Sensitivity to high salinity in tetraploid citrus seedlings increases with water availability and correlates with expression of candidate genes
Wafa Mouhaya A B D , Thierry Allario A B D , Javier Brumos B , Fernando Andrés B , Yann Froelicher A , François Luro C , Manuel Talon B , Patrick Ollitrault A and Raphaël Morillon A B EA Centre de Cooperation Internationale en Recherche Agronomique pour le Développement (CIRAD), UPR Amélioration Génétique des Espèces Multiplication Végétative, Instituto Valenciano de Investigaciones Agrarias (IVIA), 46113 Moncada, Valencia, Spain.
B Centro de Genomica, Instituto Valenciano de Investigaciones Agrarias (IVIA), 46113 Moncada, Valencia, Spain.
C Unité Génétique et Ecophysiologie de la Qualité des Agrumes, Station Institut Nationale de la Recherche Agronomique de San Giuliano, 20230 San Giuliano, France.
D These authors contributed equally to the paper.
E Corresponding author. Email: raphael.morillon@cirad.fr
Functional Plant Biology 37(7) 674-685 https://doi.org/10.1071/FP10035
Submitted: 18 February 2010 Accepted: 23 May 2010 Published: 2 July 2010
Abstract
We investigated tolerance to high salinity in well-irrigated diploid and tetraploid citrus. Comparisons were made between two diploids (2×) of trifoliate orange (Poncirus trifoliata (L.) Raf.) and willow leaf mandarin (Citrus deliciosa Ten), their respective doubled diploids (4×) and the allotetraploid (FLHORAG1) obtained from the protoplast fusion of trifoliate orange and Willow leaf mandarin. Salinity stress was applied by progressively increasing the concentration of NaCl from 50 mM to 400 mM for 8 weeks. Two-year-old plants were watered daily. Maximum quantum yield of PSII, and leaf and root chloride and sodium content were monitored. We previously reported that under moderate saline stress, citrus 4× genotypes were more tolerant that the 2×, but under these experimental conditions, 4× seedlings were certainly more sensitive to salt stress than 2×, as they accumulated more toxic ions and were more affected than 2×. Chloride accumulation in 4× leaves was greater and the maximum quantum yield of PSII was more reduced in 4× than in 2×. The expression of several candidate genes involved in signal transduction, sodium and chloride transport, osmotic adjustment, regulation of the stomata opening and detoxification processes were also investigated by quantitative real-time reverse transcription-PCR. A high correlation was observed between phenotype of sensitivity to stress and gene expression changes.
Additional keywords: polyploidy, rootstocks, salt stress.
Acknowledgements
We thank C. Jacquemond, F. Curk (Unité GEQA, INRA, San Giuliano, France) for providing 2× seedlings. The authors thank the anonymous reviewers for their comments that helped to improve the manuscript. Funding was provided by Comité Mixte Inter Universitaire Franco-Marocain’ (Program Volubilis, MA/05/1 37) for WM and European INCO project (FP6 – 2003 - INCO – DEV – 2 n° 015453) for TA. Interreg IIIA program, Ministerio de Educacion y Ciencia, AGL 2007–65437-C04–01/AGR provided funding.
Adams KL,
Cronn R,
Percifield R, Wendel JF
(2003) Genes duplicated by polyploidy show unequal contributions to the transcriptome and organ-specific reciprocal silencing. Proceedings of the National Academy of Sciences of the United States of America 100, 4649–4654.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Alós E,
Cercós M,
Rodrigo MJ,
Zacarías L, Talón M
(2006) Regulation of color break in citrus fruits. Changes in pigment profiling and gene expression induced by gibberellins and nitrate, two ripening retardants. Journal of Agricultural and Food Chemistry 54, 4888–4895.
| Crossref | GoogleScholarGoogle Scholar | PubMed |
Apse MP,
Aharon GS,
Snedden WA, Blumwald E
(1999) Salt tolerance conferred by overexpression of a vacuolar Na+/H+ antiport in Arabidopsis. Science 285, 1256–1258.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Auger DL,
Gray AD,
Ream TS,
Kato A,
Coe EH, Birchler JA
(2005) Nonadditive gene expression in diploid and triploid hybrids of maize. Genetics 169, 389–397.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Bañuls J, Primo-Millo E
(1995) Effect of salinity on some Citrus scion–rootstock combinations. Annals of Botany 76, 97–102.
| Crossref | GoogleScholarGoogle Scholar |
Banuls J,
Serna MD,
Legaz M, Primo-Millo E
(1997) Growth and gaz exchange parameters of citrus plants stressed with different salts. Journal of Plant Physiology 150, 194–199.
|
CAS |
Barrett HC
(1985) Hybridization of citrus and related genera. Fruit Varieties Journal, University Park 39, 11–16.
Bassene JB,
Froelicher Y,
Dhuique-Mayer C,
Mouhaya W,
Mar Ferrer R,
Gema Ancillo G,
Morillon R,
Navarro L, Ollitrault P
(2009) Non-additive phenotypic and transcriptomic inheritance in a citrus allotetraploid somatic hybrid between C. reticulate and C. limon: the case of pulp carotenoid biosynthesis pathway. Plant Cell Reports 28, 1689–1697.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Brumós J,
Colmenero-Flores JM,
Conesa A,
Izquierdo P,
Sánchez G,
Iglesias DJ,
López-Climent MF,
Gómez-Cadenas A, Talon M
(2009) Membrane transporters and carbon metabolism implicated in chloride homeostasis differentiate salt stress responses in tolerant and sensitive Citrus rootstocks. Functional & Integrative Genomics 9, 293–309.
| Crossref | GoogleScholarGoogle Scholar | PubMed |
Bryan GJ,
McNicoll J,
Ramsay G,
Meyer RC, Jong WSD
(1999) Polymorphic simple sequence repeat markers in chloroplast genomes of solanaceous plants. Theoretical and Applied Genetics 99, 859–867.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
Chomczynski P, Sacchi N
(1987) Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Analytical Biochemistry 162, 156–159.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Colmenero-Flores JM,
Martinez G,
Gamba G,
Vazquez N,
Iglesias DJ,
Brumos J, Talon M
(2007) Identification and functional characterization of cation–chloride cotransporters in plants. The Plant Journal 50, 278–292.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Cooper WC, Gorton BS
(1952) Toxicity and accumulation of chloride salts in Citrus on various rootstocks. Proceedings of the American Society of Horticultural Science 59, 143–146.
|
CAS |
Dai X,
Xu Y,
Ma Q,
Xu W,
Wang T,
Xue Y, Chong K
(2007) Overexpression of a R1R2R3 MYB gene, OsMYB3R–2, increases tolerance to freezing, drought, and salt stress in transgenic Arabidopsis. Plant Physiology 143, 1739–1751.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Davenport SV,
Gallego SM,
Benavides MP, Tomaro ML
(2004) Behaviour of antioxidant defense system in the adaptive response to salt stress in Helianthus annuus L. cells. Plant Growth Regulation 40, 81–88.
| Crossref | GoogleScholarGoogle Scholar |
Dure L
(1993) A repeating 1 1-mer amino acid motif and plant dessication. The Plant Journal 3, 363–369.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Elgazzar A,
Wallace A, Hemaidan N
(1965) Sodium distribution in rough lemon and trifoliate orange seedlings. Soil Science 99, 387–391.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
Feierabend J,
Schaan C, Hertwig B
(1992) Photoinactivation of catalase occurs under both high- and low-temperature stress conditions and accompanies photoinhibition of photosystem II. Plant Physiology 100, 1554–1561.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Froelicher Y,
Bassene JB,
Jedidi-Neji E,
Dambier D,
Morillon R,
Bernardini G,
Costantino G, Ollitrault P
(2007) Induced parthenogenesis in mandarin for haploid production: induction procedures and genetic analysis of plantlets. Plant Cell Reports 26, 937–944.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Froelicher Y,
Dambier D,
Bassene JB,
Costantino G, Lotfy S ,
et al
.
(2008) Characterization of microsatellite markers in mandarin orange (Citrus reticulata Blanco). Molecular Ecology Resources 8, 119–122.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
Froelicher Y,
Mouhaya W,
Bassene JP,
Costantino G,
Kamiri M,
Luro F,
Morillon R, Ollitrault P
(2010) New universal mitochondrial PCR markers reveal new information on maternal citrus phylogeny. Tree Genetics & Genomes in press. ,
Gancel AL,
Grimplet J,
Sauvage FX,
Ollitrault P, Brillouet JM
(2006) Predominant expression of diploid mandarin leaf proteome in two citrus mandarin-derived somatic allotetraploid hybrids. Journal of Agricultural and Food Chemistry 54, 6212–6218.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Grosser JW,
Louzada ES,
Gmitter FG, Chandler JL
(1994) Somatic hybridization of complementary citrus rootstocks: five new hybrids. HortScience 29, 812–813.
Grosser JW,
Ollitrault P, Olivares-Fuster O
(2000) Somatic hybridization in Citrus: an effective tool to facilitate variety improvement. In Vitro Cellular & Developmental Biology. Plant 36, 434–449.
| Crossref | GoogleScholarGoogle Scholar |
Guo WW,
Cheng YJ, Deng XX
(2002) Regeneration and molecular characterization of intergeneric somatic hybrids between Citrus reticulata and Poncirus trifoliate. Plant Cell Reports 20, 829–834.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
Guo Y,
Qiu QS,
Quintero FJ,
Pardo JM,
Ohta M,
Zhang C,
Schumaker KS, Zhua JK
(2004) Transgenic evaluation of activated mutant alleles of SOS2 reveals a critical requirement for its kinase activity and C-terminal regulatory domain for salt tolerance in Arabidopsis thaliana. The Plant Cell 16, 435–449.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Hundertmark M, Hincha DK
(2008) LEA (Late Embryogenesis Abundant) proteins and their encoding genes in Arabidopsis thaliana. BMC Genomics 9, 118.
| Crossref | GoogleScholarGoogle Scholar | PubMed |
Iglesias DJ,
Cercós M,
Colmenero-Flores JM,
Naranjo MA,
Ríos G,
.
(2008) Physiology of citrus fruiting. Brazilian Journal of Plant Physiology 19, 333–362.
Jellings AJ, Leech RM
(1984) Anatomical variation in first leaves on nine Triticum genotypes and its relationship to photosynthetic capacity. New Phytologist 96, 371–382.
| Crossref | GoogleScholarGoogle Scholar |
Leitch AR, Leitch IJ
(2008) Genomic plasticity and the diversity of polyploid plants. Science 320, 481–483.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Li WL,
Berlyn GP, Ashton MS
(1996) Polyploids and their structural and physiological characteristics relative to water deficit in Betula papyrifera (Betulaceae). American Journal of Botany 83, 15–20.
| Crossref | GoogleScholarGoogle Scholar |
Luro F,
Costantino G,
Terol J,
Argout X,
Allario T,
Wincker P,
Talon M,
Ollitrault P, Morillon R
(2008) Transferability of the EST-SSRs developed on Nules clementine (Citrus clementina Hort ex Tan) to other Citrus species and their effectiveness for genetic mapping. BMC Genomics 9, 287.
| Crossref | GoogleScholarGoogle Scholar | PubMed |
Moons A,
Bauw G,
Prinsen E,
Van Montagu M, Straeten DVD
(1995) Molecular and physiological responses to abscisic acid and salts in roots of salt-sensitive and salt-tolerant Indica rice varieties. Plant Physiology 107, 177–186.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Moya JL,
Primo-Millo E, Talon M
(1999) Morphological factors determining salt tolerance in citrus seedlings: the shoot to root ratio modulates passive root uptake of chloride ions and their accumulation in leaves. Plant, Cell & Environment 22, 1425–1433.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
Moya JL,
Gómez-Cadenas A,
Primo-Millo E, Talon M
(2003) Chloride absorption in salt-sensitive Carrizo citrange and salt-tolerant Cleopatra mandarin citrus rootstocks is linked to water usage. Journal of Experimental Botany 54, 825–833.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Morillon R, Chrispeels MJ
(2001) The role of ABA and the transpiration stream in the regulation of the osmotic water permeability of leaf cells. Proceedings of the National Academy of Sciences of the United States of America 98, 14 138–14 143.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Naot D,
Ben-Hayyim G,
Eshdat Y, Holland D
(1995) Drought, heat and salt stress induce the expression of a citrus homologue of an atypical late-embryogenesis Lea5 gene. Plant Molecular Biology 27, 619–622.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Ollitrault P,
Dambier D,
Seker M, Froelicher Y
(2000) Rootstock breeding by somatic hybridisation for the Mediterranean citrus industry. Acta Horticulturae 535, 157–162.
Pustovoitova TN,
Eremin GV,
Rassvetaeva EG,
Zhdanova NE, Zholkevich VN
(1996) Drought resistance, recovery capacity, and phytohormone content in polyploid plum leaves. Russian Journal of Plant Physiology: a Comprehensive Russian Journal on Modern Phytophysiology 43, 232–235.
|
CAS |
Riddle NC,
Jiang H,
An L,
Doerge RW, Birchler JA
(2010) Gene expression analysis at the intersection of ploidy and hybridity in maize. Theoretical and Applied Genetics 120, 341–353.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Rodrigo MJ,
Alquezar B, Zacarías L
(2006) Cloning and characterization of two 9-cisepoxycarotenoid dioxygenase genes, differentially regulated during fruit maturation and under stress conditions, from orange (Citrus sinensis L. Osbeck). Journal of Experimental Botany 57, 633–643.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Romero-Aranda R,
Bondada BR,
Syvertsen JP, Grosser JW
(1997) Leaf characteristics and net gas exchange of diploid and autotetraploid Citrus. Annals of Botany 79, 153–160.
| Crossref | GoogleScholarGoogle Scholar |
Romero-Aranda R,
Moya JL,
Tadeo FR,
Legaz F,
Primo-Millo E, Talon M
(1998) Physiological and anatomical disturbances induced by chloride salts in sensitive and tolerant citrus: beneficial and detrimental effects of cations. Plant, Cell & Environment 21, 1243–1253.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
Sakamoto A, Murata N
(2002) The role of glycine betaine in the protection of plants from stress: clues from transgenic plants. Plant, Cell & Environment 25, 163–171.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Saleh B,
Allario T,
Dambier D,
Ollitrault P, Morillon R
(2008) Tetraploid citrus rootstocks are more tolerant to salt stress than diploid. Comptes Rendues de l ‘Académie des Science Biologies 331, 703–710.
| Crossref | GoogleScholarGoogle Scholar |
Shi H,
Lee B-H,
Wu S-J, Zhu J-K
(2003) Overexpression of a plasma membrane Na+/H+ antiporter gene improves salt tolerance in Arabidopsis thaliana. Nature Biotechnology 21, 81–85.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Stupar RM,
Bhaskar PB,
Yandell BS,
Rensink WA, Hart AL ,
et al
.
(2007) Phenotypic and transcriptomic changes associated with potato autopolyploidization. Genetics 176, 2055–2067.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Subbarao GV,
Wheeler RM,
Levine LH, Stutte GW
(2001) Glycine betaine accumulation, ionic and water relations of red-beet at contrasting levels of sodium supply. Journal of Plant Physiology 158, 767–776.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Syvertsen JP,
Lee LS, Grosser JW
(2000) Limitations on growth and net gas exchange of diploid and tetraploid Citrus rootstock cultivars grown at elevated CO2. Journal of the American Society for Horticultural Science 125, 228–234.
Tadeo F,
Cercós M,
Colmenero-Flores JM,
Iglesias DI, Naranjo MA ,
et al
.
(2008) Molecular physiology of development and quality of citrus. Advances in Botanical Research 47, 147–223.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
Terol J,
Conesa A,
Colmenero JM,
Cercos M, Tadeo FR ,
et al
.
(2007) Analyses of 13 000 unique Citrus clusters associated with fruit quality, production and salinity tolerance. BMC Genomics 5, 8–31.
Talon M, Gmitter FG
(2008) Citrus genomics. International Journal of Plant Genomics ,
| Crossref | GoogleScholarGoogle Scholar | PubMed |
Tozlu I,
Moore GA, Guy CL
(2000) Effects of increasing NaCl concentration on stem elongation, dry mass production, and macro- and micro-nutrient accumulation in Poncirus trifoliata. Plant Physiology 27, 35–42.
|
CAS |
Ueda A,
Kathiresan A,
Inada M,
Narita Y,
Nakamura T,
Shi W,
Takabe T, Bennett J
(2004) Osmotic stress in barley regulates expression of a different set of genes than salt stress does. Journal of Experimental Botany 55, 2213–2218.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Ward R, Durrett R
(2004) Subfunctionalization: how often does it occur? How long does it take? Theoretical Population Biology 66, 93–100.
| Crossref | GoogleScholarGoogle Scholar | PubMed |
Warner DA,
Ku MSB, Edwards GE
(1987) Photosynthesis, leaf anatomy, and cellular constituents in polyploidy C4 grass Panicum virgatum. Plant Physiology 84, 461–466.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Weising K, Gardner RC
(1999) A set of conserved PCR primers for the analysis of simple sequence repeat polymorphisms in chloroplast genomes of dicotyledonousangiosperms. Genome 42, 9–19.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Xiong Y-C,
Li F-M, Zhang T
(2006) Performance of wheat crops with different chromosome ploidy: root-sourced signals, drought tolerance, and yield performance. Planta 224, 710–718.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |
Zekri M, Parsons LR
(1992) Salinity tolerance of citrus rootstocks: effect of salt on root and leaf mineral concentrations. Plant and Soil 147, 171–181.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
Zhang XY,
Hu CG, Yao JL
(2010) Tetraploidization of diploid Dioscorea results in activation of the antioxidant defense system and increased heat tolerance. Journal of Plant Physiology 167, 88–94.
| Crossref | GoogleScholarGoogle Scholar |
CAS |
PubMed |