研究进展[J].生物学通报,49(8):5-10. (Tao H P.2014.Research progress of nuclear localization signals[J].Biological Bulletin,49(8):5-10.) [6] 袁继红,龙欢,田明芳,等.2013.猪Prox1基因的染色体定位和亚细胞定位的研究[J].畜牧兽医学报,44(3):351-357. (Ji-Hong Y,Huan L,Ming-Fang T,et al.2013.Study on chromosomal and subcellular localization for porcineProx1gene[J].Acta Veterinaria et Zootechnica Sinica,2013,44(3):351-357.) [7] 袁超,高洪波,邓珊珊,等.2019.鸡Prox1基因的克隆与生物信息学分析[J].中国家禽,41(23):15-20. (Yuan C,Gao H B,Deng S S,et al.2019.Cloning and bioinformatics analysis of chickenProx1gene[J].Chinese Poultry,41(23):15-20.) [8] 袁志栋,沈孝宙,李建凡,等.2004.核定位信号肽用于基因转移的研究进展[J].生命科学,3(1):52-57. (Yuan Z D,Shen X Z,Li J F,et al.2004.Research progress of nuclear localization signal peptide for gene transfer[J].Life Sciences,3(1):52-57.) [9] Armour S M,Remsberg J R,Damle M,et al.2017.An HDAC3-PROX1 corepressor module acts on HNF4α to control hepatic triglycerides[J].Nature Communications,8(1):549-561. [10] Audette D S,Anand D,So T,et al.2015.Prox1 and fibroblast growth factor receptors form a novel regulatory loop controlling lens fiber differentiation and gene expression[J].Development,143(2):318-332. [11] Chen X,Patel T P,Simirskii V I,et al.2008.PCNA interacts with Prox1 and represses its transcriptional activity[J].Molecular Vision,14(2):2076-2086. [12] Chen X,Taube J R,Simirskii V I,et al.2008.Dual roles for Prox1 in the regulation of the chicken βB1-crystallin promoter[J].Investigative Ophthalmology & Visual Science,49(4):1542-1552. [13] Coleman N A,Peeples M E.1993.The matrix protein ofNewcastle disease viruslocalizes to the nucleus via a bipartite nuclear localization signal[J].Virology,195(2):596-607. [14] Duan Z,Deng S,Ji X,et al.2019.Nuclear localization ofNewcastle disease virusmatrix protein promotes virus replication by affecting viral RNA synthesis and transcription and inhibiting host cell transcription[J].Veterinary Research,50(1):22-42. [15] Duan Z,Xu H,Ji X,Zhao J,et al.2018.Importin α5 negatively regulates importin β1-mediated nuclear import ofNewcastle disease virusmatrix protein and viral replication and pathogenicity in chicken fibroblasts[J].Virulence,9(1):783-803. [16] Duan J H,Tang Z Q,Mu H,et al.2016.Nuclear import of prototype foamy virus transactivator Bel1 is mediated by KPNA1,KPNA6 and KPNA7[J].International Journal of Molecular Medicine,38(2):399-406. [17] Dudas J,Elmaouhoub A,Mansuroglu T,et al.2006.Prospero-related homeobox 1 (Prox1) is a stable hepatocyte marker during liver development,injury and regeneration,and is absent from 'oval cells'[J].Histochemistry and Cell Biollogy,126(5):549-562. [18] Elsir T,Smits A,Lindström M S,et al.2012.Transcription factor PROX1:Its role in development and cancer[J].Cancer and Metastasis Reviews,31(3-4):793-805. [19] Fahrenkrog B,Aebi U.2003.The nuclear pore complex:Nucleocytoplasmic transport and beyond[J].Nature Reviews Molecular Cell Biology,4(10):757-766. [20] Goto T,Elbahrawy A,Furuyama K,et al.2017.Liver-specific Prox1 inactivation causes hepatic injury and glucose intolerance in mice[J].FEBS Letters,591(4):624-635. [21] Goldfarb D S,Corbett A H,Mason D A,et al.2004.Importin alpha:A multipurpose nuclear-transport receptor[J].Trends in Cell Biology,14(9):505-514. [22] Kaltezioti V,Kouroupi G,Oikonomaki M,et al.2010.Prox1 regulates the notch1-mediated inhibition of neurogenesis[J].PLOS Biology,8(12):565-585. [23] Kalderon D,Roberts B L,Richardson W D,et al.1984.A short amino acid sequence able to specify nuclear location[J].Cell,39(3):499-509. [24] Kivelä R,Salmela I,Nguyen Y H,et al.2016.The transcription factor Prox1 is essential for satellite cell differentiation and muscle fiber-type regulation[J].Nature Communications,7(3):13124-13136. [25] Kapinos L E,Huang B,Rencurel C,et al.2017.Karyopherins regulate nuclear pore complex barrier and transport function[J].Journal of Cell Biology,216(11):3609-3624. [26] Papoutsi M,Dudas J,Jürgen Becker,et al.2007.Gene regulation by homeobox transcription factor Prox1 in murine hepatoblasts[J].Cell and Tissue Research,330(3):209-220. [27] Petchey L K,Risebro C A,Vieira J M,et al.2014.Loss of Prox1 in striated muscle causes slow to fast skeletal muscle fiber conversion and dilated cardiomyopathy[J].Proceedings of the National Academy of Sciences of the USA,111(26):9515-9520. [28] Qin J,Zhai J,Hong R,et al.2009.Prospero-related homeobox protein (Prox1) inhibitsHepatitis B virusreplication through repressing multiplecisregulatory elements[J].Journal of General Virology,90(5):1246-1255. [29] Qin J,Gao D M,Jiang Q F,et al.2004.Prospero-related homeobox (Prox1) is a corepressor of human liver receptor homolog-1 and suppresses the transcription of the cholesterol 7-alpha-hydroxylase gene[J].Molecular Endocrinology,18(10):2424-2439. [30] Risebro C A,Searles R G,Melville A A,et al.2009.Prox1 maintains muscle structure and growth in the developing heart[J].Development,136(4):699. [31] Ryter J M,Doe C Q,Matthews B W.2002.Structure of the DNA binding region of prospero reveals a novel homeo-prospero domain[J].Structure,10(11):1541-1549. [32] Stergiopoulos A,Elkouris M,Politis P K.2015.Prospero-related homeobox 1 (Prox1) at the crossroads of diverse pathways during adult neural fate specification[J].Frontiers in Cellular Neuroscience,26(8):454-462. [33] Wigle J T,Oliver G.1999.Prox1 function is required for the development of the murine lymphaticsystem[J].Cell,98(6):769-778. [34] Wigle J T,Chowdhury K,Gruss P,et al.1999.Prox1 function is crucial for mouse lens-fibre elongation[J].Nature Genetics,21(3):318-332. [35] Yousef M S,Matthews B W.2005.Structural basis of Prospero-DNA interaction:Implications for transcription regulation in developing cells[J].Structure,13(4):601-607. [36] Yoo J,Kang J,Lee H N,et al.2010.Kaposin-B enhances thePROX1mRNA stability during lymphatic reprogramming of vascular endothelial cells byKaposi's sarcoma herpes virus[J].PLOS Pathogens,6(8):1046-1060. [37] Yeung P L,Chen L Y,Tsai S C,et al.2008.Daxx contains two nuclear localization signals and interacts with importin α3[J].Journal of Cellular Biochemistry,103(2):456-470. [38] Zinovieva R D,Duncan M K,Johnson T R,et al.1996.Structure and chromosomal localization of the human homeobox geneProx1[J].Genomics,35(3):517-522. |
[1] |
WANG Kun-Peng, LIU Pan-Pan, XIA Yin-He, GAO Dong-Sheng, CHANG Hong-Tao, WANG Xin-Wei, LI Yong-Tao, ZHAO Jun, CHEN Lu, WANG Chuan-Qing, YANG Xia. Construction of Recombinant Lactobacillus casei Expressing Gallibacterium anatis flfA and Evaluation of Oral Immune Effect[J]. 农业生物技术学报, 2020, 28(9): 1652-1661. |
|
|
|
|