| 659 | 1 | 83 |
| 下载次数 | 被引频次 | 阅读次数 |
在植物中，GRAS转录因子通过参与光信号通路和赤霉素通路等多种信号传导途径来响应植物非生物胁迫过程已被广泛研究，但对其在生物胁迫中的响应机制研究较少。本研究基于新疆野苹果（Malus sieversii）基因组，通过TBtools、Pfam和NCBI-CDD分析软件，对新疆野苹果GRAS家族基因（MsGRAS）进行筛选与验证；随后利用Protein Paramter Calc分析MsGRAS家族成员蛋白的理化性质，用MEGA构建系统发育树；最后利用PlantCARE和Graphics软件分析MsGRAS的2 000 bp顺式作用元件。此外，分析MsGRAS基因与栽培苹果（Malus domesitica）MdGRAS基因的共线性关系。通过腐烂病菌（Valsa mali）侵染后的全长转录组数据和RT-qPCR实验对MsGRAS基因家族在响应新疆野苹果抵抗腐烂病菌过程中的表达模式进行分析和验证。结果显示：（1）在新疆野苹果中共鉴定获得83个GRAS基因，涵盖DELLA、LS、SCR、SCL3和DLT等11个亚家族；蛋白结构域预测结果表明，83个MsGRAS蛋白均包含GRAS结构域，其中有4个蛋白单独具有DELLA结构域。顺式作用元件分析显示，81个MsGRAS基因的启动子区域含有多种激素相关元件。MsGRAS基因家族共线性分析结果显示，65个MsGRAS基因和68个MdGRAS基因之间存在152对共线性关系，在栽培苹果的驯化过程中丢失了18个MsGRAS基因。（2）RNA-seq数据表明，有18个MsGRAS基因在腐烂病感染过程中差异表达，其中LISCL亚家族中差异表达基因数量最多（6个）。（3）RT-qPCR结果表明，共有6个MsGRAS基因在新疆野苹果抵抗腐烂病感染过程中显著差异表达，其中包括4个上调的MsGRAS基因，主要为LISCL亚家族成员，并且PAT1亚家族成员Ms02G03400在腐烂病菌侵染5天时，其表达量上调50倍。综上结果表明，PAT1和LISCL亚家族成员在新疆野苹果抗腐烂病过程中可能发挥重要作用，可作为后续抗病基因研究的候选基因。该研究结果为新疆野苹果GRAS基因响应腐烂病菌侵染的功能和机理研究奠定了基础。
Abstract：In plants, GRAS transcription factors have been widely studied in response to abiotic stress processes by participating in various signaling pathways such as light signaling pathway and gibberellin pathway, but their response mechanisms in biological stress have been less studied. Research on GRAS transcription factor has a long tradition. There has been less previous evidence for mechanisms of biological stress. Based on the genome of Malus sieversii, GRAS family gene(MsGRAS) of M. sieversii was screened and verified by TBtools, Pfam and NCBI-CDD analysis software. Then Protein Paramter Calc was used to analyze the physical and chemical properties of MsGRAS family members proteins, and MEGA was used to construct phylogenetic tree. Finally, PlantCARE and Graphics software were used to analyze the 2 000 bp cis-acting elements of MsGRAS. In addition, the collinearity of the MsGRAS gene to the MdGRAS gene in M. domesitica was analyzed. The full-length transcriptome data and RT-qPCR assay were used to analyze and verify the expression patterns of MsGRAS gene family in response to the resistance of Malus sieversii to Valsa mali infection.The full-length transcriptome data and RT-qPCR assay were used to analyze and verify the expression patterns of MsGRAS gene family in response to the resistance of Malus sieversii to Valsa mali. The results show that:(1) A total of 83 GRAS genes were identified in Malus sieversii including 11 subfamilies such as DELLA, LS, SCR, SCL3 and DLT subfamily. The results of protein domain prediction showed that all 83 MsGRAS proteins contained GRAS domains, among which four proteins had DELLA domains alone. Cis-acting element analysis showed that the promoter regions of 81 MsGRAS genes contained multiple hormone-related elements. The collinear analysis of MsGRAS gene family showed that there were 152 pairs of collinear relationships between 65 MsGRAS genes and 68MdGRAS genes. 18 MsGRAS genes were lost during the domestication of cultivated apples.(2) RNA-seq data showed that 18 MsGRAS genes were differentially expressed during canker disease infection, among which the LISCL subfamily had the largest number of differently expressed genes(6).(3) RT-qPCR results showed that six MsGRAS genes were significantly differently expressed during the resistance to canker disease in M. sieversii. Including four upregulated MsGRAS genes, which were mainly members of the LISCL subfamily, and PAT1 subfamily member Ms02G03400(50-fold increase) was up-regulaed under the Valsa mali infection at 5 dpi. In conclusion, PAT1 and LISCL may play an important role in the resistance of M. sieversii to canker disease, and can be used as candidate genes for future studies on resistance genes. The results of this study laid a foundation for the study on the function and mechanism of GRAS gene response to Valsa mali in M. sieversii.
胡小平，张吉光，陈婧，等，2010．新疆野苹果和秦冠的抗黑星病特性．植物病理学报，40(6):609-614.[HUX P,ZHANG J G,CHEN J,et al.,2010.Resistance characteristics of Xinjiang wild apple and Qinguan cultivars to apple scab.Acta Phytopathologica Sinica,40(6):609-614.]
柯希望，2013．黑腐皮壳侵染苹果的组织细胞学及转录组学研究，博士学位论文．杨凌：西北农林科技大学．[KE X W,2013.Histocytology and transcriptomics studies on infection of Malus domestica cv.Fuji by Valsa mali,Dissertation for Ph.D.Yangling:Northwest A&F University.]
刘晓洁，李小双，张燕，等，2019．划布轮法鉴定新疆野苹果树腐烂病抗性方法的建立．西北农业学报，28(4):674-679.[LIU X J,LI X S,ZHANG Y,et al.,2019.The pattern wheel innoculation method used to identify resistance of Xinjiang wild apple trees(Malus sieversii)to Valsa canker.Acta Agriculturae Boreali-occidentalis Sinica,28(4):674-679.]
于婷婷，李欢，宁源生，等，2023．苹果GRAS全基因组鉴定及其对生长素的响应分析．园艺学报，50(2):397-409.[YU T T,LI H,NING Y S,et al.,2023.Genome-wide identification of GRAS gene family in apple and expression analysis of its response to auxin.Acta Horticulturae Sinica,50(2):397-409.]
张艳敏，冯涛，张春雨，等，2009．新疆野苹果研究进展．园艺学报，36(3):447-452.[ZHANG Y M,FENG T,ZHANG C Y,et al.,2009.Advances in research of the Malus sieversii(Lebed.)Roem.Acta Horticulturae Sinica,36(3):447-452.]
BALLESTER A R,NORELLI J,BURCHARD E,et al.,2017.Transcriptomic response of resistant(PI613981-Malus sieversii)and susceptible("royal gala")genotypes of apple to blue mold(Penicillium expansum)infection.Front.Plant Sci.,8:1981.
CHEN C,LU L L,MA S Y,et al.,2023.Analysis of PAT1subfamily members in the GRAS family of upland cotton and functional characterization of GhSCL13-2A in Verticilli‐um dahliae resistance.Plant Cell Rep.,42(3):487-504.
CHEN K,RAJEWSKY N,2007.The evolution of gene regulation by transcription factors and microRNAs.Nat.Rev.Genet.,8(2):93-103.
COMINGS D E,RIGGS A D,1971.Molecular mechanisms of chromosome pairing,folding and function.Nature,233(5314):48-50.
DUAN N B,BAI Y,SUN H H,et al.,2017.Genome re-sequencing reveals the history of apple and supports a two-stage model for fruit enlargement.Nat.Commun.,8:249.
FAN S,ZHANG D,GAO C,et al.,2017.Identification,classification,and expression analysis of GRAS gene family in Malus domestica.Front.Physiol.,8:253.
FODE B,SIEMSEN T,THUROW C,et al.,2008.The Arabi‐dopsis GRAS protein SCL14 interacts with classⅡTGAtranscription factors and is essential for the activation of stress-induciblepromoters.PlantCell,20(11):3122-3135.
HARSHMAN J M,EVANS K M,ALLEN H,et al.,2017.Fire blight resistance in wild accessions of Malus sieversii.Plant Dis.,101(10):1738-1745.
HIRSCH S,OLDROYD G E D,2009.GRAS-domain transcription factors that regulate plant development.Plant Signal.Behav.,4(8):698-700.
HUANG W,XIAN Z Q,KANG X,et al.,2015.Genome-wide identification,phylogeny and expression analysis of GRASgene family in tomato.BMC Plant Biol.,15(1):1-18.
KIDOKORO S,SHINOZAKI K,YAMAGUCHI-SHINOZAKIK,2022.Transcriptional regulatory network of plant cold-stress responses.Trends Plant Sci.,27(9):922-935.
LEE M H,KIM B,SONG S K,et al.,2008.Large-scale analysis of the GRAS gene family in Arabidopsis thaliana.Plant Mol.Biol.,67(6):659-670.
LIU X J,LI X S,BOZOROV T A,et al.,2020.Characterization and pathogenicity of six Cytospora strains causing stem canker of wild apple in the Tianshan Forest,China.For.Path.,50(3):e12587.
LIU X J,LI X S,WEN X J,et al.,2021.PacBio full-length transcriptome of wild apple(Malus sieversii)provides insights into canker disease dynamic response.BMC Genom.,22(1):52.
LIU Y F,WANG W,2021.Characterization of the GRAS gene family reveals their contribution to the high adaptability of wheat.PeerJ,9:e10811.
LIVAK K J,SCHMITTGEN T D,2001.Analysis of relative gene expression data using real-time quantitative PCR and the 2-ΔΔCTmethod.Methods,25(4):402-408.
LU L,DIAO Z J,YANG D W,et al.,2022.The 14-3-3 protein GF14c positively regulates immunity by modulating the protein homoeostasis of the GRAS protein OsSCL7 in rice.Plant Cell Environ.,45(4):1065-1081.
MA H S,LIANG D,SHUAI P,et al.,2010.The salt-and drought-inducible poplar GRAS protein SCL7 confers salt and drought tolerance in Arabidopsis thaliana.J.Exp.Bot.,61(14):4011-4019.
NAVARRO L,BARI R,ACHARD P,et al.,2008.DELLAs control plant immune responses by modulating the balance of jasmonic acid and salicylic acid signaling.Curr.Biol.,18(9):650-655.
PENG X,FENG C,WANG Y T,et al.,2022.miR164gMsNAC022 acts as a novel module mediating drought response by transcriptional regulation of reactive oxygen species scavenging systems in apple.Hortic.Res.,9:uhac192.
SUN X P,JIAO C,SCHWANINGER H,et al.,2020.Phased diploid genome assemblies and pan-genomes provide insights into the genetic history of apple domestication.Nat.Genet.,52(12):1423-1432.
TIAN C G,WAN P,SUN S H,et al.,2004.Genome-wide analysis of the GRAS gene family in rice and Arabidopsis.Plant Mol.Biol.,54(4):519-532.
TO V T,SHI Q,ZHANG Y Y,et al.,2020.Genome-wide analysis of the GRAS gene family in barley(Hordeum vul‐gare L.).Genes,11(5):553.
WANG L,DING X L,GAO Y Q,et al.,2020.Genome-wide identification and characterization of GRAS genes in soybean(Glycine max).BMC Plant Biol.,20(1):415.
WANG X Y,MANZOOR M A,WANG M N,et al.,2022.Integrative analysis of the GRAS genes from Chinese white pear(Pyrus bretschneideri):a critical role in leaf regeneration.Front.Plant Sci.,13:898786.
WANG Y X,LIU Z W,WU Z J,et al.,2018.Genome-wide identification and expression analysis of GRAS family transcription factors in tea plant(Camellia sinensis).Sci.Rep.,8:3949.
YANG M L,ZHANG Y X,ZHANG H H,et al.,2017.Identification of MsHsp20 gene family in Malus sieversii and functional characterization of MsHsp16.9 in heat tolerance.Front.Plant Sci.,8:1761.
ZHANG H A,MI L M,XU L,et al.,2019.Genome-wide identification,characterization,interaction network and expression profile of GRAS gene family in sweet orange(Cit‐rus sinensis).Sci.Rep.,9:2156.
ZHANG Y,BOZOROV T A,LI D X,et al.,2020.An efficient in vitro regeneration system from different wild apple(Malus sieversii)explants.Plant Meth.,16(1):56.
基本信息:
DOI：10.13417/j.gab.042.000808
中图分类号:S436.61
引用信息:
[1]张雪纯,赵明奇,贾珊珊,等.新疆野苹果GRAS转录因子家族全基因组鉴定与响应腐烂病菌侵染的表达分析[J].基因组学与应用生物学,2023,42(08):808-820.DOI:10.13417/j.gab.042.000808.
基金信息:
国家自然科学基金地区联合重点项目（U1903206）和国家自然科学基金地区科学基金项目（32160074）共同资助
2023-06-04
2023-06-04
2023-06-04