玉米ZmcpSRP54基因可变剪接分析
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摘要
cpSRP54基因在植物叶绿体发育中发挥重要作用。本研究用生物信息学方法对玉米ZmcpSRP54蛋白进行了进化分析,利用RT-PCR方法对玉米ZmcpSRP54基因全长cDNA进行了克隆,进而对其进行可变剪接分析。结果表明,玉米ZmcpSRP54蛋白与其它7个物种的cpSRP54蛋白有很高的相似性。鉴定到玉米ZmcpSRP54基因25个不同转录本,1个为标准剪接转录本,其它24个为可变剪接转录本。24个可变剪接转录本共使用了17种非标准剪接位点,4种可变剪接方式,主要以可变的3′端位点、可变的5′端位点和外显子跳跃3种可变剪接方式为主。预测出18种不同长度的ORF,编码18种不同长度的多肽。11个多肽其保守结构域氨基酸序列发生部分缺失,6个多肽其保守结构域氨基酸序列发生全部缺失,1个编码全新的多肽。这些结果将为玉米ZmcpSRP54基因的功能研究提供重要信息。
The cpSRP54 gene plays an important role during chloroplast development in plant. In this study, the bioinformatics method was used for the evolutionary analysis of maize ZmcpSRP54 protein, the RT-PCR method was used for the cloning of full-length cDNA of maize ZmcpSRP54 gene, and then the bioinformatics methods were used for the analysis of alternative splicing. Phylogenetic analysis revealed that the maize ZmcpSRP54 protein was conserved with those cpSRP54 proteins in the other seven species. A total of 25 different transcripts of ZmcpSRP54 were obtained, and 1 was a standard splicing transcript and the other 24 were alternative splicing(AS) transcripts. A total of 17 non-standard splice sites and 4 AS forms were used for the 24 AS transcripts, and alternative 3′ splice site, alternative 5′ splice site and exon skipping were the most prevalent AS forms. A total of 18 ORFs with different lengths were predicted for these 24 AS transcripts, encoding 18 polypeptides with different lengths, and 11 of which had part of the amino acid sequences in the conserved region deleted, 6 of which had all of the amino acid sequences in the conserved region deleted, while 1 of which encoded another new polypeptide. These results would provide important information for the functional study of the ZmcpSRP54 gene in maize.
The cpSRP54 gene plays an important role during chloroplast development in plant. In this study, the bioinformatics method was used for the evolutionary analysis of maize ZmcpSRP54 protein, the RT-PCR method was used for the cloning of full-length cDNA of maize ZmcpSRP54 gene, and then the bioinformatics methods were used for the analysis of alternative splicing. Phylogenetic analysis revealed that the maize ZmcpSRP54 protein was conserved with those cpSRP54 proteins in the other seven species. A total of 25 different transcripts of ZmcpSRP54 were obtained, and 1 was a standard splicing transcript and the other 24 were alternative splicing(AS) transcripts. A total of 17 non-standard splice sites and 4 AS forms were used for the 24 AS transcripts, and alternative 3′ splice site, alternative 5′ splice site and exon skipping were the most prevalent AS forms. A total of 18 ORFs with different lengths were predicted for these 24 AS transcripts, encoding 18 polypeptides with different lengths, and 11 of which had part of the amino acid sequences in the conserved region deleted, 6 of which had all of the amino acid sequences in the conserved region deleted, while 1 of which encoded another new polypeptide. These results would provide important information for the functional study of the ZmcpSRP54 gene in maize.
引文
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[17]Seo P J,Park M J,Park C M.Alternative splicing of transcription factors in plant responses to low temperature stress:mechanisms and functions[J].Planta,2013,237(6):1415-1424.
[18]Wang Z F,Burge C B.Splicing regulation:from aparts list of regulatory elements to an integrated splicingcode[J].RNA,2008,14(5):802-813.
[19]Marquez Y,Brown J W S,Simpson C,et al.Comprehensive analysis of alternative splicing in rice and comparative analyses with Arabidopsis[J].Genome Res.,2012,22(6):1184-1195.
[20]Rauch H B,Patrick T L,Klusman K M,et al.Discovery and expression analysis of alternative splicing events conserved among plant SR proteins[J].Mol.Biol.Evol.,2014,31(3):605-613.
[21]E Z G,Wang L,Zhou J H.Splicing and alternative splicing in rice and humans[J].BMB Reports,2013,46(9):439-447.
[22]Reddy A S N,Marquez Y,Kalyna M,et al.Complexity of the alternative splicing landscape in plants[J].Plant Cell,2013,25:3657-3683.
[23]Chen Q Y,Han Y J,Liu H J,et al.Genome-wide association analyses reveal the importance of alternative splicing in diversifying gene function and regulating phenotypic variation in maize[J].Plant Cell,2018,30(7):1404-1423.
[24]Lopez A.Alternative splicing of pre-mRNA:developmental consequences and mechanisms of regulation[J].Annu.Rev.Genet.,1998,32:279-305.
[25]Gupta S,Wang B B,Stryker G A,et al.Two novel arginine/serine(SR)proteins in maize are differentially spliced and utilize non-canonical splice sites[J].Biochimica et Biophysica Acta,2005,1728:105-111.
[2]Tzvetkova-Chevolleau T,Hutin C,Noel L D,et al.Canonical signal recognition particle components can be bypassed for posttranslational protein targeting in chloroplasts[J].Plant Cell,2007,19:1635-1648.
[3]林添资,孙立亭,景德道,等.一个水稻黄绿叶突变体ygl14(t)的鉴定及基因定位[J].核农学报,2018,32(2):216-226.
[4]Amin P,Sy D A C,Pilgrim M L,et al.Arabidopsis mutants lacking the 43-and 54-kilodalton subunits of the chloroplast signal recognition particle have distinct phenotypes[J].Plant Physiol.,1999,121(1):61-70.
[5]Zhang F T,Luo X D,Hu B L,et al.YGL138(t),encoding a putative signal recognition particle 54 k Da protein,is involved in chloroplast development of rice[J].Rice,2013,6:7.
[6]邢永强,刘国庆,蔡禄.Pre-mRNA选择性剪接的调控及选择性剪接数据库[J].中国生物化学与分子生物学报,2016,32(1):17-28.
[7]Luco R F,Allo M,Schor I E,et al.Epigenetics in alternative pre-mRNA splicing[J].Cell,2011,144(1):16-26.
[8]Tress M L,Abascal F,Valencia A.Alternative splicing may not be the key to proteome complexity[J].Trends Biochem.Sci.,2017,42:98-110.
[9]Pan Q,Shai O,Lee L J,et al.Deep surveying of alternative splicing complexity in the human transcriptome by highthroughput sequencing[J].Nat.Genet.,2008,40:1413-1415.
[10]Werneke J M,Chatfield J M,Ogren W L.Alternative mRNAsplicing genernatesthe two ribulose bisphosphate carboxylase/oxygenase activase polypeptides in spinach and Arabidopsis[J].Plant Cell,1989,1:815.
[11]Syed N H,Kalyna M,Marquez Y,et al.Alternative splicing in plants-coming of age[J].Trends Plant Sci.,2012,17:616.
[12]Barbazuk W B,Yan F,Mc Ginnis K M.Genome-wide analyses of alternative splicing in plants:opportunities and challenges[J].Genome Res.,2008,18(9):1381-1392.
[13]Xu S H,Zhang Z B,Jing B B,et al.Transportin-SR is required for proper splicing of resistance genes and plant immunity[J].PLo S Genet.,2011,7:e1002159.
[14]Mastrangelo A M,Marone D,LaidòG,et al.Alternative splicing:enhancing ability to cope with stress via transcriptome plasticity[J].Plant Sci.,2012,185:40-49.
[15]Dubrovina A S,Kiselev K V,Zhuravlev Y N.The role of canonical and noncanonical pre-mRNA splicing in plant stress responses[J].Biomed.Res.Int.,2013,2013:264314.
[16]Carvalho R F,Feij2o C V,Duque P.On the physiological significance of alternative splicing events in higher plants[J].Protoplasma,2013,250(3):639-650.
[17]Seo P J,Park M J,Park C M.Alternative splicing of transcription factors in plant responses to low temperature stress:mechanisms and functions[J].Planta,2013,237(6):1415-1424.
[18]Wang Z F,Burge C B.Splicing regulation:from aparts list of regulatory elements to an integrated splicingcode[J].RNA,2008,14(5):802-813.
[19]Marquez Y,Brown J W S,Simpson C,et al.Comprehensive analysis of alternative splicing in rice and comparative analyses with Arabidopsis[J].Genome Res.,2012,22(6):1184-1195.
[20]Rauch H B,Patrick T L,Klusman K M,et al.Discovery and expression analysis of alternative splicing events conserved among plant SR proteins[J].Mol.Biol.Evol.,2014,31(3):605-613.
[21]E Z G,Wang L,Zhou J H.Splicing and alternative splicing in rice and humans[J].BMB Reports,2013,46(9):439-447.
[22]Reddy A S N,Marquez Y,Kalyna M,et al.Complexity of the alternative splicing landscape in plants[J].Plant Cell,2013,25:3657-3683.
[23]Chen Q Y,Han Y J,Liu H J,et al.Genome-wide association analyses reveal the importance of alternative splicing in diversifying gene function and regulating phenotypic variation in maize[J].Plant Cell,2018,30(7):1404-1423.
[24]Lopez A.Alternative splicing of pre-mRNA:developmental consequences and mechanisms of regulation[J].Annu.Rev.Genet.,1998,32:279-305.
[25]Gupta S,Wang B B,Stryker G A,et al.Two novel arginine/serine(SR)proteins in maize are differentially spliced and utilize non-canonical splice sites[J].Biochimica et Biophysica Acta,2005,1728:105-111.
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