Small RNA Transcriptome of Hibiscus Syriacus Provides Insights into the Potential Influence of microRNAs in Flower Development and Terpene Synthesis

  • Kim, Taewook (Department of Agricultural Biotechnology, Seoul National University) ;
  • Park, June Hyun (Department of Agricultural Biotechnology, Seoul National University) ;
  • Lee, Sang-gil (Program in Applied Life Chemistry, Seoul National University) ;
  • Kim, Soyoung (Department of Agricultural Biotechnology, Seoul National University) ;
  • Kim, Jihyun (Program in Applied Life Chemistry, Seoul National University) ;
  • Lee, Jungho (Green Plant Institute) ;
  • Shin, Chanseok (Department of Agricultural Biotechnology, Seoul National University)
  • Received : 2017.05.26
  • Accepted : 2017.07.11
  • Published : 2017.08.31


MicroRNAs (miRNAs) are essential small RNA molecules that regulate the expression of target mRNAs in plants and animals. Here, we aimed to identify miRNAs and their putative targets in Hibiscus syriacus, the national flower of South Korea. We employed high-throughput sequencing of small RNAs obtained from four different tissues (i.e., leaf, root, flower, and ovary) and identified 33 conserved and 30 novel miRNA families, many of which showed differential tissuespecific expressions. In addition, we computationally predicted novel targets of miRNAs and validated some of them using 5' rapid amplification of cDNA ends analysis. One of the validated novel targets of miR477 was a terpene synthase, the primary gene involved in the formation of disease-resistant terpene metabolites such as sterols and phytoalexins. In addition, a predicted target of conserved miRNAs, miR396, is SHORT VEGETATIVE PHASE, which is involved in flower initiation and is duplicated in H. syriacus. Collectively, this study provides the first reliable draft of the H. syriacus miRNA transcriptome that should constitute a basis for understanding the biological roles of miRNAs in H. syriacus.


development;flowering initiation;Hibiscus syriacus;microRNA;small RNA;terpene synthesis


Supported by : Rural Development Administration


  1. Afzal, A.J., Wood, A.J., and Lightfoot, D.A. (2008). Plant receptorlike serine threonine kinases: roles in signaling and plant defense. Mol. Plant-Microbe Int. 21, 507-517.
  2. Aukerman, M.J., and 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.
  3. Barkan, A., and Small, I. (2014). Pentatricopeptide repeat proteins in plants. Annu. Rev. Plant Biol. 65, 415-442.
  4. Boualem, A., Laporte, P., Jovanovic, M., Laffont, C., Plet, J., Combier, J.-P., Niebel, A., Crespi, M., and Frugier, F. (2008). MicroRNA166 controls root and nodule development in Medicago truncatula. Plant J. 54, 876-887.
  5. Camacho, C., Coulouris, G., Avagyan, V., Ma, N., Papadopoulos, J., Bealer, K., and Madden, T.L. (2009). BLAST+: architecture and applications. BMC Bioinformatics 10, 421-421.
  6. Casadevall, R., Rodriguez, R.E., Debernardi, J.M., Palatnik, J.F., and Casati, P. (2013). Repression of growth regulating factors by the MicroRNA396 inhibits cell proliferation by UV-B radiation in arabidopsis leaves. Plant Cell 25, 3570-3583.
  7. Cuperus, J.T., Fahlgren, N., and Carrington, J.C. (2011). Evolution and functional diversification of MIRNA cenes. Plant Cell 23, 431-442.
  8. Dai, X., and Zhao, P.X. (2011). psRNATarget: a plant small RNA target analysis server. Nucleic Acids Res. 39, W155-W159.
  9. Gutierrez, L., Bussell, J.D., Păcurar, D.I., Schwambach, J., Păcurar, M., and Bellini, C. (2009). Phenotypic plasticity of adventitious rooting in arabidopsis is controlled by complex regulation of AUXIN RESPONSE FACTOR transcripts and microRNA abundance. Plant Cell 21, 3119-3132.
  10. Hwang, D.-G., Park, J.H., Lim, J.Y., Kim, D., Choi, Y., Kim, S., Reeves, G., Yeom, S.-I., Lee, J.-S., Park, M., et al. (2013). The Hot Pepper (Capsicum annuum) MicroRNA Transcriptome reveals novel and conserved targets: a foundation for understanding microRNA functional roles in hot pepper. Plos One 8, e64238.
  11. Kim, J., Jung, J.-H., Reyes, J.L., Kim, Y.-S., Kim, S.-Y., Chung, K.-S., Kim, J.A., Lee, M., Lee, Y., Narry Kim, V., et al. (2005). microRNAdirected cleavage of ATHB15 mRNA regulates vascular development in Arabidopsis inflorescence stems. Plant J. 42, 84-94.
  12. Kim, J., Park, J.H., Lim, C.J., Lim, J.Y., Ryu, J.-Y., Lee, B.-W., Choi, J.-P., Kim, W.B., Lee, H.Y., Choi, Y., et al. (2012). Small RNA and transcriptome deep sequencing proffers insight into floral gene regulation in Rosa cultivars. BMC Genomics 13, 657.
  13. Kim, Y.-M., Kim, S., Koo, N., Shin, A.-Y., Yeom, S.-I., Seo, E., Park, S.- J., Kang, W.-H., Kim, M.-S., Park, J., et al. (2017). Genome analysis of Hibiscus syriacus provides insights of polyploidization and indeterminate flowering in woody plants. DNA Res. 24, 71-80.
  14. Koyama, T., Furutani, M., Tasaka, M., and Ohme-Takagi, M. (2007). TCP transcription factors control the morphology of shoot lateral organs via negative regulation of the expression of boundary-specific genes in arabidopsis. Plant Cell 19, 473-484.
  15. Kozomara, A., and Griffiths-Jones, S. (2014). miRBase: annotating high confidence microRNAs using deep sequencing data. Nucleic Acids Res. 42, D68-D73.
  16. Langmead, B., Trapnell, C., Pop, M., and Salzberg, S.L. (2009). Ultrafast and memory-efficient alignment of short DNA sequences to the human genome. Genome Biol. 10, R25.
  17. Lee, J.H., Yoo, S.J., Park, S.H., Hwang, I., Lee, J.S., and Ahn, J.H. (2007). Role of SVP in the control of flowering time by ambient temperature in Arabidopsis. Genes Dev. 21, 397-402.
  18. Li, Y., Zhang, Q., Zhang, J., Wu, L., Qi, Y., and Zhou, J.-M. (2010). Identification of microRNAs involved in pathogen-associated molecular pattern-triggered plant innate immunity. Plant Physiol. 152, 2222-2231.
  19. Meyers, B.C., Axtell, M.J., Bartel, B., Bartel, D.P., Baulcombe, D., Bowman, J.L., Cao, X., Carrington, J.C., Chen, X., and Green, P.J. (2008). Criteria for annotation of plant MicroRNAs. Plant Cell 20, 3186-3190.
  20. Moxon, S., Jing, R., Szittya, G., Schwach, F., Rusholme Pilcher, R.L., Moulton, V., and Dalmay, T. (2008). Deep sequencing of tomato short RNAs identifies microRNAs targeting genes involved in fruit ripening. Genome Res. 18, 1602-1609.
  21. Nag, A., King, S., and Jack, T. (2009). miR319a targeting of TCP4 is critical for petal growth and development in Arabidopsis. Proc. Natl. Acad. Sci. USA 106, 22534-22539.
  22. Nozawa, M., Miura, S., and Nei, M. (2012). Origins and evolution of microRNA genes in plant species. Genome Biol. Evol. 4, 230-239.
  23. Pantaleo, V., Szittya, G., Moxon, S., Miozzi, L., Moulton, V., Dalmay, T., and Burgyan, J. (2010). Identification of grapevine microRNAs and their targets using high-throughput sequencing and degradome analysis. Plant J. 62, 960-976.
  24. Park, J.H., and Shin, C. (2014). MicroRNA-directed cleavage of targets: mechanism and experimental approaches. BMB Rep. 47, 417-423.
  25. Park, J.H., and Shin, C. (2015). The role of plant small RNAs in NBLRR regulation. Brief. Func. Genomics. 14, 268-274.
  26. Peng, J., Xia, Z., Chen, L., Shi, M., Pu, J., Guo, J., and Fan, Z. (2014). Rapid and Efficient Isolation of High-Quality Small RNAs from Recalcitrant Plant Species Rich in Polyphenols and Polysaccharides. Plos One 9, e95687.
  27. Rodriguez, R.E., Mecchia, M.A., Debernardi, J.M., Schommer, C., Weigel, D., and Palatnik, J.F. (2010). Control of cell proliferation in Arabidopsis thaliana by microRNA miR396. Development 137, 103-112.
  28. Rogers, K., and Chen, X. (2013). Biogenesis, Turnover, and mode of action of plant microRNAs. Plant Cell Online.
  29. Si-Ammour, A., Windels, D., Arn-Bouldoires, E., Kutter, C., Ailhas, J., Meins, F., and Vazquez, F. (2011). miR393 and secondary siRNAs regulate expression of the TIR1/AFB2 auxin receptor clade and auxinrelated development of Arabidopsis leaves. Plant Physiol. 157, 683-691.
  30. Trotta, E. (2014). On the normalization of the minimum free energy of RNAs by sequence length. Plos One 9, e113380.
  31. Vidal, E.A., Araus, V., Lu, C., Parry, G., Green, P.J., Coruzzi, G.M., and Gutierrez, R.A. (2010). Nitrate-responsive miR393/AFB3 regulatory module controls root system architecture in Arabidopsis thaliana. Proc. Natl. Acad. Sci. USA 107, 4477-4482.
  32. Voinnet, O. (2009). Origin, biogenesis, and activity of plant microRNAs. Cell 136, 669-687.
  33. Wang, K., Senthil-Kumar, M., Ryu, C.-M., Kang, L., and Mysore, K.S. (2012). Phytosterols play a key role in plant innate immunity against bacterial pathogens by regulating nutrient efflux into the apoplast. Plant Physiol. 158, 1789-1802.
  34. Williams, L., Grigg, S.P., Xie, M., Christensen, S., and Fletcher, J.C. (2005). Regulation of Arabidopsis shoot apical meristem and lateral organ formation by microRNA miR166g and its AtHD-ZIP target genes. Development 132, 3657-3668.
  35. Wu, G., Park, M.Y., Conway, S.R., Wang, J.-W., Weigel, D., and Poethig, R.S. (2009). The sequential action of miR156 and miR172 regulates developmental timing in Arabidopsis. Cell 138, 750-759.
  36. Wu, M.-F., Tian, Q., and Reed, J.W. (2006). Arabidopsis microRNA167 controls patterns of ARF6 and ARF8 expression, and regulates both female and male reproduction. Development 133, 4211-4218.
  37. Yang, C.-Y., Huang, Y.-H., Lin, C.-P., Lin, Y.-Y., Hsu, H.-C., Wang, C.-N., Liu, L.-Y.D., Shen, B.-N., and Lin, S.-S. (2015). MicroRNA396-targeted SHORT VEGETATIVE PHASE is required to repress flowering and is related to the development of abnormal flower symptoms by the phyllody symptoms1 effector. Plant Physiol. 168, 1702-1716.
  38. Yu, N., Niu, Q.-W., Ng, K.-H., and Chua, N.-H. (2015). The role of miR156/SPLs modules in Arabidopsis lateral root development. Plant J. 83, 673-685.
  39. Zhang, W., Gao, S., Zhou, X., Xia, J., Chellappan, P., Zhou, X., Zhang, X., and Jin, H. (2010). Multiple distinct small RNAs originate from the same microRNA precursors. Genome Biol. 11, R81-R81.
  40. Zhou, M., Li, D., Li, Z., Hu, Q., Yang, C., Zhu, L., and Luo, H. (2013). Constitutive expression of a miR319 gene alters plant development and enhances salt and drought tolerance in transgenic creeping bentgrass. Plant Physiol. 161, 1375-1391.
  41. Zhu, Q.-H., and Helliwell, C.A. (2011). Regulation of flowering time and floral patterning by miR172. J. Exp. Bot. 62, 487-495.
  42. Rubio-Somoza, I., and Weigel, D. (2013). Coordination of flower maturation by a regulatory circuit of three microRNAs. PLoS Genet. 9, e1003374.

Cited by

  1. Transcriptomic analyses of rice (Oryza sativa) genes and non-coding RNAs under nitrogen starvation using multiple omics technologies vol.19, pp.1, 2018,