Distinctive physiological and molecular responses to cold stress among cold-tolerant and cold-sensitive Pinus halepensis seed sources

[EN] BackgroundForest species ranges are confined by environmental limitations such as cold stress. The natural range shifts of pine forests due to climate change and proactive-assisted population migration may each be constrained by the ability of pine species to tolerate low temperatures, especially in northern latitudes or in high altitudes. The aim of this study is to characterize the response of cold-tolerant versus cold-sensitive Pinus halepensis (P. halepensis) seedlings at the physiological and the molecular level under controlled cold conditions to identify distinctive features which allow us to explain the phenotypic difference. With this objective gas-exchange and water potential was determined and the photosynthetic pigments, soluble sugars, glutathione and free amino acids content were measured in seedlings of different provenances under control and cold stress conditions.ResultsGlucose and fructose content can be highlighted as a potential distinctive trait for cold-tolerant P. halepensis seedlings. At the amino acid level, there was a significant increase and accumulation of glutathione, proline, glutamic acid, histidine, arginine and tryptophan along with a significant decrease of glycine.ConclusionOur results established that the main difference between cold-tolerant and cold-sensitive seedlings of P. halepensis is the ability to accumulate the antioxidant glutathione and osmolytes such as glucose and fructose, proline and arginine.This study is a part of the research project: "Application of molecular biology techniques in forest restoration in Mediterranean environments, PAID-05-11" funded by the Universitat Politecnica de Valencia (UPV), program for supporting R&D of new multidisciplinary research lines. The authors are grateful to the Ministerio de Economia y Competitividad AGL2014-57431-P and BIO2016-77776-P. AV was supported by project Survive-2 (CGL2015-69773-C2-2-P MINECO/FEDER) by the Spanish Government and Prometeo program (DESESTRES Generalitat Valenciana). CEAM is funded by Generalitat Valenciana. None of the funding bodies was involved in the design of the study; collection, analysis, and interpretation of data; and in writing of the manuscript which were performed entirely by the signing authors.Taïbi, K.; Campo García, ADD.; Vilagrosa, A.; Belles Albert, JM.; López-Gresa, MP.; López-Nicolás, JM.; Mulet, JM. (2018). Distinctive physiological and molecular responses to cold stress among cold-tolerant and cold-sensitive Pinus halepensis seed sources. BMC Plant Biology. 18:1-11. https://doi.org/10.1186/s12870-018-1464-511118IPCC: Climate Change 2007. Impacts, Adaptation and Vulnerability Contribution of Working Group II to the Fourth Assessment Report of the IPCC (978 0521 88010–7 Hardback; 978 0521 70597–4 Paperback).Giorgi F, Lionello P. Climate change projections for the Mediterranean region. Glob Planet Change. 2008;63:90–104.Petoukhov V, Semenov VA. A link between reduced Barents-Kara Sea ice and cold winter extremes over northern continents. J Geophys Res. -Atmos. 2010;115:D21111.Kodra E, Steinhaeuser K, Ganguly AR. Persisting cold extremes under 21st-century warming scenarios. Geophys Res Lett. 2011;38:L08705.Kreyling J. Winter climate change: a critical factor for temperate vegetation performance. Ecology. 2010;91:1939–48.Yildiz D, Nzokou P, Deligoz A, Koc I, Genc M. Chemical and physiological responses of four Turkish red pine (Pinus brutia Ten.) provenances to cold temperature treatments. Eur J Forest Res. 2014;133:809. https://doi.org/10.1007/s10342-014-0798-2 .Bannister P, Neuner G. Frost resistance and the distribution of conifers. In: Bigras FJ, Colombo SJ, editors. Conifer Cold Hardiness. Tree Physiology, vol. 1. Dordrecht: Springer; 2001. p. 3–21.Thomashow MF. Plant cold acclimation, freezing tolerance genes and regulatory mechanisms. Annu Rev Plant Phys. 1999;50:571–99.Kreyling J, Wiesenberg GLB, Thiel D, Wohlfart C, Huber G, Walter J, Jentsch A, Konnert M, Beierkuhnlein C. Cold hardiness of Pinus nigra Arnold as influenced by geographic origin, warming, and extreme summer drought. Env Exp Bot. 2010;78:99–108.Alonso-Blanco C, Gomez-Mena C, Llorente F, Koornneef M, Salinas J, Martínez-Zapater JM. Genetic and molecular analyses of natural variation indicate CBF2 as a candidate gene for underlying a freezing tolerance quantitative trait locus in Arabidopsis. Plant Phys. 2005;139:1304–12. https://doi.org/10.1104/pp.105.068510 .Colombo SJ, Zhao S, Blumwald E. Frost hardiness gradients in shoots and roots of Picea mariana seedlings. Scand J For Res. 1995;10:32–6.Porcel R, Bustamante A, Ros R, Serrano R, Mulet JM. BvCOLD1: a novel aquaporin from sugar beet (Beta vulgaris L.) involved in boron homeostasis and abiotic stress. Plant Cell Environ. 2018:1–14. https://doi.org/10.1111/pce.13416 .Pardos M, Royo A, Gil L, Pardos JA. Effect of nursery location and outplanting date on field performance of Pinus halepensis and Quercus ilex seedlings. Forestry. 2003;76:67–81.Puértolas J, Gil L, Pardos JA. Effects of nitrogen fertilization and temperature on frost hardiness of Aleppo pine (Pinus halepensis mill .) seedlings assessed by chlorophyll fluorescence. Forestry. 2005;78:501–11.Taïbi K, del Campo AD, Mulet JM, Flors J, Aguado A. Testing Aleppo pine seed sources response to climate change by using trial sites reflecting future conditions. New For. 2014. https://doi.org/10.1007/s11056-014-9423-y .Taïbi K, del Campo AD, Aguado A, Mulet JM. The effect of genotype by environment interaction, phenotypic plasticity and adaptation on Pinus halepensis reforestation establishment under expected climate drifts. Ecol Eng. 2015;84:218–28. https://doi.org/10.1016/j.ecoleng.2015.09.005 .Taïbi K, del Campo AD, Vilagrosa A, Bellés JM, López-Gresa MP, Pla D, Calvete JJ, López-Nicolás JM, Mulet JM. Drought tolerance in Pinus halepensis seed sources as identified by distinctive physiological and molecular markers. Front Plant Sci. 2017;8:1202. https://doi.org/10.3389/fpls.2017.01202 .Baquedano FJ, Valladares F, Castillo FJ. Phenotypic plasticity blurs ecotypic divergence in the response of Quercus coccifera and Pinus halepensis to water stress. Eur J Forest Res. 2008;127:495–506.Van den Ende W, Valluru R. Sucrose, sucrosyl oligosaccharides, and oxidative stress: scavenging and salvaging? J Exp Bot. 2009;60:9–18.Keunen E, Peshev D, Vangronsveld J, Van den Ende W, Cuypers A. Plant sugars are crucial players in the oxidative challenge during abiotic stress: extending the traditional concept. Plant Cell Environ. 2013;36:1242–55.Mulet JM, Llopis-Torregrosa V, Primo C, Marqués MC, Yenush L. Endocytic regulation of alkali metal transport proteins in mammals, yeast and plants. Curr Genet. 2013;59:207. https://doi.org/10.1007/s00294-013-0401-2 .Van den Ende W. Multifunctional fructans and raffinose family oligosaccharides. Front Plant Sci. 2013;4:247.Cao YY, Yang MT, Li X, Zhou ZQ, Wang XJ, Bai JG. Exogenous sucrose increases chilling tolerance in cucumber seedlings by modulating antioxidant enzyme activity and regulating proline and soluble sugar contents. Sci Hortic. 2014;179:67–77.Mulet JM, Alemany B, Ros R, Calvete JJ, Serrano R. Expression of a plant serine O-acetyltransferase in Saccharomyces cerevisiae confers osmotic tolerance and creates an alternative pathway for cysteine biosynthesis. Yeast. 2004;21(4):303–12.Pyngrope S, Bhoomika K, Dubey RS. Reactive oxygen species, ascorbate–glutathione pool, and enzymes of their metabolism in drought-sensitive and tolerant indica rice (Oryza sativa L.) seedlings subjected to progressing levels of water deficit. Protoplasma. 2013;250:585–600.Chen THH, Uemura M, Fujikawa S. Cold hardiness in plants: molecular genetics, cell biology, and physiology. Wallingford: CABI Publishing UK; 2006.Odlum K, Blake T, Kim Y, Glerum C. Influence of photoperiod and temperature on frost hardiness and free amino acid concentrations in black spruce seedlings. Tree Physiol. 1993;13:275–82. https://doi.org/10.1093/treephys/13.3.275 .Kim YT, Glerum C. Seasonal free amino-acid fluctuations in red pine and white spruce needles. Can J For Res. 1995;25:697–703. https://doi.org/10.1139/x95-077 .Vicent I, Navarro A, Mulet JM, Sharma S, Serrano R. Uptake of inorganic phosphate is a limiting factor for Saccharomyces cerevisiae during growth at low temperatures. FEMS Yeast Res. 2015;15:1–13. https://doi.org/10.1093/femsyr/fov008 .Angelcheva L, Mishra Y, Antti H, Kjellsen TD, Funk C, Strimbeck RG, et al. Metabolomic analysis of extreme freezing tolerance in Siberian spruce (Picea obovata). New Phytol. 2014;204:545–55. https://doi.org/10.1111/nph.12950 .Villar-Salvador P, Peñuelas Rubira JL, Vallas CJ. Dessication patterns of Pinus halepensis seedlings grown in different types of containers. Cuad Soc Esp Cien For. 2004;17:93–9.Hoagland DR, Arnon DI. The water-culture method for growing plants without soil, Vol. 347, 2nd Edn, Berkeley: University of California, College of Agriculture, Agricultural Experiment Station, 32; 1950.Maxwell K, Johnson GN. Chlorophyll fluorescence- a practical guide. J Exp Bot. 2008;51:659–68.Lichtenthaler HK. Chlorophylls and carotenoids: pigments of photosynthetic biomembranes. Method Enzymol. 1987;148:350–82.Fayos J, Bellés JM, López-Gresa MP, Primo J, Conejero V. Induction of gentisic acid 5-O-beta-D-xylopyranoside in tomato and cucumber plants infected by different pathogens. Phytochemistry. 2006;67:142–8