利用酵母双杂交系统筛选与大豆花叶病毒核内含体蛋白互作的大豆寄主因子

doi:10.3864/j.issn.0578-1752.2025.19.001

摘要/Abstract

摘要：

【目的】由大豆花叶病毒（soybean mosaic virus，SMV）引起的大豆花叶病毒病是大豆最具危害的病毒性病害之一，严重影响大豆的产量和品质。利用酵母文库筛选与SMV核内含体蛋白（NIa-Pro和NIb）互作的寄主蛋白，为深入解析SMV的侵染机制及大豆抗病机制提供理论依据及新思路。【方法】首先，从大豆花叶病毒病株系SMV-HN中分别克隆NIa-Pro和NIb，将其重组至pGBKT7酵母载体以构建诱饵质粒，通过酵母双杂交文库筛选技术鉴定与2个病毒功能蛋白互作的大豆寄主蛋白。其次，克隆编码外膜孔蛋白16（outer envelope pore protein 16-1，OEP16）的寄主蛋白基因GmOEP16，并通过酵母双杂交（Y2H）和荧光素酶互补试验（LCA）明确GmOEP16与NIa-Pro和NIb的互作关系。再次，利用荧光定量PCR分析GmOEP16在SMV侵染和外源激素诱导下的表达模式。最后，采用病毒诱导的基因瞬时沉默技术（VIGS）对GmOEP16在SMV病程反应中的功能进行验证。【结果】成功构建pGBKT7-NIa-Pro和pGBKT7-NIb诱饵质粒，分别筛选到12个与NIa-Pro和NIb互作的大豆寄主蛋白。利用Y2H试验，验证NIa-Pro与GmOEP16和GmDEG5，以及NIb与GmOEP16、GmZC3H18和GmAHP1存在相互作用；进一步通过LCA试验，明确GmOEP16与NIa-Pro和NIb均可发生相互作用。表达模式分析显示，GmOEP16可以响应SMV的诱导，并可在反应初期快速地响应水杨酸和脱落酸的诱导。VIGS试验表明，有效沉默GmOEP16后，相较于对照组，沉默植株叶片没有出现明显的感病症状，且SMV外壳蛋白（coat protein，CP）基因SMV-CP的表达量显著降低，即大豆增强了对SMV的抗性，说明该基因可负调控大豆对SMV的抗性。【结论】成功构建了pGBKT7-NIa-Pro和pGBKT7-NIb诱饵载体，并各筛选得到12个与它们互作的大豆寄主蛋白。其中，GmOEP16与SMV核内含体蛋白NIa-Pro和NIb存在相互作用；GmOEP16可响应SMV的诱导且负调控SMV抗性，促进SMV的侵染。

关键词: 大豆花叶病毒, 核内含体蛋白, 酵母文库筛选, 外膜孔蛋白16, 病毒诱导的基因沉默

Abstract:

【Objective】Soybean mosaic virus (SMV) is one of the most damaging viral diseases of soybean, which seriously affects soybean yield and quality. Identification of host proteins interacting with SMV nuclear inclusion proteins (NIa-Pro and NIb) using yeast two-hybrid library screening, aiming to establish a theoretical foundation and propose novel perspectives insights into the molecular mechanisms of SMV infection and soybean resistance.【Method】Firstly, the coding sequences of NIa-Pro and NIb were cloned from the SMV strain SMV-HN and recombined into the pGBKT7 vector to construct the bait plasmids, and then soybean proteins interacting with the two viral functional proteins were identified by yeast library screening. Secondly, the host gene GmOEP16 encoding Outer Envelope Pore Protein 16 (OEP16) was cloned, and the interactions of GmOEP16 with NIa-Pro and NIb were clarified by yeast two-hybrid (Y2H) and luciferase complementation assay (LCA). Quantitative real-time PCR (qRT-PCR) was used to analyse the expression pattern of GmOEP16 under SMV treatment and exogenous hormone induction. Finally, virus-induced gene silencing (VIGS) was used to validate the function of GmOEP16 gene in SMV disease response.【Result】pGBKT7-NIa-Pro and pGBKT7-NIb recombinant plasmids were successfully constructed, and 12 soybean host proteins were screened for interactions with NIa-Pro and NIb, respectively. The Y2H assay was further used to verify that NIa-Pro interacted with GmOEP16 and GmDEG5, and NIb interacted with GmOEP16, GmZC3H18 and GmAHP1. The LCA assay was further used to clarify that GmOEP16 interacted with both NIa-Pro and NIb. Expression analysis revealed that GmOEP16 was induced by SMV infection and responded rapidly to salicylic acid (SA) and abscisic acid (ABA) stimuli during early response. The VIGS assay showed that effectively silencing of GmOEP16 resulted in no obvious susceptibility phenotype in leaf tissues relative to the wild-type controls. Meanwhile, the expression of SMV-CP was significantly reduced in the GmOEP16-silenced plants, suggesting that the soybean resistance to SMV was enhanced. Collectively, these findings demonstrated that GmOEP16 could function as a negative regulator of SMV resistance in soybean.【Conclusion】The pGBKT7-NIa-Pro and pGBKT7-NIb bait vectors were successfully constructed, and each 12 soybean host proteins that respectively interacted with pGBKT7-NIa-Pro and pGBKT7-NIb were identified. Among them, GmOEP16 interacted with both NIa-Pro and NIb. GmOEP16 responded to SMV induction and negatively regulated SMV resistance, which promoted SMV infection on soybeans.

Key words: soybean mosaic virus, nuclear inclusion proteins, yeast library screening, outer envelope pore protein 16, virus-induced gene silencing

于哲, 周芳雪, 刘润发, 田雅琦, 吉好木哈, 王勇翔, 冯雯蜜, 牟可欣, 井妍, 李海燕. 利用酵母双杂交系统筛选与大豆花叶病毒核内含体蛋白互作的大豆寄主因子[J]. 中国农业科学, 2025, 58(19): 3799-3813.

YU Zhe, ZHOU FangXue, LIU RunFa, TIAN YaQi, JIHAO MuHa, WANG YongXiang, FENG WenMi, MOU KeXin, JING Yan, LI HaiYan. Screening for Soybean Host Factors that Interact with Soybean Mosaic Virus Nuclear Inclusion Proteins Using the Yeast Two-Hybrid System[J]. Scientia Agricultura Sinica, 2025, 58(19): 3799-3813.

0
/ / 推荐

导出引用管理器 EndNote|Reference Manager|ProCite|BibTeX|RefWorks

链接本文: https://www.chinaagrisci.com/CN/10.3864/j.issn.0578-1752.2025.19.001

https://www.chinaagrisci.com/CN/Y2025/V58/I19/3799

图/表 10

图1

表1

图2

表2

图3

图4

图5

表3

GmOEP16启动子中的部分顺式作用元件"

元件 Element	序列 Sequences	作用 Function
HD-Zip 1	TAAT	参与叶绿体中层细胞分化的元素 Element involved in differentiation of the palisade mesophyll cells
ARE	ATTG	厌氧诱导所必需的顺式作用调控元件 Cis-acting regulatory element essential for the anaerobic induction
TCT-motif	TCTTAC	光敏元件的一部分 Part of a light responsive element
I-box	TATAATAAT	光敏元件的一部分 Part of a light responsive element
AE-box	AGAAACTT	光响应模块的一部分 Part of a module for light response
Gap-box	CAAATGAA(A/G)A	光敏元件的一部分 Part of a light responsive element
TATA-box	TATA(A/T)A(A/T)	转录起点-30附近的核心启动子元件 Core promoter element around -30 of transcription start
GATA-motif	AAGGATAAGG	光敏元件的一部分 Part of a light responsive element
TC-rich repeats	ATTCTCTAAC	参与防御和应激反应的顺式作用元件 Cis-acting element involved in defense and stress responsiveness
ABRE	CGCACGTGTC	参与脱落酸反应的顺式作用元件 Cis-acting element involved in the abscisic acid responsiveness
AT1-motif	AATTATTTTTTATT	光响应模块的一部分 Part of a light responsive module
TCA-element	CCATCTTTTT	参与水杨酸反应的顺式作用元件 Cis-acting element involved in salicylic acid responsiveness
GCN4_motif	TGAGTCA	参与胚乳表达的顺式调控元件 Cis-regulatory element involved in endosperm expression
MBS	CAACTG	涉及干旱诱导性的MYB结合位点 MYB binding site involved in drought-inducibility
Box 4	ATTAAT	参与光响应的保守DNA模块的一部分 Part of a conserved DNA module involved in light responsiveness
CAAT-box	CAAT	启动子和增强子区域的共同顺式作用元件 Common cis-acting element in promoter and enhancer regions
MRE	AACCTAA	参与光响应的MYB结合位点 MYB binding site involved in light responsiveness

表3

图6

图7

参考文献 62

[1]	LIU Y C, DU H L, LI P C, SHEN Y T, PENG H, LIU S L, ZHOU G A, ZHANG H K, LIU Z, SHI M, et al. Pan-genome of wild and cultivated soybeans. Cell, 2020, 182(1): 162-176.e13. doi: S0092-8674(20)30618-8 pmid: 32553274
[2]	WHITHAM S A, QI M S, INNES R W, MA W B, LOPES-CAITAR V, HEWEZI T. Molecular soybean-pathogen interactions. Annual Review of Phytopathology, 2016, 54: 443-468. doi: 10.1146/annurev-phyto-080615-100156 pmid: 27359370
[3]	CHENG R X, MEI R X, YAN R, CHEN H Y, MIAO D, CAI L N, FAN J Y, LI G R, XU R, LU W G, et al. A new distinct geminivirus causes soybean stay-green disease. Molecular Plant, 2022, 15(6): 927-930. doi: 10.1016/j.molp.2022.03.011 pmid: 35358702
[4]	XU F, FENG C H, LIU L L, SHI R J, HAN S, SONG Y L, WANG J M, HAN Z H, ZHANG J J, LI Y H, et al. First report of Fusarium falciforme causing root rot of soybean (Glycine max) in Henan, China. Plant Disease, 2023, 107(7): 2244.
[5]	MALAPI-NELSON M, WEN R H, OWNLEY B H, HAJIMORAD M R. Co-infection of soybean with soybean mosaic virus and alfalfa mosaic virus results in disease synergism and alteration in accumulation level of both viruses. Plant Disease, 2009, 93(12): 1259-1264.
[6]	WEI Z, JIANG C, MAO C, ZHANG H, MIAO R, CHEN J, SUN Z. Occurrence of soybean yellow common mosaic virus in soybean in China showing yellow common mosaic disease. Plant Disease, 2021, 105(4): 1236.
[7]	YANG Y Q, LIN J, ZHENG G J, ZHANG M C, ZHI H J. Recombinant soybean mosaic virus is prevalent in Chinese soybean fields. Archives of Virology, 2014, 159(7): 1793-1796. doi: 10.1007/s00705-014-1980-z pmid: 24445813
[8]	王大刚, 黄志平, 杨勇, 李杰坤, 吴倩. 大豆花叶病毒抗性基因及分子标记研究进展. 植物遗传资源学报, 2024, 25(6): 882-897.
	WANG D G, HUANG Z P, YANG Y, LI J K, WU Q. Progress on studies of resistance genes and molecular markers of soybean mosaic virus. Journal of Plant Genetic Resources, 2024, 25(6): 882-897. (in Chinese)
[9]	ISHIBASHI K, SARUTA M, SHIMIZU T, SHU M, ANAI T, KOMATSU K, YAMADA N, KATAYOSE Y, ISHIKAWA M, ISHIMOTO M, et al. Soybean antiviral immunity conferred by dsRNase targets the viral replication complex. Nature communications, 2019, 10(1): 4033. doi: 10.1038/s41467-019-12052-5 pmid: 31562302
[10]	ZHANG P P, DU H Y, WANG J, PU Y X, YANG C Y, YAN R J, YANG H, CHENG H, YU D Y. Multiplex CRISPR/Cas9-mediated metabolic engineering increases soya bean isoflavone content and resistance to soya bean mosaic virus. Plant Biotechnology Journal, 2020, 18(6): 1384-1395. doi: 10.1111/pbi.13302 pmid: 31769589
[11]	ADAMS M J, ANTONIW J F, BEAUDOIN F. Overview and analysis of the polyprotein cleavage sites in the family Potyviridae. Molecular Plant Pathology, 2005, 6(4): 471-487. doi: 10.1111/j.1364-3703.2005.00296.x pmid: 20565672
[12]	LIU J Z, FANG Y, PANG H X. The current status of the soybean- Soybean mosaic virus (SMV) pathosystem. Frontiers in Microbiology, 2016, 7: 1906.
[13]	李文霞. 大豆花叶病毒组分蛋白6K1、NIa-VPg、NIa-Pro、NIb和CP的互作蛋白鉴定与分析[D]. 呼和浩特: 内蒙古大学, 2023.
	LI W X. Components of soybean mosaic virus protein 6K1, NIa-VPg, NIa-pro, NIb and CP interactions of protein identification and analysis[D]. Hohhot: Inner Mongolia University, 2023. (in Chinese)
[14]	宋英培. 影响大豆花叶病毒传播的因素及抗SMV相关基因的表达与定位分析[D]. 南京: 南京农业大学, 2015.
	SONG Y P. Factors affecting smv transmission and analysis of smv resistance related genes[D]. Nanjing: Nanjing Agricultural University, 2015. (in Chinese)
[15]	NUNNA H, QU F, TATINENI S. P3 and NIa-pro of turnip mosaic virus are independent elicitors of superinfection exclusion. Viruses, 2023, 15(7): 1459.
[16]	CASTEEL C L, YANG C L, NANDURI A C, DE JONG H N, WHITHAM S A, JANDER G. The NIa-Pro protein of Turnip mosaic virus improves growth and reproduction of the aphid vector, Myzus persicae (green peach aphid). The Plant Journal, 2014, 77(4): 653-663. doi: 10.1111/tpj.12417 pmid: 24372679
[17]	ANINDYA R, SAVITHRI H S. Potyviral NIa proteinase, a proteinase with novel deoxyribonuclease activity. Journal of Biological Chemistry, 2004, 279(31): 32159-32169. doi: 10.1074/jbc.M404135200 pmid: 15163663
[18]	KOZIEŁ E, SUROWIECKI P, PRZEWODOWSKA A, BUJARSKI J J, OTULAK-KOZIEŁ K. Modulation of expression of PVY^NTN RNA-dependent RNA polymerase (NIb) and heat shock cognate host protein HSC70 in susceptible and hypersensitive potato cultivars. Vaccines, 2021, 9(11): 1254.
[19]	SHEN W T, SHI Y, DAI Z J, WANG A M. The RNA-dependent RNA polymerase NIb of potyviruses plays multifunctional, contrasting roles during viral infection. Viruses, 2020, 12(1): 77.
[20]	DU K T, PENG D Z, WU J Q, ZHU Y B, JIANG T, WANG P, CHEN X, JIANG S J, LI X D, CAO Z Y, FAN Z F, ZHOU T. Maize splicing-mediated mRNA surveillance impeded by sugarcane mosaic virus-coded pathogenic protein NIa-Pro. Science Advances, 2024, 10(34): eadn3010.
[21]	JIA Z X, RUI P H, FANG X X, HAN K L, YU T Q, LU Y W, ZHENG H Y, CHEN J P, YAN F, WU G W. Proteolysis of host DEAD-box RNA helicase by potyviral proteases activates plant immunity. New Phytologist, 2025, 245(4): 1655-1672. doi: 10.1111/nph.20318 pmid: 39611543
[22]	GE L H, JIA M X, SHAN H Y, GAO W F, JIANG L, CUI H G, CHENG X F, UZEST M, ZHOU X P, WANG A M, et al. Viral RNA polymerase as a SUMOylation decoy inhibits RNA quality control to promote potyvirus infection. Nature Communications, 2025, 16: 157.
[23]	GAO L, LUO J, DING X, WANG T, HU T, SONG P, ZHAI R, ZHANG H, ZHANG K, LI K, ZHI H. Soybean RNA interference lines silenced for eIF4E show broad potyvirus resistance. Molecular plant pathology, 2020, 21(3): 303-317. doi: 10.1111/mpp.12897 pmid: 31860775
[24]	BWALYA J, WIDYASARI K, VÖLZ R, KIM K H. Chloroplast- related host proteins interact with NIb and NIa-Pro of soybeans mosaic virus and induce resistance in the susceptible cultivar. Virus Research, 2023, 336: 199205.
[25]	LUO J, ZHOU J J, ZHANG J Z. Aux/IAA gene family in plants: Molecular structure, regulation, and function. International Journal of Molecular Sciences, 2018, 19(1): 259.
[26]	JOHNSON R R, WAGNER R L, VERHEY S D, WALKER- SIMMONS M K. The abscisic acid-responsive kinase PKABA1 interacts with a seed-specific abscisic acid response element-binding factor, TaABF, and phosphorylates TaABF peptide sequences. Plant Physiology, 2002, 130(2): 837-846. doi: 10.1104/pp.001354 pmid: 12376648
[27]	ZHOU S Q, HUANG K R, ZHOU Y, HU Y Q, XIAO Y C, CHEN T, YIN M Q, LIU Y, XU M L, JIANG X C. Degradome sequencing reveals an integrative miRNA-mediated gene interaction network regulating rice seed vigor. BMC Plant Biology, 2022, 22(1): 269. doi: 10.1186/s12870-022-03645-2 pmid: 35650544
[28]	WU L J, TIAN Z D, ZHANG J H. Functional dissection of auxin response factors in regulating tomato leaf shape development. Frontiers in Plant Science, 2018, 9: 957. doi: 10.3389/fpls.2018.00957 pmid: 30022995
[29]	CHEN J, JIANG S L, YANG G B, LI L J, LI J, YANG F J. The MYB transcription factor SmMYB113 directly regulates ethylene-dependent flower abscission in eggplant. Plant Physiology and Biochemistry, 2024, 209: 108544.
[30]	ALCÂNTARA A, BOSCH J, NAZARI F, HOFFMANN G, GALLEI M, UHSE S, DARINO M A, OLUKAYODE T, REUMANN D, BAGGALEY L, et al. Systematic Y2H screening reveals extensive effector-complex formation. Frontiers in Plant Science, 2019, 10: 1437. doi: 10.3389/fpls.2019.01437 pmid: 31803201
[31]	CAI N, NONG X Q, LIU R, MCNEILL M R, WANG G J, ZHANG Z H, TU X B. The conserved cysteine-rich secretory protein MaCFEM85 interacts with MsWAK16 to activate plant defenses. International Journal of Molecular Sciences, 2023, 24(4): 4037.
[32]	SHAO W, SHI G F, CHU H, DU W J, ZHOU Z K, WURIYANGHAN H. Development of an NLR-ID toolkit and identification of novel disease-resistance genes in soybean. Plants, 2024, 13(5): 668.
[33]	QI Z Q, MENG X L, XU M, DU Y, YU J J, SONG T Q, PAN X Y, ZHANG R S, CAO H J, YU M N, et al. A novel Pik allele confers extended resistance to rice blast. Plant, Cell & Environment, 2024, 47(12): 4800-4814.
[34]	CARTER E W, PERAZA O G, WANG N. The protein interactome of the Citrus Huanglongbing pathogen Candidatus Liberibacter asiaticus. Nature Communications, 2023, 14: 7838.
[35]	DAGVADORJ B, OUTRAM M A, WILLIAMS S J, SOLOMON P S. The necrotrophic effector ToxA from Parastagonospora nodorum interacts with wheat NHL proteins to facilitate Tsn1-mediated necrosis. The Plant Journal, 2022, 110(2): 407-418.
[36]	XIAO H G, LORD E, SANFAÇON H. Proteolytic processing of plant proteins by potyvirus NIa proteases. Journal of Virology, 2022, 96(2): e0144421.
[37]	YANG C L, LIU Q Y, PENG M, CHEN X L, ZHU W L, CHEN X H, LI Q Y, ZENG D G, ZHAO Y Z. Penaeus stylirostris densovirus proteins CP and NS1 interact with peritrophin of Litopenaeus vannamei. Fish & Shellfish Immunology, 2020, 106: 357-364.
[38]	宾羽, 张琦, 王春庆, 赵晓春, 宋震, 周常勇. 利用酵母双杂交系统筛选与柑橘黄化脉明病毒CP互作的寄主因子. 中国农业科学, 2023, 56(10): 1881-1892. doi: 10.3864/j.issn.0578-1752.2023.10.006.
	BIN Y, ZHANG Q, WANG C Q, ZHAO X C, SONG Z, ZHOU C Y. Screening of the host factors interacting with CP of Citrus yellow vein clearing virus by yeast two-hybrid system. Scientia Agricultura Sinica, 2023, 56(10): 1881-1892. doi: 10.3864/j.issn.0578-1752.2023.10.006. (in Chinese)
[39]	SONG P W, ZHI H J, WU B Y, CUI X Y, CHEN X. Soybean Golgi SNARE 12 protein interacts with soybean mosaic virus encoded P3N-PIPO protein. Biochemical and Biophysical Research Communications, 2016, 478(4): 1503-1508. doi: 10.1016/j.bbrc.2016.08.103 pmid: 27553272
[40]	ZHANG H, CHENG G Y, YANG Z T, WANG T, XU J S. Identification of sugarcane host factors interacting with the 6K2 protein of the sugarcane mosaic virus. International Journal of Molecular Sciences, 2019, 20(16): 3867.
[41]	何勇, 范晓珠, 陈新月, 段书静, 胡婷婷, 谢如雪, 王宇晴, 陈静. 利用酵母双杂交系统筛选与辣椒轻斑驳病毒126 kDa蛋白互作的辣椒寄主因子. 中国农业科学, 2024, 57(15): 2986-2996. doi: 10.3864/j.issn.0578-1752.2024.15.006.
	HE Y, FAN X Z, CHEN X Y, DUAN S J, HU T T, XIE R X, WANG Y Q, CHEN J. Screening and verification of pepper host factors interacting with the 126 kDa protein of pepper mild mottle virus by yeast two-hybrid system. Scientia Agricultura Sinica, 2024, 57(15): 2986-2996. doi: 10.3864/j.issn.0578-1752.2024.15.006. (in Chinese)
[42]	冯雯蜜, 周芳雪, 于哲, 牟可欣, 井妍, 李海燕. 大豆抗花叶病毒病基因GmRHF1的克隆及功能分析. 中国农业科学, 2024, 57(23): 4632-4645. doi: 10.3864/j.issn.0578-1752.2024.23.005.
	FENG W M, ZHOU F X, YU Z, MOU K X, JING Y, LI H Y. Cloning and functional analysis of gm RHF1 gene against soybean mosaic virus. Scientia Agricultura Sinica, 2024, 57(23): 4632-4645. doi: 10.3864/j.issn.0578-1752.2024.23.005. (in Chinese)
[43]	ZHOU F X, FENG W M, MOU K X, YU Z, ZENG Y C, ZHANG W P, ZHOU Y G, LI Y X, GAO H T, XU K H, et al. Genome-wide analysis and expression profiling of soybean RbcS family in response to plant hormones and functional identification of GmRbcS 8 in soybean mosaic virus. International Journal of Molecular Sciences, 2024, 25(17): 9231.
[44]	HINNAH S C, HILL K, WAGNER R, SCHLICHER T, SOLL J. Reconstitution of a chloroplast protein import channel. EMBO Journal, 1997, 16(24): 7351-7360. doi: 10.1093/emboj/16.24.7351 pmid: 9405364
[45]	KIM J, NA Y J, PARK S J, BAEK S H, KIM D H. Biogenesis of chloroplast outer envelope membrane proteins. Plant Cell Reports, 2019, 38(7): 783-792. doi: 10.1007/s00299-019-02381-6 pmid: 30671649
[46]	REINBOTHE S, QUIGLEY F, SPRINGER A, SCHEMENEWITZ A, REINBOTHE C. The outer plastid envelope protein Oep16:Role as precursor translocase in import of protochlorophyllide oxidoreductase A. Proceedings of the National Academy of Sciences of the United States of America, 2004, 101(7): 2203-2208.
[47]	POLLMANN S, SPRINGER A, BUHR F, LAHROUSSI A, SAMOL I, BONNEVILLE J M, TICHTINSKY G, VON WETTSTEIN D, REINBOTHE C, REINBOTHE S. A plant Porphyria related to defects in plastid import of protochlorophyllide oxidoreductase A. Proceedings of the National Academy of Sciences of the United States of America, 2007, 104(6): 2019-2023.
[48]	CAI H R, ZHAO B K, LIANG K X, YUAN P G, ZHANG C Z, YANG S M, DUAN S J, JIN H L, WANG P, LIU B, et al. The Arabidopsis chloroplast protein HHL 1 regulates AvrRpt2-triggered immunity via light-dependent reactive oxygen species homeostasis. Journal of Integrative Plant Biology, 2025, 66(8): 2151-2166.
[49]	CHENG D J, XU X J, YAN Z Y, TETTEY C K, FANG L, YANG G L, GENG C, TIAN Y P, LI X D. The chloroplast ribosomal protein large subunit 1 interacts with viral polymerase and promotes virus infection. Plant Physiology, 2021, 187(1): 174-186.
[50]	PATEL R, HSU S C, BÉDARD J, INOUE K, JARVIS P. The Omp85-related chloroplast outer envelope protein OEP80 is essential for viability in Arabidopsis. Plant Physiology, 2008, 148(1): 235-245.
[51]	旷永洁, 柳浪, 严芳, 任斌, 闫大琦, 张大伟, 林宏辉, 周焕斌. 水稻与病原物互作中植物激素功能的研究进展. 生物技术通报, 2018, 34(2): 74-86. doi: 10.13560/j.cnki.biotech.bull.1985.2017-1104
	KUANG Y J, LIU L, YAN F, REN B, YAN D Q, ZHANG D W, LIN H H, ZHOU H B. Functions of phytohormones during the interactions between rice and pathogens. Biotechnology Bulletin, 2018, 34(2): 74-86. (in Chinese)
[52]	BLÁZQUEZ M A, NELSON D C, WEIJERS D. Evolution of plant hormone response pathways. Annual Review of Plant Biology, 2020, 71: 327-353. doi: 10.1146/annurev-arplant-050718-100309 pmid: 32017604
[53]	SAMOL I, BUHR F, SPRINGER A, POLLMANN S, LAHROUSSI A, ROSSIG C, VON WETTSTEIN D, REINBOTHE C, REINBOTHE S. Implication of the oep16-1 mutation in a flu-independent, singlet oxygen-regulated cell death pathway in Arabidopsis thaliana. Plant and Cell Physiology, 2011, 52(1): 84-95.
[54]	PUDELSKI B, SCHOCK A, HOTH S, RADCHUK R, WEBER H, HOFMANN J, SONNEWALD U, SOLL J, PHILIPPAR K. The plastid outer envelope protein OEP16 affects metabolic fluxes during ABA-controlled seed development and germinationOpen Access. Journal of Experimental Botany, 2012, 63(5): 1919-1936. doi: 10.1093/jxb/err375 pmid: 22155670
[55]	DREA S C, LAO N T, WOLFE K H, KAVANAGH T A. Gene duplication, exon gain and neofunctionalization of OEP16-related genes in land plants. The Plant Journal, 2006, 46(5): 723-735.
[56]	NIELSEN E, AKITA M, DAVILA-APONTE J, KEEGSTRA K. Stable association of chloroplastic precursors with protein translocation complexes that contain proteins from both envelope membranes and a stromal Hsp100 molecular chaperone. EMBO Journal, 1997, 16(5): 935-946. doi: 10.1093/emboj/16.5.935 pmid: 9118955
[57]	DUY D, SOLL J, PHILIPPAR K. Solute channels of the outer membrane: From bacteria to chloroplasts. Biological Chemistry, 2007, 388(9): 879-889. doi: 10.1515/BC.2007.120 pmid: 17696771
[58]	SEEDORF M, SOLL J. Copper chloride, an inhibitor of protein import into chloroplasts. FEBS Letters, 1995, 367(1): 19-22. doi: 10.1016/0014-5793(95)00529-i pmid: 7601278
[59]	GÜNSEL U, KLÖPFER K, HÄUSLER E, HITZENBERGER M, BÖLTER B, SPERL L E, ZACHARIAS M, SOLL J, HAGN F. Structural basis of metabolite transport by the chloroplast outer envelope channel OEP21. Nature Structural & Molecular Biology, 2023, 30(6): 761-769.
[60]	POHLMEYER K, SOLL J, GRIMM R, HILL K, WAGNER R. A high-conductance solute channel in the chloroplastic outer envelope from pea. The Plant Cell, 1998, 10(7): 1207-1216.
[61]	SAMOL I, ROSSIG C, BUHR F, SPRINGER A, POLLMANN S, LAHROUSSI A, VON WETTSTEIN D, REINBOTHE C, REINBOTHE S. The outer chloroplast envelope protein OEP16-1 for plastid import of NADPH: Protochlorophyllide oxidoreductase a in Arabidopsis thaliana. Plant and Cell Physiology, 2011, 52(1): 96-111.
[62]	POHLMEYER K, SOLL J, STEINKAMP T, HINNAH S, WAGNER R. Isolation and characterization of an amino acid-selective channel protein present in the chloroplastic outer envelope membrane. Proceedings of the National Academy of Sciences of the United States of America, 1997, 94(17): 9504-9509.

编号 Number	基因号 Gene identifier	克隆数量 No. of clones	基因描述 Gene description
1	Glyma.06G080100	1	肌动蛋白纤维包被蛋白——原肌球蛋白 Actin filament-coating protein tropomyosin
2	Glyma.02G247800	1	富丝氨酸/精氨酸剪接因子 Serine/arginine rich splicing factor
3	Glyma.11G072700	2	含FG-GAP重复蛋白 FG-GAP repeat-containing protein
4	Glyma.13G219400	3	叶绿体类蛋白酶5 Protease do-like 5, chloroplastic
5	Glyma.02G184800	5	外膜孔蛋白16-1，叶绿体 Outer envelope pore protein 16-1, chloroplastic
6	Glyma.05G225000	1	线粒体加工肽酶/加工增强肽酶 Mitochondrial processing peptidase/Processing enhancing peptidase
7	Glyma.13G162800	1	无机二磷酸酶/焦磷酸磷酸水解酶 Inorganic diphosphatase/Pyrophosphate phosphohydrolase
8	Glyma.07G034800	2	亚油酸13S-脂氧合酶/亚油酸9S-脂氧合酶/亚油酸9-脂氧合酶 Linoleate 13S-lipoxygenase/lipoxidase//linoleate 9s-lipoxygenase/linoleate 9-lipoxygenase
9	Glyma.06G118000	2	蓝藻和叶绿体NDH-1亚基M（NdhM） Cyanobacterial and plastid NDH-1 subunit M (NdhM)
10	Glyma.02G002000	1	甲基-6-植烷基-1,4-羟醌甲基转移酶，叶绿体 Methyl-6-phytyl-1,4-hydroquinone methyltransferase, chloroplastic
11	Glyma.09G010700	1	溶质运载家族成员13 Solute carrier family 13 member
12	Glyma.04G166700	1	肽脯氨酰顺反异构酶 Peptidyl-prolyl cis-trans isomerase

编号 Number	基因号 Gene identifier	克隆数量 No. of clones	基因描述 Gene description
1	Glyma.17G159000	1	生长因子蛋白相关 Growth factor protein-related
2	Glyma.15G275600	1	光系统Ⅱ5 kDa蛋白，叶绿体 PhotosystemⅡ 5 kDa protein, chloroplastic
3	Glyma.11G102300	2	含FG-GAP重复蛋白 FG-GAP repeat-containing protein
4	Glyma.18G002800	1	硒结合蛋白 Selenium-binding protein
5	Glyma.06G172000	1	肽酶Ⅰ锌金属蛋白酶（M18） AminopeptidaseⅠ zinc metalloprotease (M18)
6	Glyma.05G135300	1	液泡磷酸合成酶亚基f Vacuolar ATP synthase subunit f
7	Glyma.13G352100	2	60S核糖体蛋白L28 60S ribosomal protein L28
8	Glyma.05G216000	3	含锌指CCCH结构域的蛋白18 Zinc finger CCCH domain-containing protein 18
9	Glyma.11G140500	1	RNA聚合酶Ⅱ转录亚基37e相关介质 Mediator of rna polymeraseⅡ transcription subunit 37e-related
10	Glyma.09G008100	1	叶绿体相关质体脂质相关蛋白3 Plastid-lipid-associated protein 3, chloroplastic- related
11	Glyma.02G184800	4	外膜孔蛋白16-1，叶绿体 Outer envelope pore protein 16-1, chloroplastic
12	Glyma.15G099800	3	含组氨酸磷酸转移蛋白1 Histidine-containing phosphotransfer protein1