金针菇寡肽转运蛋白编码基因fvopt1与fvopt2的鉴定与表达分析
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摘要
基于金针菇(Flammulina velutipes)的基因组和转录组数据,通过本地BLAST方法,获得2个寡肽转运蛋白编码基因,命名为fvopt1和fvopt2,并对其结构、编码蛋白的基本性质、系统进化与表达模式进行分析。结果表明:fvopt1和fvopt2分别含有13和18个外显子,编码蛋白FVOPT1与FVOPT2各含774和772个氨基酸,相对分子质量为86120.25与86192.24,等电点为7.46与7.92,且均无信号肽。保守结构域分析与系统进化分析结果表明,FVOPT1与FVOPT2为寡肽转运蛋白家族成员。实时荧光定量PCR结果表明:在菌丝阶段,fvopt1在无氮培养基中的表达量高于其在含氮培养基中的表达量;在含氮培养基中,fvopt1在整个子实体发育阶段(原基、菌柄和菌盖)中表达量上调,fvopt2只在原基中高表达,推测fvopt1和fvopt2可能与子实体和原基的发育有关。
Using the local BLAST, two genes encoding oligopeptide transporter, fvoptl and fvopt2, were identified from the genome and transcriptome sequences of Flammulina velutipes. Gene structures, basic characteristics of deduced proteins, phylogenetic evolution and expression patterns of both genes were analyzed. The results showed that fvoptl and fvopt2 contained 13 and 18 exons, respectively; and their deduced proteins FVOPT1 and FVOPT2 consisted of 774 and 772 amino acids, respectively. Molecular weights of FVOPT1 and FVOPT2 were 86120.25 and 86192.24; their isoelectric points were 7.46 and 7.92,respectively. Neither protein contained a signal peptide. Conserved domain analysis and phylogenetic analysis revealed that FVOPT1 and FVOPT2 belonged to the oligopeptide transporter superfamily. Results of real time quantitative PCR indicated that the expression level of fvoptl during the mycelium stage was higher on nitrogen-free medium than on nitrogen-containing medium. When cultivated on nitrogen-containing medium,the expression level of fvoptl was up-regulated during the fruiting body development stage, including primordium, stipe and pileus. On the other hand, fvopt2 was only highly expressed in primordium.Therefore, fvopt1 and fvopt2 were hypothesized to be functionally important to the development of fruiting body and primordium, respectively.
Using the local BLAST, two genes encoding oligopeptide transporter, fvoptl and fvopt2, were identified from the genome and transcriptome sequences of Flammulina velutipes. Gene structures, basic characteristics of deduced proteins, phylogenetic evolution and expression patterns of both genes were analyzed. The results showed that fvoptl and fvopt2 contained 13 and 18 exons, respectively; and their deduced proteins FVOPT1 and FVOPT2 consisted of 774 and 772 amino acids, respectively. Molecular weights of FVOPT1 and FVOPT2 were 86120.25 and 86192.24; their isoelectric points were 7.46 and 7.92,respectively. Neither protein contained a signal peptide. Conserved domain analysis and phylogenetic analysis revealed that FVOPT1 and FVOPT2 belonged to the oligopeptide transporter superfamily. Results of real time quantitative PCR indicated that the expression level of fvoptl during the mycelium stage was higher on nitrogen-free medium than on nitrogen-containing medium. When cultivated on nitrogen-containing medium,the expression level of fvoptl was up-regulated during the fruiting body development stage, including primordium, stipe and pileus. On the other hand, fvopt2 was only highly expressed in primordium.Therefore, fvopt1 and fvopt2 were hypothesized to be functionally important to the development of fruiting body and primordium, respectively.
引文
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[17] WILES AM, CAI H, NAIDER F, et al. Nutrient regulation of oligopeptide transport in Saccharomyces cerevisiae[J]. Microbiology, 2006, 152(10):3133-3145.
[18] DESAI PR, THAKUR A, GANGULI D, et al. Glutathione utilization by Candida albicans requires a functional glutathione degradation(DUG)pathway and OPT7, an unusual member of the oligopeptide transporter family[J]. Journal of Biological Chemistry, 2011, 286(48):41183-41194.
[19] TSAY YF, CHIU CC, TSAI CB, et al. Nitrate transporters and peptide transporters[J]. FEBS Letters, 2007,581(12):2290-2300.
[20] LUCIC E, FOURREY C, KOHLER A,et al. A gene repertoire for nitrogen transporters in Laccaria bicolor[J]. New Phytologist, 2008, 180(2):343-364.
[21] REUSS O, MORSCHHAUSER J. A family of oligopeptide transporters is required for growth of Candida albicans on proteins[J]. Molecular Microbiology, 2006, 60(3):795-812.
[2] STEINER HY, NAIDER F, BECKER JM. The PTR family:a new group of peptide transporters[J]. Molecular Microbiology, 1995, 16(5):825-834.
[3] GOMOLPLITINANT KM, SAIER MH. Evolution of the oligopeptide transporter family[J]. The Journal of Membrane Biology, 2011, 240(2):89-110.
[4] THAKUR A, KAUR J, BACHHAWAT AK. Pgtl, a glutathione transporter from the fission yeast Schizosaccharomyces pombe[J]. FEMS Yeast Research, 2008, 8(6):916-929.
[5] DWORECK T, WOLF K, ZIMMERMANN M. SpOPTl, a member of the oligopeptide family(OPT)of the fission yeast Schizosaccharomyces pombe, is involved in the transport of glutathione through the outer membrane of the cell[J]. Yeast, 2009, 26(1):67-73.
[6] KAUR J, SRIKANTH CV, BACHHAWAT AK. Differential roles played by the native cysteine residues of the yeast glutathione transporter, Hgtlp[J]. FEMS Yeast Research, 2009, 9(6):849-866.
[7] ELBAZ-ALON Y, MORGAN B, CLANCY A, et al. The yeast oligopeptide transporter Opt2 is localized to peroxisomes and affects glutathione redox homeostasis[J]. FEMS Yeast Research, 2014, 14(7):1055-1067.
[8] REUβO, MORSCHHAUSER J. A family of oligopeptide transporters is required for growth of Candida albicans on proteins[J]. Molecular Microbiology, 2006, 60(3):795-812.
[9] LIN H, ZHANG Z, ZHANG MQ, et al. ZOOM Zillions of oligos mapped[J]. Bioinformatics, 2008, 24(21):2431-2437.
[10] HU B,JIN JP,GUO AY,et al. GSDS 2.0:an upgraded gene feature visualization server[J]. Bioinformatics,2015, 31(8):1296-1297.
[11] YAMADA M, SAKURABA S, SHIBATA K, et al. Cloning and characterization of a gene coding for a hydrophobin, Fv-hydl, specifically expressed during fruiting body development in the basidiomycete Flammulina velutipes[J]. Applied Microbiology and Biotechnology, 2005, 67(2):240-246.
[12] KIM HI,LEE C S,PARK YJ. Further characterization of hydrophobin genes in genome of Flammulina velutipes[J]. Mycoscience, 2016, 57(5):320-325.
[13]姚建,董振辉,马雪西,等.一种基于免费在线工具的定量PCR引物设计方法[J].基因组学与应用生物学,2015,34(12):2779-2784.
[14] WILES AM. The oligopeptide transport family:informatic and experimental characterization[D]. Knoxville:University of Tennessee, 2005.
[15] XIANG Q, SHEN K, YU X, et al. Analysis of the oligopeptide transporter gene family in Ganoderma lucidum:structure, phylogeny, and expression patterns[J]. Genome, 2016, 60(4):293-302.
[16] YEN MR, TSENG YH, SAIER JR, et al. Maize yellow stripel, an iron-phytosiderophore uptake transporter, is a member of the oligopeptide transporter(OPT)family[J]. Microbiology, 2001,147(11):2881-2883.
[17] WILES AM, CAI H, NAIDER F, et al. Nutrient regulation of oligopeptide transport in Saccharomyces cerevisiae[J]. Microbiology, 2006, 152(10):3133-3145.
[18] DESAI PR, THAKUR A, GANGULI D, et al. Glutathione utilization by Candida albicans requires a functional glutathione degradation(DUG)pathway and OPT7, an unusual member of the oligopeptide transporter family[J]. Journal of Biological Chemistry, 2011, 286(48):41183-41194.
[19] TSAY YF, CHIU CC, TSAI CB, et al. Nitrate transporters and peptide transporters[J]. FEBS Letters, 2007,581(12):2290-2300.
[20] LUCIC E, FOURREY C, KOHLER A,et al. A gene repertoire for nitrogen transporters in Laccaria bicolor[J]. New Phytologist, 2008, 180(2):343-364.
[21] REUSS O, MORSCHHAUSER J. A family of oligopeptide transporters is required for growth of Candida albicans on proteins[J]. Molecular Microbiology, 2006, 60(3):795-812.