Kurşun (II) Asetat ve Salisilik Asidin Medicago sativa L. Bitkisinde Gelişim, Biyokimyasal Parametreler ve miRNA156 Gen Anlatımı Üzerine Etkileri

Year 2025, Volume: 12 Issue: 1, 74 - 86

Mustafa Akçay

Abstract

Bitkiler bulundukları çevre içinde büyüme, gelişim ve fotosentezini engelleyen farklı şekillerde meydana gelebilen olumsuz koşullara maruz kalabilmektedirler. Kurşun (II) asetat, bitkilerin büyüme ve gelişmesini sınırlayan toksik bir ağır metaldir. Salisilik asit, bitkiler tarafından sentezlenebilen ve bitkilerin büyüme ve gelişiminde önemli rol oynayan bir fitohormondur. Bu çalışmada, materyal olarak Türkiye’de tarımı en fazla yapılan ve hayvan beslenmesinde kullanılan yem bitki türü baklagillerinden yonca (Medicago sativa L.) bitkisinin Erzurum genotipi kullanılmış ve kurşun (II) asetat (Pb(CH3COO)2), salisilik asit (SA) ve kombinasyon uygulamalarının bitki üzerinde çeşitli bitki büyüme parametreleri, biyokimyasal analiz ve miRNA ifadesi üzerine etkileri araştırılmıştır. Bitkiler, 25oC sıcaklık 16/8 fotoperiyot ve %60 neme ayarlanmış bitki büyütme kabini içerisinde 28 gün boyunca büyütülmüş ve ardından uygulama aşamasına geçilmiştir. Bitkilere uygulanan konsantrasyon uygulamaları kurşun (II) asetat (100 ve 1000 ppm), salisilik asit (0.5 mM ve 2 mM) ve kombinasyonları şeklinde olup, konsantrasyonlar bitkilerin toprağına 8 gün boyunca 40 ml çözelti şeklinde uygulanmıştır. Bu süre sonunda bitkiler hasat edilerek yapılan uygulamaların kök ve bitki uzunlukları (cm) ile kök ve yaprak yaş ağırlıkları (g), MDA içeriği ve miRNA156 gen anlatım seviyesi üzerine olan etkileri araştırılmıştır. Çalışma sonucunda, kurşun (II) asetat (100 ve 1000 ppm), uygulamalarının bitkinin kök ve bitki uzunlukları ile yaprak ve kök yaş ağırlıklarını azalttığı, MDA içeriğini şiddetli artırdığı ve miRNA156 gen anlatım seviyesini aşağı regüle ederek bitkinin büyüme ve gelişmesini olumsuz etkilediği, buna karşın salisilik asit (0,5 mM ve 2 mM) uygulamaları bitkinin kök ve bitki uzunlukları ile yaprak ve kök yaş ağırlıklarını artırdığı, MDA içeriğini azalttığı ve miRNA156 gen anlatım seviyesini yukarı regüle ederek bitkinin büyüme ve gelişmesini sağladığı tespit edilmiştir.

Keywords

Medicago sativa L., Kurşun (II) asetat, Salisilik asit, Lipit peroksidasyonu, miRNA156

Effects of Lead (II) Acetate and Salicylic Acid on Growth, Biochemical Parameters and miRNA156 Gene Expression in Medicago sativa L.

Abstract

Plants can be exposed to unfavourable conditions in their environment that can occur in different ways that prevent them from growing, developing and photosynthesising. Lead (II) acetate is a toxic heavy metal that limits the growth and development of plants. Salicylic acid is a phytohormone that can be synthesised by plants and plays an important role in plant growth and development. In this study, Erzurum genotype of alfalfa (Medicago sativa L.), one of the most widely cultivated forage legume plants in Türkiye and used in animal nutrition, was used as material and the effects of lead (II) acetate (Pb(CH3COO)2), salicylic acid (SA) and combination treatments on various plant growth parameters, biochemical analyses and miRNA expression were investigated. Plants were grown for 28 days in a plant growth chamber adjusted to 25oC temperature, 16/8 photoperiod and 60% humidity and then the application phase was started. Concentrations of lead (II) acetate (100 and 1000 ppm), salicylic acid (0.5 mM and 2 mM) and their combinations were applied to the soil of the plants as 40 ml solution for 8 days. At the end of this period, the plants were harvested and the effects of the treatments on root and plant lengths (cm), root and leaf wet weights (g), MDA content and miRNA 156 gene expression level were investigated. As a result of the study, lead (II) acetate (100 and 1000 ppm) treatments decreased the root and plant lengths and leaf and root wet weights, increased the MDA content severely and down-regulated the miRNA156 gene expression level and negatively affected the growth and development of the plant, On the other hand, salicylic acid (0.5 mM and 2 mM) treatments increased root and plant lengths, leaf and root wet weights, decreased MDA content, and up-regulated miRNA156 gene expression level and promoted plant growth and development.

Keywords

Medicago sativa L., Lead (II) acetate, Salicylic acid, Lipid peroxidation, miRNA156

References

• AbdElgawad, H., Zinta, G., Hamed, B. A., Selim, S., Beemster, G., Hozzein, W. N. ve Abuelsoud, W. 2020. Maize roots and shoots show distinct profiles of oxidative stress and antioxidant defense under heavy metal toxicity. Environmental Pollution, 258, 113705.
• Ahmad, P., Ozturk, M. ve Gucel, S. 2012. Oxidative damage and antioxidants induced by heavy metal stress in two cultivars of mustard (Brassica juncea L.) plants. Fresenius Environ Bull, 21(10), 2953-2961.
• Akinci, I.E., Akinci, S. ve Yilmaz, K. 2010. Response of tomato (Solanum lycopersicum L.) to lead toxicity: Growth, element uptake, chlorophyll and water content. African Journal of Agricultural Research, 5(6), 416-423.
• Alloway, B.J. 2013. Sources of heavy metals and metalloids in soils. Heavy metals in soils: trace metals and metalloids in soils and their bioavailability, 11-50.
• Altun, H. ve Orcan, P. 2024. Responses to exogenous elicitor treatment in lead-stressed Oryza sativa L. BMC Plant Biology, 24(1), 897.
• Alzahrani, Y., Alharby, H.F., Hakeem, K.R. ve Alsamadany, H. 2020. Modulating effect of EDTA and SDS on growth, biochemical parameters and antioxidant defense system of Dahlia variabilis grown under cadmium and lead-induced stress. Notulae Botanicae Horti Agrobotanici Cluj-Napoca, 48(2), 906-923.
• Arteca, R.N. 1999. Plant growth substances: principles and applications. Springer Science & Business Media. New York, 128 s.
• Avcıoğlu, R., Geren, H., Tamkoç, A. ve Karadağ, Y. 2009. Yonca (Medicago sp. L.). Yem Bitkileri Cilt II: Baklagil Yem Bitkileri. Tarım ve Köyişleri Bakanlığı, Tarımsal Üretim ve Geliştirme Genel Müdürlüğü, İzmir, 293-295.
• Bagautdinova, Z.Z., Omelyanchuk, N., Tyapkin, A.V., Kovrizhnykh, V.V., Lavrekha, V.V. ve Zemlyanskaya, E.V. 2022. Salicylic acid in root growth and development. International Journal of Molecular Sciences, 23(4), 2228.
• Bhatti, K.H., Sehrish Anwar, S.A., Khalid Nawaz, K.N., Khalid Hussain, K.H., Siddiqi, E.H., Sharif, R.U. ve Aneela Khalid, A.K. 2013. Effect of heavy metal Lead (Pb) stress of different concentration on wheat (Triticum aestivum L.). Middle East Journal of Scientific Research, 14(2), 148-154.
• Chen, Y.E., Cui, J.M., Li, G.X., Yuan, M., Zhang, Z.W., Yuan, S. ve Zhang, H.Y. 2016. Effect of salicylic acid on the antioxidant system and photosystem II in wheat seedlings. Biologia plantarum, 60(1), 139-147.
• Dere, S. ve Doğan, M. 2020. Kurşun Uygulamasının Yerfıstığı (Arachis hypogaea L.)’ndaki Morfolojik ve Fizyolojik Etkileri. Türkiye Tarımsal Araştırmalar Dergisi, 7(3), 233-245.
• Deswal, M. ve Laura, J.S. 2018. Effect of heavy metals cadmium, nickel and lead on the seed germination and early seedling growth of Pisum sativum. Research Journal of Life Sciences, Bioinformatics, Pharmaceutikal and Chemical Sciences, 4, 368-383.
• Dimkpa, C.O. ve Bindraban, P.S. 2016. Fortification of micronutrients for efficient agronomic production: a review. Agronomy for Sustainable Development, 36(1), 7.
• Ding, Y., Chen, Z. ve Zhu, C. 2011. Microarray-based analysis of cadmium-responsive microRNAs in rice (Oryza sativa). Journal of experimental botany, 62(10), 3563-3573.
• Duffus, J.H. 2002. "Heavy metals" a meaningless term? (IUPAC Technical Report). Pure and applied chemistry, 74(5), 793-807.
• Eun, S.O., Shik Youn, H. ve Lee, Y. 2000. Lead disturbs microtubule organization in the root meristem of Zea mays. Physiologia plantarum, 110(3), 357-365.
• Fu, Z.Q., Yan, S., Saleh, A., Wang, W., Ruble, J., Oka, N. ve Dong, X. 2012. NPR3 and NPR4 are receptors for the immune signal salicylic acid in plants. Nature, 486(7402), 228-232.
• Gan, C.D., Chen, T. ve Yang, J.Y. 2020. Remediation of vanadium contaminated soil by alfalfa (Medicago sativa L.) combined with vanadium-resistant bacterial strain. Environmental Technology & Innovation, 20, 101090.
• Gao, J., Luo, M., Peng, H., Chen, F. ve Li, W. 2019. Characterization of cadmium-responsive MicroRNAs and their target genes in maize (Zea mays) roots. BMC Molecular Biology, 20, 1-9.
• Gezer, E. 2011. Arpa (hordeum vulgare l.) Bitkisinin bazı çeşitlerinde ağır metal stresi etkilerinin fizyolojik olarak araştırılması., Sakarya Üniversitesi, Yüksek Lisans Tezi, 1-43
• Ghani, A., Shah, A.U. ve Akhtar, U. 2010. Effect of lead toxicity on growth, chlorophyll and lead (Pb) content of two varieties of maize (Zea mays L.). Pakistan Journal of Nutrition, 9(9): 887891.
• Godbold, D.L. ve Kettner, C. 1991. Lead influences root growth and mineral nutrition of Picea abies seedlings. Journal of plant physiology, 139(1), 95-99.
• Gottesfeld, P., Were, F.H., Adogame, L., Gharbi, S., San, D., Nota, M.M. ve Kuepouo, G. 2018. Soil contamination from lead battery manufacturing and recycling in seven African countries. Environmental research, 161, 609-614.
• Hara, M., Furukawa, J., Sato, A., Mizoguchi, T. ve Miura, K. 2012. Abiotic stress and role of salicylic acid in plants. Abiotic stress responses in plants: metabolism, productivity and sustainability, 235-251.
• He, Q., Zhu, S. ve Zhang, B. 2014. MicroRNA–target gene responses to lead-induced stress in cotton (Gossypium hirsutum L.). Functional & integrative genomics, 14, 507-515.
• Heath, R.L. ve Packer, L. 1968. Photoperoxidation in isolated chloroplasts: I. Kinetics and stoichiometry of fatty acid peroxidation. Archives of biochemistry and biophysics, 125(1), 189-198.
• Hussain, A., Abbas, N., Arshad, F., Akram, M., Khan, Z.I., Ahmad, K. ve Mirzaei, F. 2013. Effects of diverse doses of Lead (Pb) on different growth attributes of Zea-Mays L. Agricultural Sciences, 4(5), 262-265.
• Jones-Rhoades, M.W. 2012. Conservation and divergence in plant microRNAs. Plant molecular biology, 80, 3-16.
• Kaya, A.R., Eryigit, T., Uslu, O.S., Gedik, O. Ve Tuncturk, M. 2019. Effects of lead on seed germination and seedling growth in different sesame (Sesamum İndicum) genotypes. Fresenius Environmental Bulletin, 28(9), 6574-6579.
• Kıran, S., Özkay, F., Kuşvuran, Ş. ve Ellialtıoğlu, Ş. 2015. Kurşunun kıvırcık salata (lactuca sativa var. Crispa) bitkisinin bazı morfolojik ve biyokimyasal özelliklerine etkisi. Iğdır Üniversitesi Fen Bilimleri Enstitüsü Dergisi, 5(1), 83-88.
• Kieffer, P., Dommes, J., Hoffmann, L., Hausman, J.F. ve Renaut, J. 2008. Quantitative changes in protein expression of cadmium‐exposed poplar plants. Proteomics, 8(12), 2514-2530.
• Kopittke, P.M., Asher, C.J., Blamey, F.P.C. ve Menzies, N.W. 2007. Toxic effects of Pb 2+ on the growth and mineral nutrition of signal grass (Brachiaria decumbens) and Rhodes grass (Chloris gayana). Plant and Soil, 300, 127-136.
• Li, X., Schuler, M.A. ve Berenbaum, M.R. 2002. Jasmonate and salicylate induce expression of herbivore cytochrome P450 genes. Nature, 419(6908), 712-715.
• Li, Z., Ma, Z., Hao, X., Rensing, C. Ve Wei, G. 2014. Genes conferring copper resistance in Sinorhizobium meliloti CCNWSX0020 also promote the growth of Medicago lupulina in copper-contaminated soil. Applied and Environmental Microbiology, 80(6), 1961-1971.
• Livak K.J ve Schmittgen T.D. 2001 Analysis of relative gene expression data using real-time quantitative PCR and the 2− ΔΔCT method. Methods, 25(4), 402-408.
• Małkowski, E., Kita, A., Galas, W., Karcz, W. Ve Kuperberg, J.M. 2002. Lead distribution in corn seedlings (Zea mays L.) and its effect on growth and the concentrations of potassium and calcium. Plant Growth Regulation, 37, 69-76.
• Min Yang, Z. ve Chen, J. 2013. A potential role of microRNAs in plant response to metal toxicity. Metallomics, 5(9), 1184-1190.
• Nriagu, J.O. 1992. Toxic metal pollution in Africa. Science of the Total Environment, 121, 1-37.
• Obroucheva, N.V., Bystrova, E.I., Ivanov, V.B., Antipova, O.V. ve Seregin, I.V. 1998. Root growth responses to lead in young maize seedlings. In Root Demographics and Their Efficiencies in Sustainable Agriculture, Grasslands and Forest Ecosystems: Proceedings of the 5th Symposium of the International Society of Root Research, held 14–18 July 1996 at Madren Conference Center, Clemson University, Clemson, South Carolina, USA, Springer Netherlands, 445-456.
• Ouzounidou, G., Moustakas, M. ve Eleftheriou, E.P. 1997. Physiological and ultrastructural effects of cadmium on wheat (Triticum aestivum L.) leaves. Archives of Environmental Contamination and Toxicology, 32, 154-160.
• Ozkose, A. 2018. Effect of environment x cultivar interaction on protein and mineral contents of alfalfa (Medicago sativa L.) in central Anatolia, Turkey. Sains Malays, 47(3), 551-562.
• Pourrut, B., Jean, S., Silvestre, J. Ve Pinelli, E. 2011. Lead-induced DNA damage in Vicia faba root cells: potential involvement of oxidative stress. Mutation Research/Genetic Toxicology and Environmental Mutagenesis, 726(2), 123-128.
• Prasad, M.N.V., Hagemeyer, J., Devi, S.R. ve Prasad, M.N.V. 1999. Membrane lipid alterations in heavy metal exposed plants. Heavy metal stress in plants: from molecules to ecosystems, Berlin Heidelberg, 99-116.
• Radović, J., Sokolović, D. Ve Marković, J.J.B.A.H. 2009. Alfalfa-most important perennial forage legume in animal husbandry. Biotechnology in Animal Husbandry, 25(5-6-1), 465-475.
• Rai, P.K., Lee, S.S., Zhang, M., Tsang, Y.F. ve Kim, K.H. 2019. Heavy metals in food crops: Health risks, fate, mechanisms, and management. Environment international, 125, 365-385.
• Rascio, N. ve Navari-Izzo, F. 2011. Heavy metal hyperaccumulating plants: how and why do they do it? And what makes them so interesting?. Plant science, 180(2), 169-181.
• Raskin, I. 1992. Role of salicylic acid in plants. Annual review of plant biology, 43(1), 439-463.
• Shriram, V., Kumar, V., Devarumath, R.M., Khare, T.S. ve Wani, S.H. 2016. MicroRNAs as potential targets for abiotic stress tolerance in plants. Frontiers in Plant Science, 7, 817.
• Shu, Y., Liu, Y., Li, W., Song, L., Zhang, J. Ve Guo, C. 2016. Genome-wide investigation of microRNAs and their targets in response to freezing stress in Medicago sativa L., based on high-throughput sequencing. G3:Genes, Genomes, Genetics, 6(3), 755-765.
• Song, W., Zheng, A., Shao, H., Chu, L., Brestic, M. ve Zhang, Z. 2012. The alleviative effect of salicylic acid on the physiological indices of the seedling leaves in six different wheat genotypes under lead stress. Plant Omics, 5(5), 486-493.
• Song, Y., Lv, J., Ma, Z. ve Dong, W. 2019. The mechanism of alfalfa (Medicago sativa L.) response to abiotic stress. Plant Growth Regulation, 89, 239-249.
• Tabande, L., Sepehri, M., Yasrebi, J., Zarei, M., Ghasemi-Fasaei, R. ve Khatabi, B. 2022. A comparison between the function of Serendipita indica and Sinorhizobium meliloti in modulating the toxicity of zinc oxide nanoparticles in alfalfa (Medicago sativa L.). Environmental Science and Pollution Research, 1-14.
• Xie, K., Wu, C. ve Xiong, L. 2006. Genomic organization, differential expression, and interaction of SQUAMOSA promoter-binding-like transcription factors and microRNA156 in rice. Plant physiology, 142(1), 280-293.
• Xu, M., Hu, T., Zhao, J., Park, M.Y., Earley, K.W., Wu, G. ve Poethig, R.S. 2016. Arabidopsis thaliana'daki miR156 tarafından düzenlenen SQUAMOSA PROMOTER BINDING PROTEIN-BENZERİ (SPL) genlerinin gelişimsel işlevleri. PLoS genetiği , 12 (8), e1006263.
• Yang, Y., Wei, X., Lu, J., You, J., Wang, W. Ve Shi, R. 2010. Lead-induced phytotoxicity mechanism involved in seed germination and seedling growth of wheat (Triticum aestivum L.). Ecotoxicology and environmental safety, 73(8), 1982-1987.
• Yilmaz, H.Ş. 2015. Bazı ağır metal uygulamalarının sorgum (Sorghum bicolor L.) bitkisinin enzim aktivitesi üzerine etkisi. Erciyes Üniversitesi, Fen Bilimleri Enstitüsü, Yüksek Lisans Tezi, 1-74.
• Yolcu, H. ve Tan, M. 2008. Ülkemiz yem bitkileri tarımına genel bir bakış. Tarım Bilimleri Dergisi, 14(3), 303-312.
• Zhang, F., Liu, M., Li, Y., Che, Y. ve Xiao, Y. 2019. Effects of arbuscular mycorrhizal fungi, biochar and cadmium on the yield and element uptake of Medicago sativa. Science of the Total Environment, 655, 1150-1158.
• Zhou, Z.S., Huang, S.Q. ve Yang, Z.M. 2008. Bioinformatic identification and expression analysis of new microRNAs from Medicago truncatula. Biochemical and biophysical research communications, 374(3), 538-542.