Kuraklık Stresi Altındaki Buğday Çeşitlerinde Salisilik Asit Uygulamalarının miRNA İfadesi Üzerine Etkileri

Öz

Türkiye’de buğday yetiştiriciliği yapılan alanlarda bitki büyümesi, kuraklık stresi gibi çevresel bir faktörden yüksek oranda etkilenmektedir ve bu abiyotik stres tarımsal ürünlerin verimini önemli derecede düşürmektedir. Bitki büyümesini uyaran kimyasalların bitkilere uygulanması stres toleransını arttırmak için kolay, düşük maliyetli, düşük riskli ve etkili bir yaklaşımdır ve bunlar içerisinde salisilik asit kuraklık stresi, tuzluluk gibi olumsuz çevre koşullarına bitkilerin uyum sağlamasında ve bitkilerin strese dayanıklılık geliştirmesinde ön plana çıkmaktadır. Bu çalışmada kuraklığa toleranslı çeşit olarak Gün91, hassas çeşit olarak Ankara98 çeşiti kullanılmıştır ve bitkilere iki farklı doz (0.1 mM, 0.5mM) salisilik asit uygulaması yapılmıştır. Salisilik asit uygulamasından bir hafta sonra kuraklık stresi uygulaması gerçekleştirilmiştir. Kuraklık stresi uygulamasının sonunda uygulama ve kontrol saksılarından toplanan yaprak örneklerinden RNA’lar izole edilmiş ve miR156, miR169, miR172, miR319 ile mir398 ve bu miRNA’lara ait hedef genlerin ifade seviyeleri belirlenmiştir. Kuraklık stres koşulları altında salisilik asit ile muamele edilen yaprak dokularında; miR156 ve miR172'nin ekspresyon seviyesi kuraklık stresine toleranslı Gün91 çeşidinde yüksek iken, miR169 seviyesi hassas Ankara98 çeşidinde daha yüksek olarak belirlenmiştir. miR319 ifade seviyesi Gün91 çeşidinde 0.5mM salisilik asit+kuraklık uygulamasında daha yüksek iken, Ankara98 çeşidinde 0.1mM salisilik asit+kuraklık uygulamasında daha yüksek olarak gözlenmiştir. miR398 ekspresyon seviyesi, hem Gün91, hem de Ankara98 çeşitlerinde salisilik asit ile kuraklık stresinin birlikte uygulandığı bitki gruplarında miR398 ifade seviyesi kontrol grubuna göre belirgin azalma göstermiştir. Salisilik asit uygulamasının tek başına yapıldığı gruplarda ise miR398 ifade seviyesi daha yüksek olarak belirlenmiştir. Sonuç olarak; bu çalışma ile salisilik asidin strese yanıtta önemli bir sinyal molekülü olduğu ve stres koşullarındaki etkiyi azalttığı ve stres tolerans mekanizmasının uygulanan salisilik asit ile kuraklığa toleranslı ve hassas buğday çeşitlerinde farklı olduğu vurgulanmıştır.

Anahtar Kelimeler

• Buğday,Kuraklık,miRNA,Salisilik asit

The Effects of Salicylic Acid Applications on miRNA Expression in Wheat Varieties Under Drought Stress

Abstract

Wheat cultivation in Turkey; is highly affected by environmental factors, such as drought stress, which reduces yields significantly. Applying chemicals that stimulate plant growth is an easy, low-cost, low-risk and effective approach to increase stress tolerance, and salicylic acid is prominent in adapting plants to adverse environmental conditions such as drought stress, salinity, and improving plants' tolerance to stress. In the study, Gün91 was used as drought tolerant variety and Ankara98 was used as the sensitive variety and two different doses (0.1 mM, 0.5mM) of salicylic acid were applied to the plants. Drought stress application was performed one week after salicylic acid application and RNAs were isolated from leaf samples collected from application and control pots, and expression levels of miR156, miR169, miR172, miR319 and mir398 and target genes belonging to these miRNAs were determined. While the expression level of miR156 and miR172 was higher in Day91 with tolerance to drought stress in leaf tissues treated with salicylic acid under drought stress conditions; the level of miR169 was higher in the sensitive Ankara98 variety. In Gün91 variety, miR319 expression level was higher in 0.5 mM salicylic acid + drought application, whereas in Ankara98 variety was higher in 0.1 mM salicylic acid + drought application. The miR398 expression level showed a significant decrease in both Gün91 and Ankara98 varieties in the plant groups where salicylic acid and drought stress were applied together compared to the control group. As a result; with this study, it was emphasized that salicylic acid is an important signal molecule in response to stress and it mitigates the effect in stress conditions, and the tolerance mechanism of drought tolerant and sensitive varieties with salicylic acid application were different.

Keywords

• Wheat,Drought,miRNA,Salicylic acid

Kaynakça

1. Alonso-Peral, M. M., Sun, C., Millar, A. A. (2012). MicroRNA159 can act asa switch or tuning microRNA independently of its abundance in Arabidopsis. PLoS ONE, 7, e34751.
2. Bhupinder, S., Usha, K. (2003). Salicylic acid induced physiological and biochemical changes in wheat seedlings under water stress. Plant Growth Regulation, 39, 137-141.
3. Brenchley, R., Spannagl, M., Pfeifer, M., Barker, G. L. A., D’Amore1, R., Allen, A. M., McKenzie, N., Kramer, M., Kerhornou, A., Bolser, D., Kay, S., Waite, D., Trick, M., Bancroft, I., Gu, Y., Huo, N., Luo, M. C., Sehgal, S., Gill, B., Kianian, S., Anderson, O., Kersey, P., Dvorak, J., McCombie, W. R., Hall, A., Mayer, K. F. X., Edwards, K. J., Bevan, M. W., Hall, N. (2012). Analysis of the bread wheat genome using whole-genome shotgun sequencing. Nature, 2012, 705-710.
4. Flavell, R. B., Smith, D. B. (1976). Nucleotide sequence organisation in the wheat genome. Heredity: an International Journal of Genetics, 37, 231-252.
5. Frazier, T., Sun, G., Burklew, C., Zhang, B. (2011). Salt and Drought Stresses Induce the Aberrant Expression of microRNA Genes in Tobacco. Molecular Biotechnology, 49, 159-65.
6. Hamada, A. M, Al-Hakimi, A. M. A. (2001). Salicylic acid versus salinity-drought- induced stres on wheat seedlings. Rostlinna Vyroba, 47, 444-450.
7. Hwang, E. W., Shin, S. J., Park, S. C., Jeong, M. J., Kwon, H. B. (2011). Identification of miR172 family members and their putative targets responding to drought stress in Solanum tuberosum. Genes and Genomics, 33, 105-110.
8. Inal, B., Turktas, M., Eren, H., Ilhan, E., Okay, S., Atak, M., Erayman, M., Unver, T. (2014). Genome-wide fungal stress responsive miRNA expression in wheat. Planta, 240, 1287-1298.

9. Kantar, M., Unver, T., Budak, H. (2010). Regulation of Barley miRNAs Upon Dehydration Stres Correlated with Target Gene Expression. Functional Integrative Genomics, 10, 493-507.
10. Liu, H. H., Tian, X., Li, Y. J., Wu, C. A., Zheng, C. C. (2008). Microarray-based analysis of stress-regulated microRNAs in Arabidopsis thaliana. RNA, 14, 836-43.
11. Livak K. J., Schmittgen T. D. (2001). Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCT method. Methods, 25, 402–408.
12. Mehmood, Z., Ashiq, M., Noorka, I. R., Ali, A., Tabasum, S., Iqbal, M. S. (2014). Chemical Control of Monocot Weeds in Wheat (Triticum aestivum L.). American Journal of Plant Sciences, 5, 1272-1276.
13. Miyashita, K., Tanakamaru, S., Maitani, T., Kimura, K. (2005). Recovery Responses of Photosynthesis, Transpiration, and Stomatal Conductance in Kidney Bean Following Drought Stress. Environmental and Experimental Botany, 53, 205-214.
14. Rahaie, M., Xue, G. P., Naghavi, M. R., Alizadeh, H., Schenk, P. M. (2010). A MYB gene from wheat (Triticum aestivum L.) is up-regulated during salt and drought stresses and differentially regulated between salt-tolerant and sensitive genotypes. Plant Cell Reports, 29, 835-844.
15. Ramanjulu S., Yang-Fang, L., Guru, J. (2012). Funtions of microRNAs in plant stress responses. Trends in Plant Science, 4, 196-203.
16. Senaratna, T., Touchell, D., Bunn, E., Dixon, K. (2000). Acetyl salicylic acid (Aspirin) and salicylic acid induce multiple stress tolerance in bean and tomato plants. Plant Growth Regulation, 30, 157-161.
17. Sunkar, R., Li, Y. F., Jagadeeswaran, G. (2012). Functions of microRNAs in plant stress responses. Trends in Plant Science, 17, 196-203.
18. Shi, Q., Bao, Z., Zhu, Z., Ying, Q., Qian, Q. (2006). Effects of different treatments of salicylic acid on heat tolerance, chlorophyll fluorescence and antioxidant enzyme activity in seedlings of Cucumis sativa L. Plant Growth Regulation, 48, 127-135.
19. Shi, Q., Zhu, Z. (2008). Effects of exogenous salicylic acid on manganese toxicity, element contents and antioxidative system in cucumber. Enviromental and Experimental Botany, 63, 317-326.
20. Snyman, M., Cronjé, M. J. (2008). Modulation of heat shock factors accompanies salicylic acid-mediated potentiation of Hsp70 in tomato seedlings. Journal of Experimental Botany, 59, 2125-2132.
21. Sunkar, R., Li, Y. F., Jagadeeswaran, G. (2012). Functions of microRNAs in plant stres responses. Trends in Plant Science, 17, 196-203.
22. Xin, M. W. Y., Yao, Y., Song, N., Hu, Z., Qin, D., Xie, C., Peng, H., Ni, Z., Sun, Q. (2011). Identification and characterization of wheat long non-protein coding RNAs responsive to powdery mildew infection and heat stress by using microarray analysis and SBS sequencing. BMC Plant Biology, 11, 1-13.
23. Yang, J., Zhang, N., Ma, C., Qu, Y., Si, H., Wang, D. (2013). Prediction and verification of microRNAs related to prolin accumulation under drought stress in potato. Computational Biology and Chemistry, 46, 48-54.
24. Zhang, N., Yang, J., Wang, Z., Wen, Y., Wang, J., He, W., Liu, B., Si, H., Wang, D. (2014). Identification of Novel and Conserved MicroRNAs Related to Drought Stress in Potato by Deep Sequencing. PLoS ONE, 9, e95489.
25. Zhou, M., Li, D., Li, Z., Hu, Q., Yang, C., Zhu, L., Luo, H. (2013). Constitutive expression of a miR319 gene alters plant development and enhances salt and drought tolerance in transgenic creeping bentgrass. Plant Physiology, 161, 1375-1391.
26. Zhua, C., Dinga, Y. Liua, H. (2011). miR398 and Plant Stress Responses. Physiologia Plantarum, 143, 1-9.