Skip to main content
SpringerLink
Log in

Identification of tomato miRNAs responsive to root colonization by endophytic Pochonia chlamydosporia

  • Genomics, transcriptomics, proteomics
  • Published:
Applied Microbiology and Biotechnology Aims and scope Submit manuscript

Abstract

The molecular mechanisms active during the endophytic phase of the fungus Pochonia chlamydosporia are still poorly understood. In particular, few data are available on the links between the endophyte and the root response, as modulated by noncoding small RNAs. In this study, we describe the microRNAs (miRNAs) that are differentially expressed (DE) in the roots of tomato, colonized by P. chlamydosporia. A genome-wide NGS expression profiling of small RNAs in roots, either colonized or not by the fungus, showed 26 miRNAs upregulated in inoculated roots. Their predicted target genes are involved in the plant information processing system, which recognizes, percepts, and transmits signals, with higher representations in processes such as apoptosis and plant defense regulation. RNAseq data showed that predicted miRNA target genes were downregulated in tomato roots after 4, 7, 10, and 21 days post P. chlamydosporia inoculation. The differential expression of four miRNAs was further validated using qPCR analysis. The P. chlamydosporia endophytic lifestyle in tomato roots included an intricate network of miRNAs and targets. Data provide a first platform of DE tomato miRNAs after P. chlamydosporia colonization. They indicated that several miRNAs are involved in the host response to the fungus, playing important roles for its recognition as a symbiotic microorganism, allowing endophytism by modulating the host defense reaction. Data also indicated that endophytism affects tRNA fragmentation. This is the first study on miRNAs induced by P. chlamydosporia endophytism and related development regulation effects in Solanum lycopersicum.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

References

  • Antolin-Llovera M, Ried MK, Binder A, Parniske M (2012) Receptor kinase signaling pathways in plant-microbe interactions. Annu Rev Phytopathol 50:451–473

    Article  CAS  PubMed  Google Scholar 

  • Asman A, Vetukuri RR, Jahan SN, Fogelqvist J, Corcoran P, Avrova AO, Whisson SC, Dixelius C (2014) Fragmentation of tRNA in Phytophthora infestans asexual life cycle stages and during host plant infection. BMC Microbiol 14:308

    Article  PubMed  PubMed Central  Google Scholar 

  • Barturen G, Rueda A, Hamberg M, Alganza A, Lebron R, Kotsyfakis M, Shi B, Koppers-Lalic D, Hackenberg M (2014) sRNAbench: profiling of small RNAs and its sequence variants in single or multi-species high-throughput experiments. Methods Next Gener Seq 1:21–31

    Google Scholar 

  • Bordallo JJ, Lopez-Llorca LV, Jansson HB, Salinas J, Persmark L, Asensio L (2002) Colonization of plant roots by egg-parasitic and nematode-trapping fungi. New Phytol 154:491–499

    Article  Google Scholar 

  • Branscheid A, Sieh D, Pant BD, May P, Devers EA, Elkrog A, Schauser L, Scheible WR, Krajinski F (2010) Expression pattern suggests a role of MiR399 in the regulation of the cellular response to local Pi increase during arbuscular mycorrhizal symbiosis. Mol Plant-Microbe Interact 23:915–926

    Article  CAS  PubMed  Google Scholar 

  • Chen C, Ridzon DA, Broomer AJ, Zhou Z, Lee DH, Nguyen JT, Barbisin M, Xu NL, Mahuvakar VR, Andersen MR, Lao KQ, Livak KJ, Guegler KJ (2005) Real-time quantification of microRNAs by stem–loop RT–PCR. Nucleic Acids Res 33:e179

    Article  PubMed  PubMed Central  Google Scholar 

  • Combier JP, Frugier F, de Billy F, Boualem A, El-Yahyaoui F (2006) MtHAP2–1 is a key transcriptional regulator of symbiotic nodule development regulated by microRNA169 in Medicago truncatula. Genes Dev 20:3084–3088

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Dai X, Zhao PX (2011) psRNATarget: a plant small RNA target analysis server. Nucleic Acids Res 39:W155–W159

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Devers EA, Branscheid A, May P, Krajinski F (2011) Stars and symbiosis: microRNA- and microRNA*-mediated transcript cleavage involved in arbuscular mycorrhizal symbiosis. Plant Physiol 156:1990–2010

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Du Z, Zhou X, Ling Y, Zhang Z, Su Z (2010) agriGO: a GO analysis toolkit for the agricultural community. Nucleic Acids Res 38:W64–W70

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Escudero N, Lopez-Llorca LV (2012) Effects on plant growth and root-knot nematode infection of an endophytic GFP transformant of the nematophagous fungus Pochonia chlamydosporia. Symbiosys 57:33–42

    Article  Google Scholar 

  • Fei Q, Xia R, Meyers BC (2013) Phased, secondary, small interfering RNAs in post-transcriptional regulatory networks. Plant Cell 25:2400–2415. https://doi.org/10.1105/tpc.113.114652

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Gebetsberger J, Polacek N (2013) Slicing tRNAs to boost functional ncRNA diversity. RNA Biol 10:1798–1806

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Gu M, Liu W, Meng Q, Zhang W, Chen A, Sun S (2014) Identification of microRNAs in six solanaceous plants and their potential link with phosphate and mycorrhizal signaling. J Integr Plant Biol 56:1164–1178. https://doi.org/10.1111/jipb.12233

    Article  CAS  PubMed  Google Scholar 

  • Gu M, Xu K, Chen A, Zhu Y, Tang G, Xu G (2010) Expression analysis suggests potential roles of microRNAs for phosphate and arbuscular mycorrhizal signaling in Solanum lycopersicum. Physiol Plant 138:226–237. https://doi.org/10.1111/j.1399-3054.2009.01320.x

    Article  CAS  PubMed  Google Scholar 

  • Hackenberg M, Rodriguez-Ezpeleta N, Aransay AM (2011) miRanalyzer: an update on the detection and analysis of microRNAs in high-throughput sequencing experiments. Nucleic Acids Res 39:132–138

    Article  Google Scholar 

  • Hanlon MT, Coenen C (2011) Genetic evidence for auxin involvement in arbuscular mycorrhizal initiation. New Phytol 189:701–709

    Article  PubMed  Google Scholar 

  • Hewezi T, Baum TJ (2012) Complex feedback regulations govern the expression of miRNA396 and its GRF target genes. Plant Signal Behav 7:749–751

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Hewezi T, Howe P, Maier TR, Baum TJ (2008) Arabidopsis small RNAs and their targets during cyst nematode parasitism. MPMI 21:1622–1634

    Article  CAS  PubMed  Google Scholar 

  • Hewezi T, Maier T, Nettleton D, Baum TJ (2012) Arabidopsis microRNA396-GRF1/GRF3 regulatory module acts as a developmental regulator in the reprogramming of root cells during cyst nematode infection. Plant Physiol 159:321–335

  • Holt DB, Gupta V, Meyer D, Abel NB, Andersen SU, Stougaard J (2015) Micro RNA 172 (miR172) signals epidermal infection and is expressed in cells primed for bacterial invasion in Lotus japonicus roots and nodules. New Phytol 208:241–256. https://doi.org/10.1111/nph.13445

    Article  CAS  PubMed  Google Scholar 

  • Jin W, Wu F (2015) Characterization of miRNAs associated with Botrytis cinerea infection of tomato leaves. BMC Plant Biol 15:1

    Article  PubMed  PubMed Central  Google Scholar 

  • Kal AJ, van Zonneveld AJ, Benes V, van den Berg M, Koerkamp MG, Albermann K, Strack N, Ruijter JM, Richter A, Dujon B, Ansorge W, Tabak HF (1999) Dynamics of gene expression revealed by comparison of serial analysis of gene expression transcript profiles from yeast grown on two different carbon sources. Mol Biol Cell 10:1859–1872

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kozomara A, Griffiths-Jones S (2014) miRBase: annotating high confidence microRNAs using deep sequencing data. Nucleic Acids Res 42(D1):D68–D73. https://doi.org/10.1093/nar/gkt1181

    Article  CAS  PubMed  Google Scholar 

  • Kozyrovska NO (2013) Crosstalk between endophytes and a plant host within information processing networks. Biopolym Cell 29:234–243

    Article  CAS  Google Scholar 

  • Larriba E, Jaime MD, Carbonell-Caballero J, Conesa A, Dopazo J, Nislow C, Martín-Nieto J, Lopez-Llorca LV (2014) Sequencing and functional analysis of the genome of a nematode egg-parasitic fungus, Pochonia chlamydosporia. Fungal Genet Biol 65:69–80

    Article  CAS  PubMed  Google Scholar 

  • Larriba E, Jaime MDLA, Nislow C, Martín-Nieto J, Lopez-Llorca LV (2015) Endophytic colonization of barley (Hordeum vulgare) roots by the nematophagous fungus Pochonia chlamydosporia reveals plant growth promotion and a general defense and stress transcriptomic response. J Plant Res 128(4):665–678

  • Lelandais-Briere C, Naya L, Sallet E, Calenge F, Frugier F (2009) Genome-wide Medicago truncatula small RNA analysis revealed novel microRNAs and isoforms differentially regulated in roots and nodules. Plant Cell 21:2780–2796

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • 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 U S A 109:1790–1795

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Liu S, Li JH, Wu J, Zhou KR, Zhou H, Yang JH, Qu LH (2015) StarScan: a web server for scanning small RNA targets from degradome sequencing data. Nucleic Acids Res 43:W480–W486

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2−CT method. Methods 25:402–408

    Article  CAS  PubMed  Google Scholar 

  • Maciá-Vicente J, Rosso LC, Ciancio A, Jansson H-B, Lopez-Llorca LV (2009) Colonisation of barley roots by endophytic Fusarium equiseti and Pochonia chlamydosporia: effects on plant growth and disease. Ann Appl Biol 155:391–401

    Article  Google Scholar 

  • Manzanilla-Lopez RH, Esteves I, Finetti-Sialer MM, Hirsch PR, Ward E, Devonshire J, Hidalgo-Diaz L (2013) Pochonia chlamydosporia: advances and challenges to improve its performance as a biological control agent of sedentary endo-parasitic nematodes. J Nematol 45:1–7

    PubMed  PubMed Central  Google Scholar 

  • Molnar A, Melnyk CW, Bassett A, Hardcastle TJ, Dunn R (2010) Small silencing RNAs in plants are mobile and direct epigenetic modification in recipient cells. Science 328:872–875

    Article  CAS  PubMed  Google Scholar 

  • Mortazavi A, Williams BA, McCue K, Schaeffer L, Wold B (2008) Mapping and quantifying mammalian transcriptomes by RNA-seq. Nat Methods 5:621–628

    Article  CAS  PubMed  Google Scholar 

  • Nag A, King S, Jack T (2009) miR319a targeting of TCP4 is critical for petal growth and development in Arabidopsis. Proc Natl Acad Sci USA 106:22534–22539

  • 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

    Article  PubMed  PubMed Central  Google Scholar 

  • Nath U, Crawford BCW, Carpenter R, Coen E (2003) Genetic control of surface curvature. Science 299:1404–1407

    Article  CAS  PubMed  Google Scholar 

  • Ouyang S, Park G, Atamian HS, Han CS, Stajich JE, Kaloshian I (2014) MicroRNAs suppress NB domain genes in tomato that confer resistance to Fusarium oxysporum. PLoS Pathog 10:e1004464

    Article  PubMed  PubMed Central  Google Scholar 

  • Peláez P, Sanchez F (2013) SmallRNAs in plant defense responses during viral and bacterial interactions: similarities and differences. Front Plant Sci 4:343

    Article  PubMed  PubMed Central  Google Scholar 

  • Pfaffl MW, Horgan GW, Dempfle L (2002) Relative expression software tool (REST©) for group-wise comparison and statistical analysis of relative expression results in real-time PCR. Nucleic Acids Res 30:e36

    Article  PubMed  PubMed Central  Google Scholar 

  • RNAcentral Consortium (2015) RNAcentral: an international database of ncRNA sequences. Nucleic Acids Res 43:D123–D129

    Article  Google Scholar 

  • Rodriguez PL (1998) Protein phosphatase 2C (PP2C) function in higher plants. Plant Mol Biol 38:919–927

    Article  CAS  PubMed  Google Scholar 

  • Rosso LC, Finetti-Sialer MM, Hirsch PR, Ciancio A, Kerry BR, Clark IM (2011) Transcriptome analysis shows differential gene expression in the saprotrophic to parasitic transition of Pochonia chlamydosporia. Appl Microbiol Biotechnol 90:1981–1994

    Article  CAS  PubMed  Google Scholar 

  • Rosso LC, Pentimone I, Colagiero M, Ferrara M, Nigro F, Ciancio A (2013) Expression of Meloidogyne incognita resistance genes induced by endophytic Pochonia chlamydosporia in tomato. Nematropica 43:322

    Google Scholar 

  • Rueda A, Barturen G, Lebrón R, Gómez-Martín C, Alganza Á, Oliver JL, Hackenberg M (2015) sRNAtoolbox: an integrated collection of small RNA research tools. Nucleic Acids Res 43(W1):W467–W473

  • Ruiz-Ferrer V, Voinnet O (2009) Roles of plant small RNAs in biotic stress responses. Annu Rev Plant Biol 60:485–510

    Article  CAS  PubMed  Google Scholar 

  • 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

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Stone J, Liang X, Nekl E, Stiers J (2005) Arabidopsis AtSPL14, a plant-specific SBP-domain transcription factor, participates in plant development and sensitivity to fumonisin B1. Plant J 41:744–754

  • Subramanian S, Fu Y, Sunkar R, Barbazuk W, Zhu JK (2008) Novel and nodulation-regulated microRNAs in soybean roots. BMC Genomics 9:160

    Article  PubMed  PubMed Central  Google Scholar 

  • Sun G (2011) MicroRNAs and their diverse functions in plants. Plant Mol Biol 18:17–36

    Google Scholar 

  • Takahashi H (2013) Auxin biology in roots. Plant Roots 7:49–64

    Article  CAS  Google Scholar 

  • The Tomato Genome Consortium (2012) The tomato genome sequence provides insights into fleshy fruit evolution. Nature 485:635–641

    Article  Google Scholar 

  • Valencia-Sanchez MA, Liu J, Hannon GJ, Parker R (2006) Control of translation and mRNA degradation by miRNAs and siRNAs. Genes Dev 20:515–524

    Article  CAS  PubMed  Google Scholar 

  • Valiollahi E, Farsi M, Fevereiro P, Kakhki AM (2014) Bioinformatic characterization and expression analysis of miRNAs in Solanum lycopersicum. Plant Omics J 7:108–116

    CAS  Google Scholar 

  • 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. https://doi.org/10.1186/1746-4811-3-12

    Article  PubMed  PubMed Central  Google Scholar 

  • Wang Y, Li K, Chen L, Zou Y, Liu H, Tian Y (2015) MicroRNA167-directed regulation of the auxin response factors GmARF8a and GmARF8b is required for soybean nodulation and lateral root development. Plant Physiol 168:984–999. https://doi.org/10.1104/pp.15.00265

    Article  PubMed  PubMed Central  Google Scholar 

  • Wang Y, Wang L, Zou Y, Chen L, Cai Z, Zhang S (2014) Soybean miR172c targets the repressive AP2 transcription factor NNC1 to activate ENOD40 expression and regulate nodule initiation. Plant Cell 26:4782–4801. https://doi.org/10.1105/tpc.114.131607

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Weiberg A, Wang M, Lin FM, Zhao HW, Zhang ZH, Kaloshian I (2013) Fungal small RNAs suppress plant immunity by hijacking host RNA interference pathways. Science 342:118

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Weiberg A, Wang M, Bellinger M, Jin H (2014) Small RNAs: a new paradigm in plant-microbe interactions. Annu Rev Phytopathol 52:495–516

    Article  CAS  PubMed  Google Scholar 

  • Wu P, Wu Y, Liu CC, Liu LW, Ma FF, XY W, Wu M, Hang YY, Chen JQ, Shao ZQ, Wang B (2016) Identification of arbuscular mycorrhiza (AM)-responsive microRNAs in tomato. Front Plant Sci 7:429. https://doi.org/10.3389/fpls.2016.00429

    PubMed  PubMed Central  Google Scholar 

  • Yang L, Huang H (2014) Roles of small RNAs in plant disease resistance. J Integr Plant Biol 56:962–970

    Article  CAS  PubMed  Google Scholar 

  • Ye W, Shen C-H, Lin Y, Chen P-J, Xu X (2014) Growth promotion-related miRNAs in Oncidium orchid roots colonized by the endophytic fungus Piriformospora indica. PLoS One 9:e84920

    Article  PubMed  PubMed Central  Google Scholar 

  • Yoo BC, Kragler F, Varkonyi-Gasic E, Haywood V, Archer-Evans S, Lee YM, Lough TJ, Lucas WJ (2004) A systemic small RNA signaling system in plants. Plant Cell 16:1979–2000

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Zhang W, Gao S, Zhou X (2011) Bacteria-responsive microRNAs regulate plant innate immunity by modulating plant hormone networks. Plant Mol Biol 75:93–105

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

The authors thank Dr. G. Loconsole, M. Saponari, and A. Gianpetruzzi for assistance with NGS and preparation of sRNA libraries.

Funding

This research was partially funded by projects CISIA (CNR), SELGE (Regione Puglia), BIOMED (MiPAF), and Eureka!Eurostars E!7364 “Poch_art.” I.P. gratefully acknowledges the funding of two Short Term Scientific Missions by COST Action FP1305 “BioLink.” R.L. is supported by the Ministry of Education of Spain (FPU13/05662).

Author information

Authors and Affiliations

  1. Istituto per la Protezione Sostenibile delle Piante, Consiglio Nazionale delle Ricerche, Via G. Amendola 122/D, 70126, Bari, Italy

    Isabella Pentimone, Laura C. Rosso, Mariantonietta Colagiero & Aurelio Ciancio

  2. Genetics Department & Biotechnology Institute, Biomedical Research Center (CIBM), University of Granada, Granada, Spain

    Ricardo Lebrón & Michael Hackenberg

  3. Dipartimento di Scienze del Suolo, della Pianta e degli Alimenti, Università degli Studi di Bari - Aldo Moro, Via G. Amendola 165/A, 70126, Bari, Italy

    Franco Nigro

Authors
  1. Isabella Pentimone
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Ricardo Lebrón
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Michael Hackenberg
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Laura C. Rosso
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Mariantonietta Colagiero
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Franco Nigro
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Aurelio Ciancio
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Isabella Pentimone.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

This article does not contain any studies with human participants or animals performed by any of the authors.

Electronic supplementary material

ESM 1

(PDF 1634 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Pentimone, I., Lebrón, R., Hackenberg, M. et al. Identification of tomato miRNAs responsive to root colonization by endophytic Pochonia chlamydosporia . Appl Microbiol Biotechnol 102, 907–919 (2018). https://doi.org/10.1007/s00253-017-8608-7

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00253-017-8608-7

Keywords

Search

Navigation