Abstract
Main conclusion
Overexpression of miR172a and b in tomato ( Solanum lycopersicum ) Zaofen No. 2 increased resistance to Phytophthora infestans infection by suppressing of an AP2/ERF transcription factor.
The miR172 family has been shown to participate in the growth phase transition, flowering time control, abiotic and biotic stresses by regulating the expression of a small group of AP2/ERF transcription factors. In this study, the precursors of miR172a and b were cloned from tomato, Solanum pimpinellifolium L3708. We used the degradome sequencing to determine the cleavage site of miR172 to a member of the AP2/ERF transcription factor family (Solyc11g072600.1.1). qRT-PCR results showed that the expression of AP2/ERF was negatively correlated with the expression of miR172 in S. pimpinellifolium L3708 infected with Phytophthora infestans. Overexpression of miR172a and b in S. lycopersicum Zaofen No. 2 conferred greater resistance to P. infestans infection, as evidenced by decreased disease index, lesion sizes, and P. infestans abundance. The SOD and POD play important roles in scavenging late massive ROS in plant–pathogen interaction. Malonaldehyde (MDA) is widely recognized as an indicator of lipid peroxidation. Membrane damage in plants can be estimated by measuring leakage of electrolytes, which is evaluated by determining relative electrolyte leakage (REL). Less H2O2 and O2 −, higher activities of POD and SOD, less MDA content and REL, and higher chlorophyll content and photosynthetic rate were also shown in transgenic plants after inoculation with P. infestans. Our results constitute the first step towards further investigations into the biological function and molecular mechanism of miR172-mediated silencing of AP2/ERF transcription factors in S. lycopersicum–P. infestans interaction and provide a candidate gene for breeding to enhance biotic stress-resistance in S. lycopersicum.
Similar content being viewed by others
Abbreviations
- dpi:
-
Days post inoculation
- MDA:
-
Malonaldehyde
- POD:
-
Peroxidase
- REL:
-
Relative electrolyte leakage
- ROS:
-
Reactive oxygen species
- SOD:
-
Superoxide dismutase
References
Addo-Quaye C, Eshoo TW, Bartel DP, Axtell MJ (2008) Endogenous siRNA and miRNA targets identified by sequencing of the Arabidopsis degradome. Curr Biol 18:758–762
Addo-Quaye C, Miller W, Axtell MJ (2009) CleaveLand: a pipeline for using degradome data to find cleaved small RNA targets. Bioinformatics 25:130–131
Alabi OJ, Zheng Y, Jagadeeswaran G, Sunkar R, Naidu RA (2012) High-throughput sequence analysis of small RNAs in grapevine (Vitis vinifera L.) affected by grapevine leafroll disease. Mol Plant Pathol 13:1060–1076
Anderson JP, Lichtenzveig J, Gleason C, Oliver RP, Singh KB (2010) The B-3 ethylene response factor MtERF1-1 mediates resistance to a subset of root pathogens in Medicago truncatula without adversely affecting symbiosis with rhizobia. Plant Physiol 154:861–873
Aukerman MJ, Sakai H (2003) Regulation of flowering time and floral organ identity by a microRNA and its APETALA2-like target genes. Plant Cell 15:2730–2741
Berrocal-Lobo M, Molina A (2004) Ethylene response factor 1 mediates Arabidopsis resistance to the soilborne fungus Fusarium oxysporum. Mol Plant Microbe Interact 17:763–770
Berrocal-Lobo M, Molina A, Solano R (2002) Constitutive expression of ETHYLENE-RESPONSE-FACTOR1 in Arabidopsis confers resistance to several necrotrophic fungi. Plant J 29:23–32
Black LL, Wang TC, Hanson PM, Chen JT (1996) Late blight resistance in four wild tomato accessions: effectiveness in diverse locations and inheritance of resistance. Phytopathology 86:S24
Bonde R, Murphy EF (1952) Resistance of certain tomato varieties and crosses to late blight. Maine Agric Exp Stn Bull 497:5–15
Cao Y, Wu Y, Zheng Z, Song F (2006) Overexpression of the rice EREBP-like gene OsBIERF3 enhances disease resistance and salt tolerance in transgenic tobacco. Physiol Mol Plant Pathol 67:202–211
Chen X (2004) A microRNA as a translational repressor of APETALA2 in Arabidopsis flower development. Science 303:2022–2025
Chen L, Zhang Z, Liang H, Liu H, Du L, Xu H, Xin Z (2008) Overexpression of TiERF1 enhances resistance to sharp eyespot in transgenic wheat. J Exp Bot 59:4195–4204
Chen HY, Ling JG, Wu FH, Zhang LJ, Sun ZD, Yang HQ (2013) Effect of hypobaric storage on flesh lignification, active oxygen metabolism and related enzyme activities in bamboo shoots. LWT Food Sci Technol 51:190–195
Chen J, Zheng Y, Qin L, Wang Y, Chen L, He Y, Fei Z, Lu G (2016) Identification of miRNAs and their targets through high-throughput sequencing and degradome analysis in male and female Asparagus officinalis. BMC Plant Biol 16:80
de Vries S, Kloesges T, Rose LE (2015) Evolutionarily dynamic, but robust, targeting of resistance genes by the miR482/2118 gene family in the Solanaceae. Genome Biol Evol 7:3307–3321
de Vries S, von Dahlen JK, Uhlmann C, Schnake A, Kloesges T, Rose LE (2017) Signatures of selection and host-adapted gene expression of the Phytophthora infestans RNA silencing suppressor PSR2. Mol Plant Pathol 18:110–124
Ebrahimi Khaksefidi R, Mirlohi S, Khalaji F, Fakhari Z, Shiran B, Fallahi H, Rafiei F, Budak H, Ebrahimie E (2015) Differential expression of seven conserved microRNAs in response to abiotic stress and their regulatory network in Helianthus annuus. Front Plant Sci 6:741
Fei Q, Li P, Teng C, Meyers BC (2015) Secondary siRNAs from Medicago NB-LRRs modulated via miRNA-target interactions and their abundances. Plant J 83:451–465
Foolad MR, Merk HL, Ashrafi H (2008) Genetics, genomics and breeding of late blight and early blight resistance in tomato. Crit Rev Plant Sci 27:75–107
Fry WE, Goodwin SB, Matuszak JM, Spoelman LJ, Milgroom MG, Drenth A (1992) Population genetics and intercontinental migration of Phytophthora infestans. Annu Rev Phytopathol 30:107–129
Gai YP, Li YQ, Guo FY, Yuan CZ, Mo YY, Zhang HL, Wang H, Ji XL (2014) Analysis of phytoplasma-responsive sRNAs provide insight into the pathogenic mechanisms of mulberry yellow dwarf disease. Sci Rep 4:5378
Gu YQ, Wildermuth MC, Chakravarthy S, Loh YT, Yang C, He X, Han Y, Martin GB (2002) Tomato transcription factors Pti4, Pti5, and Pti6 activate defense responses when expressed in Arabidopsis. Plant Cell 14:817–831
Gupta OP, Meena NL, Sharma I, Sharma P (2014) Differential regulation of microRNAs in response to osmotic, salt and cold stresses in wheat. Mol Biol Rep 41:4623–4629
Huang X, Wang A, Xu X, Li J, Li N (2010) Construction of genetic linkage map and QTL analysis of Phytophthora infestans resistant gene Ph-2 in tomato. Acta Hortic Sin 37:1085–1092
Huang XS, Luo T, Fu XZ, Fan QJ, Liu JH (2011) Cloning and molecular characterization of a mitogen-activated protein kinase gene from Poncirus trifoliata whose ectopic expression confers dehydration/drought tolerance in transgenic tobacco. J Exp Bot 62:5191–5206
Janga SC, Vallabhaneni S (2011) MicroRNAs as post-transcriptional machines and their interplay with cellular networks. Adv Exp Med Biol 722:59–74
Jiang Y, Guo L, Liu R, Jiao B, Zhao X, Ling Z, Luo K (2016) Overexpression of poplar PtrWRKY89 in transgenic Arabidopsis leads to a reduction of disease resistance by regulating defense-related genes in salicylate-jasmonate-dependent signaling. PLoS One 11:e0149137
Jin H (2008) Endogenous small RNAs and antibacterial immunity in plants. FEBS Lett 582:2679–2684
Jung JH, Seo YH, Seo PJ, Reyes JL, Yun J, Chua NH, Park CM (2007) The GIGANTEA-regulated microRNA172 mediates photoperiodic flowering independent of CONSTANS in Arabidopsis. Plant Cell 19:2736–2748
Kim MJ, Mutschler MA (2005) Transfer to processing tomato and characterization of late blight resistance derived from Solanum pimpinellifolium L. L3708. J Am Soc Hortic Sci 130:877–884
Kotchoni SO, Gachomo EW (2006) The reactive oxygen species network pathways: an essential prerequisite for perception of pathogen attack and the acquired disease resistance in plants. J Biosci 31:389–404
Lauter N, Kampani A, Carlson S, Goebel M, Moose SP (2005) microRNA172 down-regulates glossy15 to promote vegetative phase change in maize. Proc Natl Acad Sci USA 102:9412–9417
Lee B, Lee H, Xiong L, Zhu JK (2002) A mitochondrial complex I defect impairs cold-regulated nuclear gene expression. Plant Cell 14:1235–1251
Lee YS, Lee DY, Cho LH, An G (2014) Rice miR172 induces flowering by suppressing OsIDS1 and SNB, two AP2 genes that negatively regulate expression of Ehd1 and florigens. Rice (N Y) 7:31
Li F, Pignatta D, Bendix C, Brunkard JO, Cohn MM, Tung J, Sun H, Kumar P, Baker B (2012) MicroRNA regulation of plant innate immune receptors. Proc Natl Acad Sci USA 109:1790–1795
Li J, Luan Y, Zhai J, Liu P, Xia X (2013) Bioinformatic analysis of functional characteristics of miR172 family in tomato. J Northeast Agric Univ (English Edition) 20:19–27
Li Y, Lu YG, Shi Y, Wu L, Xu YJ, Huang F, Guo XY, Zhang Y, Fan J, Zhao JQ, Zhang HY, Xu PZ, Zhou JM, Wu XJ, Wang PR, Wang WM (2014) Multiple rice microRNAs are involved in immunity against the blast fungus Magnaporthe oryzae. Plant Physiol 164:1077–1092
Li J, Luan Y, Liu Z (2015a) SpWRKY1 mediates resistance to Phytophthora infestans and tolerance to salt and drought stress by modulating reactive oxygen species homeostasis and expression of defense-related genes in tomato. Plant Cell Tissue Organ Cult 123:67–81
Li J, Luan Y, Liu Z (2015b) Overexpression of SpWRKY1 promotes resistance to Phytophthora nicotianae and tolerance to salt and drought stress in transgenic tobacco. Physiol Plant 155:248–266
Li W, Wang T, Zhang Y, Li Y (2016) Overexpression of soybean miR172c confers tolerance to water deficit and salt stress, but increases ABA sensitivity in transgenic Arabidopsis thaliana. J Exp Bot 67:175–194
Liu D, Chen X, Liu J, Ye J, Guo Z (2012) The rice ERF transcription factor OsERF922 negatively regulates resistance to Magnaporthe oryzae and salt tolerance. J Exp Bot 63:3899–3911
Luan Y, Wang W, Liu P (2014) Identification and functional analysis of novel and conserved microRNAs in tomato. Mol Biol Rep 41:5385–5394
Luan Y, Cui J, Zhai J, Li J, Han L, Meng J (2015) High-throughput sequencing reveals differential expression of miRNAs in tomato inoculated with Phytophthora infestans. Planta 241:1405–1416
McGrath KC, Dombrecht B, Manners JM, Schenk PM, Edgar CI, Maclean DJ, Scheible WR, Udvardi MK, Kazan K (2005) Repressor- and activator-type ethylene response factors functioning in jasmonate signaling and disease resistance identified via a genome-wide screen of Arabidopsis transcription factor gene expression. Plant Physiol 139:949–959
Merk HL, Ashrafi H, Foolad MR (2012) Selective genotyping to identify late blight resistance genes in an accession of the tomato wild species Solanum pimpinellifolium. Euphytica 187:63–75
Miranda BEC, Suassuna ND, Reis A (2010) Mating type, mefenoxam sensitivity, and pathotype diversity in Phytophthora infestans isolates from tomato in Brazil. Pesq Agropec Bras 45:671–679
Mo J, Li D, Zhang H, Song F (2011) Roles of ERF transcription factors in biotic and abiotic stress response in plants. Plant Physiol J 47:1145–1154
Nair SK, Wang N, Turuspekov Y, Pourkheirandish M, Sinsuwongwat S, Chen G, Sameri M, Tagiri A, Honda I, Watanabe Y, Kanamori H, Wicker T, Stein N, Nagamura Y, Matsumoto T, Komatsuda T (2010) Cleistogamous flowering in barley arises from the suppression of microRNA-guided HvAP2 mRNA cleavage. Proc Natl Acad Sci USA 107:490–495
Naqvi AR, Haq QM, Mukherjee SK (2010) MicroRNA profiling of tomato leaf curl new delhi virus (tolcndv) infected tomato leaves indicates that deregulation of mir159/319 and mir172 might be linked with leaf curl disease. Virol J 7:281
Navarro L, Dunoyer P, Jay F, Arnold B, Dharmasiri N, Estelle M, Voinnet O, Jones JDG (2006) A plant miRNA contributes to antibacterial resistance by repressing auxin signaling. Science 312:436–439
Ouyang S, Park G, Atamian HS, Han CS, Stajich JE, Kaloshian I, Borkovich KA (2014) MicroRNAs suppress NB domain genes in tomato that confer resistance to Fusarium oxysporum. PLoS Pathog 10:e1004464
Pan WJ, Tao JJ, Cheng T, Bian XH, Wei W, Zhang WK, Ma B, Chen SY, Zhang JS (2016) Soybean miR172a improves salt tolerance and can function as a long-distance signal. Mol Plant 9:1337–1340
Park W, Li J, Song R, Messing J, Chen X (2002) CARPEL FACTORY, a dicer homolog, and HEN1, a novel protein, act in microRNA metabolism in Arabidopsis thaliana. Curr Biol 12:1484–1495
Park Y, Hwang J, Kim K, Kang J, Kim B (2013) Development of the gene-based SCARs for the Ph-3 locus, which confers late blight resistance in tomato. Sci Hortic 164:9–16
Pré M, Atallah M, Champion A, De Vos M, Pieterse CM, Memelink J (2008) The AP2/ERF domain transcription factor ORA59 integrates jasmonic acid and ethylene signals in plant defense. Plant Physiol 147:1347–1357
Qiao Y, Liu L, Xiong Q, Flores C, Wong J, Shi J, Wang X, Liu X, Xiang Q, Jiang S, Zhang F, Wang Y, Judelson HS, Chen X, Ma W (2013) Oomycete pathogens encode RNA silencing suppressors. Nat Genet 45:330–333
Qiao Y, Shi J, Zhai Y, Hou Y, Ma W (2015) Phytophthora effector targets a novel component of small RNA pathway in plants to promote infection. Proc Natl Acad Sci USA 112:5850–5855
Sewelam N, Kazan K, Thomas-Hall SR, Kidd BN, Manners JM, Schenk PM (2013) Ethylene response factor 6 is a regulator of reactive oxygen species signaling in Arabidopsis. PLoS One 8:e70289
Shi WN, Hao LL, Li J, Liu DD, Guo XQ, Li H (2014) The Gossypium hirsutum WRKY gene GhWRKY39-1 promotes pathogen infection defense responses and mediates salt stress tolerance in transgenic Nicotiana benthamiana. Plant Cell Rep 33:483–498
Shivaprasad PV, Chen HM, Patel K, Bond DM, Santos BA, Baulcombe DC (2012) A microRNA superfamily regulates nucleotide binding site-leucine-rich repeats and other mRNAs. Plant Cell 24:859–874
Spanudakis E, Jackson S (2014) The role of microRNAs in the control of flowering time. J Exp Bot 65:365–380
Steffen P, Voss B, Rehmsmeier M, Reeder J, Giegerich R (2006) RNAshapes: an integrated RNA analysis package based on abstract shapes. Bioinformatics 22:500–503
Sun G, Luan Y, Cui J (2014) Mining and characterization of miRNAs closely associated with the pathogenicity in tomato. Yichuan 36:69–76
Tian Y, Zhang H, Pan X, Chen X, Zhang Z, Lu X, Huang R (2011) Overexpression of ethylene response factor TERF2 confers cold tolerance in rice seedlings. Transgenic Res 20:857–866
Varkonyi-Gasic E, Wu R, Wood M, Walton EF, Hellens RP (2007) Protocol: a highly sensitive RT-PCR method for detection and quantification of microRNAs. Plant Methods 3:12
Wi SJ, Ji NR, Park KY (2012) Synergistic biosynthesis of biphasic ethylene and reactive oxygen species in response to hemibiotrophic Phytophthora parasitica in tobacco plants. Plant Physiol 159:251–265
Wong J, Gao L, Yang Y, Zhai J, Arikit S, Yu Y, Duan S, Chan V, Xiong Q, Yan J, Li S, Liu R, Wang Y, Tang G, Meyers BC, Chen X, Ma W (2014) Roles of small RNAs in soybean defense against Phytophthora sojae infection. Plant J 79:928–940
Wu L, Zhang Z, Zhang H, Wang XC, Huang R (2008) Transcriptional modulation of ethylene response factor protein JERF3 in the oxidative stress response enhances tolerance of tobacco seedlings to salt, drought, and freezing. Plant Physiol 148:1953–1963
Würschum T, Gross-Hardt R, Laux T (2006) APETALA2 regulates the stem cell niche in the Arabidopsis shoot meristem. Plant Cell 18:295–307
Xiong Q, Ye W, Choi D, Wong J, Qiao Y, Tao K, Wang Y, Ma W (2014) Phytophthora suppressor of RNA silencing 2 is a conserved RxLR effector that promotes infection in soybean and Arabidopsis thaliana. Mol Plant Microbe Interact 27:1379–1389
Yan J, Zhang H, Zheng Y, Ding Y (2014) Comparative expression profiling of miRNAs between the cytoplasmic male sterile line Meixiang A and its maintainer line Meixiang B during rice anther development. Planta 241:109–123
Yang L, Huang H (2014) Roles of small RNAs in plant disease resistance. J Integr Plant Biol 56:962–970
Yang L, Mu X, Liu C, Cai J, Shi K, Zhu W, Yang Q (2015) Overexpression of potato miR482e enhanced plant sensitivity to Verticillium dahliae infection. J Integr Plant Biol 57:1078–1088
Ye W, Ma W (2016) Filamentous pathogen effectors interfering with small RNA silencing in plant hosts. Curr Opin Microbiol 32:1–6
Yoshikawa T, Ozawa S, Sentoku N, Itoh J, Nagato Y, Yokoi S (2013) Change of shoot architecture during juvenile-to-adult phase transition in soybean. Planta 238:229–237
Zhang J, Zeng R, Chen J, Liu X, Liao Q (2008) Identification of conserved microRNAs and their targets from Solanum lycopersicum Mill. Gene 423:1–7
Zhang C, Liu L, Wang X, Vossen J, Li G, Li T, Zheng Z, Gao J, Guo Y, Visser RGF, Li J, Bai Y, Du Y (2014) The Ph-3 gene from Solanum pimpinellifolium encodes CC-NBS-LRR protein conferring resistance to Phytophthora infestans. Theor Appl Genet 127:1353–1364
Zhang H, Huang L, Dai Y, Liu S, Hong Y, Tian L, Huang L, Cao Z, Li D, Song F (2015) Arabidopsis AtERF15 positively regulates immunity against Pseudomonas syringae pv. tomato DC3000 and Botrytis cinerea. Front. Plant Sci 6:686
Zhou J, Zhang H, Yang Y, Zhang Z, Zhang H, Hu X, Chen J, Wang XC, Huang R (2008) Abscisic acid regulates TSRF1-mediated resistance to Ralstonia solanacearum by modifying the expression of GCC box-containing genes in tobacco. J Exp Bot 59:645–652
Zhou L, Liu Y, Liu Z, Kong D, Duan M, Luo L (2010) Genome wide identification and analysis of drought-responsive microRNAs in Oryza sativa. J Exp Bot 61:4157–4168
Acknowledgements
This work is supported by Grants from the National Natural Science Foundation of China (Nos. 31471880 and 61472061).
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Conflict of interest
The authors declare that they have no conflict of interest.
Electronic supplementary material
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Luan, Y., Cui, J., Li, J. et al. Effective enhancement of resistance to Phytophthora infestans by overexpression of miR172a and b in Solanum lycopersicum . Planta 247, 127–138 (2018). https://doi.org/10.1007/s00425-017-2773-x
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00425-017-2773-x