Candidate genes expression profiling during wilting in chickpea caused by Fusarium oxysporum f. sp. ciceris race 5

Autoři: Cristina Caballo aff001;  Patricia Castro aff002;  Juan Gil aff002;  Teresa Millan aff002;  Josefa Rubio aff001;  Jose V. Die aff002
Působiště autorů: Área de Genómica y Biotecnología, IFAPA, Alameda del Obispo, Córdoba, Spain aff001;  Department of Genetics - ETSIAM, University of Córdoba, Campus de Rabanales, Córdoba, Spain aff002
Vyšlo v časopise: PLoS ONE 14(10)
Kategorie: Research Article
doi: 10.1371/journal.pone.0224212


Chickpea production may be seriously threatened by Fusarium wilt, a disease caused by the soil-borne fungus Fusarium oxysporum f. sp. ciceris. F. oxysporum race 5 is the most important race in the Mediterranean basin. Recently, the region responsible for resistance race 5 has been delimited within a region on chromosome 2 that spans 820 kb. To gain a better understanding of this genomic region, we used a transcriptomic approach based on quantitative real-time PCR to analyze the expression profiles of 22 selected candidate genes. We used a pair of near-isogenic lines (NILs) differing in their sensitivity to Fusarium race 5 (resistant vs susceptible) to monitor the transcriptional changes over a time-course experiment (24, 48, and 72 hours post inoculation, hpi). Qualitative differences occurred during the timing of regulation. A cluster of 12 genes were induced by the resistant NIL at 24 hpi, whereas a second cluster contained 9 genes induced by the susceptible NIL at 48 hpi. Their possible functions in the molecular defence of chickpea is discussed. Our study provides new insight into the molecular defence against Fusarium race 5 and demonstrates that development of NILs is a rich resource to facilitate the detection of candidate genes. The new genes regulated here may be useful against other Fusarium races.

Klíčová slova:

Fungi – Fusarium – Fusarium oxysporum – Gene expression – Gene regulation – Polymerase chain reaction – Transcription factors – Transcriptional control


1. FAOSTAT (2017) Retrieved January 15, 2019 from

2. Chen W, Sharma HC, Hari C, Muehlbauer FJ, Frederick J, American Phytopathological Society. Compendium of chickpea and lentil diseases and pests. St. Paul, Minn: APS Press; 2011.

3. Recorbet G, Steinberg C, Olivain C, Edel V, Trouvelot S, Dumas-Gaudot E, et al. Wanted: pathogenesis-related marker molecules for Fusarium oxysporum. New Phytol. 2003;159: 73–92. doi: 10.1046/j.1469-8137.2003.00795.x

4. Cho S, Muehlbauer FJ. Genetic effect of differentially regulated fungal response genes on resistance to necrotrophic fungal pathogens in chickpea (Cicer arietinum L.). Physiological and Molecular Plant Pathology. 2004;64: 57–66. doi: 10.1016/j.pmpp.2004.07.003

5. Jiménez-Díaz RM, Castillo P, Jiménez-Gasco M del M, Landa BB, Navas-Cortés JA. Fusarium wilt of chickpeas: Biology, ecology and management. Crop Prot. 2015;73: 16–27. doi: 10.1016/j.cropro.2015.02.023

6. Kaiser W, Alcala-Jimenez A, Hervas-Vargas A, Trapero-Casas J, Jiménez-Díaz R. Screening of wild Cicer species for resistance to races 0 to 5 of Fusarium oxysporum f. sp ciceris. Plant Dis. 1994;78:962–967

7. Sharma KD, Muehlbauer FJ. Fusarium wilt of chickpea: physiological specialization, genetics of resistance and resistance gene tagging. Euphytica. 2007;157: 1–14. doi: 10.1007/s10681-007-9401-y

8. Jendoubi W, Bouhadida M, Boukteb A, Béji M, Kharrat M. Fusarium Wilt Affecting Chickpea Crop. Agriculture. 2017;7: 23. doi: 10.3390/agriculture7030023

9. Tekeoglu M, Tullu A, Kaiser WJ, Muehlbauer FJ. Inheritance and Linkage of Two Genes that Confer Resistance to Fusarium Wilt in Chickpea. Crop Sci. 2000;40: 1247. doi: 10.2135/cropsci2000.4051247x

10. Winter P, Benko-Iseppon AM, Hüttel B, Ratnaparkhe M, Tullu A, Sonnante G, et al. A linkage map of the chickpea (Cicer arietinum L.) genome based on recombinant inbred lines from a C. arietinum×C. reticulatum cross: localization of resistance genes for fusarium wilt races 4 and 5. Theor Appl Genet. 2000;101: 1155–1163. doi: 10.1007/s001220051592

11. Castro P, Pistón F, Madrid E, Millán T, Gil J, Rubio J. Development of chickpea near-isogenic lines for Fusarium wilt. Theor Appl Genet. 2010;121: 1519–1526. doi: 10.1007/s00122-010-1407-5 20652529

12. Mannur DM, Babbar A, Thudi M, Sabbavarapu MM, Roorkiwal M, Yeri SB, et al. Super Annigeri 1 and improved JG 74: two Fusarium wilt-resistant introgression lines developed using marker-assisted backcrossing approach in chickpea (Cicer arietinum L.). Mol Breeding. 2019;39: 2. doi: 10.1007/s11032-018-0908-9 30631246

13. Ashraf N, Basu S, Narula K, Ghosh S, Tayal R, Gangisetty N, et al. Integrative network analyses of wilt transcriptome in chickpea reveal genotype dependent regulatory hubs in immunity and susceptibility. Sci Rep. 2018;8: 6528. doi: 10.1038/s41598-018-19919-5 29695764

14. Gupta S, Chakraborti D, Sengupta A, Basu D, Das S. Primary metabolism of chickpea is the initial target of wound inducing early sensed Fusarium oxysporum f. sp. ciceri race I. PLoS ONE. 2010;5: e9030. doi: 10.1371/journal.pone.0009030 20140256

15. Gupta S, Bhar A, Chatterjee M, Das S. Fusarium oxysporum f.sp. ciceri race 1 induced redox state alterations are coupled to downstream defense signaling in root tissues of chickpea (Cicer arietinum L.). PLoS ONE. 2013;8: e73163. doi: 10.1371/journal.pone.0073163 24058463

16. Gupta S, Bhar A, Das S. Understanding the molecular defence responses of host during chickpea–Fusarium interplay: where do we stand? Functional Plant Biol. 2013;40: 1285. doi: 10.1071/FP13063

17. Gupta S, Bhar A, Chatterjee M, Ghosh A, Das S. Transcriptomic dissection reveals wide spread differential expression in chickpea during early time points of Fusarium oxysporum f. sp. ciceri Race 1 attack. PLoS ONE. 2017;12: e0178164. doi: 10.1371/journal.pone.0178164 28542579

18. Chatterjee M, Gupta S, Bhar A, Chakraborti D, Basu D, Das S. Analysis of root proteome unravels differential molecular responses during compatible and incompatible interaction between chickpea (Cicer arietinum L.) and Fusarium oxysporum f. sp. ciceri Race1 (Foc1). BMC Genomics. 2014;15: 949. doi: 10.1186/1471-2164-15-949 25363865

19. Kumar Y, Dholakia BB, Panigrahi P, Kadoo NY, Giri AP, Gupta VS. Metabolic profiling of chickpea-Fusarium interaction identifies differential modulation of disease resistance pathways. Phytochemistry. 2015;116: 120–129. doi: 10.1016/j.phytochem.2015.04.001 25935544

20. Kumar Y, Zhang L, Panigrahi P, Dholakia BB, Dewangan V, Chavan SG, et al. Fusarium oxysporum mediates systems metabolic reprogramming of chickpea roots as revealed by a combination of proteomics and metabolomics. Plant Biotechnol J. 2016;14: 1589–1603. doi: 10.1111/pbi.12522 26801007

21. Sharma M, Sengupta A, Ghosh R, Agarwal G, Tarafdar A, Nagavardhini A, et al. Genome wide transcriptome profiling of Fusarium oxysporum f sp. ciceris conidial germination reveals new insights into infection-related genes. Sci Rep. 2016;6: 37353. doi: 10.1038/srep37353 27853284

22. Upasani ML, Limaye BM, Gurjar GS, Kasibhatla SM, Joshi RR, Kadoo NY, et al. Chickpea-Fusarium oxysporum interaction transcriptome reveals differential modulation of plant defense strategies. Sci Rep. 2017;7: 7746. doi: 10.1038/s41598-017-07114-x 28798320

23. Caballo C, Madrid E, Gil J, Chen W, Rubio J, Millan T. Saturation of genomic region implicated in resistance to Fusarium oxysporum f. sp. ciceris race 5 in chickpea. Mol Breeding. 2019;39: 16. doi: 10.1007/s11032-019-0932-4

24. Ali L, Madrid E, Varshney RK, Azam S, Millan T, Rubio J, et al. Mapping and identification of a Cicer arietinum NSP2 gene involved in nodulation pathway. Theor Appl Genet. 2014;127: 481–488. doi: 10.1007/s00122-013-2233-3 24247237

25. Ali L, Deokar A, Caballo C, Tar’an B, Gil J, Chen W, et al. Fine mapping for double podding gene in chickpea. Theor Appl Genet. 2016;129: 77–86. doi: 10.1007/s00122-015-2610-1 26433827

26. Jendoubi W, Bouhadida M, Millan T, Kharrat M, Gil J, Rubio J, et al. Identification of the target region including the Foc0 1 /foc0 1 gene and development of near isogenic lines for resistance to Fusarium Wilt race 0 in chickpea. Euphytica. 2016;210: 119–133. doi: 10.1007/s10681-016-1712-4

27. Sharma KD, Chen W, Muehlbauer FJ. Genetics of Chickpea Resistance to Five Races of Fusarium Wilt and a Concise Set of Race Differentials for Fusarium oxysporum f. sp. ciceris. Plant Dis. 2005;89: 385–390. doi: 10.1094/PD-89-0385 30795454

28. Bhatti MA. Effects of inoculum density and temperature on root rot and wilt of chickpea. Plant Dis. 1992;76: 50. doi: 10.1094/PD-76-0050

29. Die JV, Román B. RNA quality assessment: a view from plant qPCR studies. J Exp Bot. 2012;63: 6069–6077. doi: 10.1093/jxb/ers276 23045609

30. Schmittgen TD, Livak KJ. Analyzing real-time PCR data by the comparative C(T) method. Nat Protoc. 2008;3: 1101–1108. 18546601

31. Die JV, Obrero Á, González-Verdejo CI, Román B. Characterization of the 3’:5’ ratio for reliable determination of RNA quality. Anal Biochem. 2011;419: 336–338. doi: 10.1016/j.ab.2011.08.012 21889484

32. Gurjar GS, Giri AP, Gupta VS. Gene Expression Profiling during Wilting in Chickpea Caused by Fusarium oxysporum F. sp. Ciceri. AJPS. 2012;03: 190–201. doi: 10.4236/ajps.2012.32023

33. Zuker M. Mfold web server for nucleic acid folding and hybridization prediction. Nucleic Acids Res. 2003;31: 3406–3415. doi: 10.1093/nar/gkg595 12824337

34. Ramakers C, Ruijter JM, Deprez RHL, Moorman AFM. Assumption-free analysis of quantitative real-time polymerase chain reaction (PCR) data. Neurosci Lett. 2003;339: 62–66. doi: 10.1016/s0304-3940(02)01423-4 12618301

35. Castro P, Román B, Rubio J, Die JV. Selection of reference genes for expression studies in Cicer arietinum L.: analysis of cyp81E3 gene expression against Ascochyta rabiei. Mol Breeding. 2011;29: 261–274. doi: 10.1007/s11032-010-9544-8

36. Die JV, Gil J, Millan T. Genome-wide identification of the auxin response factor gene family in Cicer arietinum. BMC Genomics. 2018;19: 301. doi: 10.1186/s12864-018-4695-9 29703137

37. Czechowski T, Stitt M, Altmann T, Udvardi MK, Scheible W-R. Genome-wide identification and testing of superior reference genes for transcript normalization in Arabidopsis. Plant Physiol. 2005;139: 5–17. doi: 10.1104/pp.105.063743 16166256

38. Die JV, Rowland LJ. Superior cross-species reference genes: a blueberry case study. PLoS ONE. 2013;8: e73354. doi: 10.1371/journal.pone.0073354 24058469

39. Vandesompele J, De Preter K, Pattyn F, Poppe B, Van Roy N, De Paepe A, et al. Accurate normalization of real-time quantitative RT-PCR data by geometric averaging of multiple internal control genes. Genome Biol. 2002;3: research0034. doi: 10.1186/gb-2002-3-7-research0034 12184808

40. Hellemans J, Mortier G, De Paepe A, Speleman F, Vandesompele J. qBase relative quantification framework and software for management and automated analysis of real-time quantitative PCR data. Genome Biol. 2007;8: R19. doi: 10.1186/gb-2007-8-2-r19 17291332

41. Consortium UniProt. UniProt: a worldwide hub of protein knowledge. Nucleic Acids Res. 2019;47: D506–D515. doi: 10.1093/nar/gky1049 30395287

42. Rubio J, Hajj-Moussa E, Kharrat M, Moreno MT, Millan T, Gil J. Two genes and linked RAPD markers involved in resistance to Fusarium oxysporum f. sp. ciceris race 0 in chickpea. Plant Breeding. 2003;122: 188–191. doi: 10.1046/j.1439-0523.2003.00814.x

43. Cobos MJ, Fernández MJ, Rubio J, Kharrat M, Moreno MT, Gil J, et al. A linkage map of chickpea (Cicer arietinum L.) based on populations from Kabuli x Desi crosses: location of genes for resistance to fusarium wilt race 0. Theor Appl Genet. 2005;110: 1347–1353. doi: 10.1007/s00122-005-1980-1 15806343

44. Iruela M, Castro P, Rubio J, Cubero JI, Jacinto C, Millán T, et al. Validation of a QTL for resistance to ascochyta blight linked to resistance to fusarium wilt race 5 in chickpea (Cicer arietinum L.). Eur J Plant Pathol. 2007;119: 29–37. doi: 10.1007/s10658-007-9121-0

45. Cobos MJ, Winter P, Kharrat M, Cubero JI, Gil J, Millan T, et al. Genetic analysis of agronomic traits in a wide cross of chickpea. Field Crops Res. 2009;111: 130–136. doi: 10.1016/j.fcr.2008.11.006

46. Halila I, Rubio J, Millán T, Gil J, Kharrat M, Marrakchi M. Resistance in chickpea (Cicer arietinum) to Fusarium wilt race ‘0’. Plant Breeding. 2009; doi: 10.1111/j.1439-0523.2009.01703.x

47. Palomino C, Fernández-Romero MD, Rubio J, Torres A, Moreno MT, Millán T. Integration of new CAPS and dCAPS-RGA markers into a composite chickpea genetic map and their association with disease resistance. Theor Appl Genet. 2009;118: 671–682. doi: 10.1007/s00122-008-0928-7 19034411

48. Bustin SA, Benes V, Garson JA, Hellemans J, Huggett J, Kubista M, et al. The MIQE guidelines: minimum information for publication of quantitative real-time PCR experiments. Clin Chem. 2009;55: 611–622. doi: 10.1373/clinchem.2008.112797 19246619

49. Wang L, Yin X, Cheng C, Wang H, Guo R, Xu X, et al. Evolutionary and expression analysis of a MADS-box gene superfamily involved in ovule development of seeded and seedless grapevines. Mol Genet Genomics. 2015;290: 825–846. doi: 10.1007/s00438-014-0961-y 25429734

50. Theißen G, Gramzow L. Structure and evolution of plant MADS domain transcription factors. Plant Transcription Factors. Elsevier; 2016. pp. 127–138.

51. Smaczniak C, Immink RGH, Angenent GC, Kaufmann K. Developmental and evolutionary diversity of plant MADS-domain factors: insights from recent studies. Development. 2012;139: 3081–3098. doi: 10.1242/dev.074674 22872082

52. Saha G, Park J-I, Jung H-J, Ahmed NU, Kayum MA, Chung M-Y, et al. Genome-wide identification and characterization of MADS-box family genes related to organ development and stress resistance in Brassica rapa. BMC Genomics. 2015;16: 178. doi: 10.1186/s12864-015-1349-z 25881193

53. Lee B, Henderson DA, Zhu J-K. The Arabidopsis cold-responsive transcriptome and its regulation by ICE1. Plant Cell. 2005;17: 3155–3175. doi: 10.1105/tpc.105.035568 16214899

54. Wei B, Cai T, Zhang R, Li A, Huo N, Li S, et al. Novel microRNAs uncovered by deep sequencing of small RNA transcriptomes in bread wheat (Triticum aestivum L.) and Brachypodium distachyon (L.) Beauv. Funct Integr Genomics. 2009;9: 499–511. doi: 10.1007/s10142-009-0128-9 19499258

55. Lee I, Seo Y-S, Coltrane D, Hwang S, Oh T, Marcotte EM, et al. Genetic dissection of the biotic stress response using a genome-scale gene network for rice. Proc Natl Acad Sci USA. 2011;108: 18548–18553. doi: 10.1073/pnas.1110384108 22042862

56. Zhao Y, Li X, Chen W, Peng X, Cheng X, Zhu S, et al. Whole-genome survey and characterization of MADS-box gene family in maize and sorghum. Plant Cell Tiss Organ Cult. 2011;105: 159–173. doi: 10.1007/s11240-010-9848-8

57. Khong GN, Pati PK, Richaud F, Parizot B, Bidzinski P, Mai CD, et al. Osmads26 negatively regulates resistance to pathogens and drought tolerance in rice. Plant Physiol. 2015;169: 2935–2949. doi: 10.1104/pp.15.01192 26424158

58. Yin W, Hu Z, Hu J, Zhu Z, Yu X, Cui B, et al. Tomato (Solanum lycopersicum) MADS-box transcription factor SlMBP8 regulates drought, salt tolerance and stress-related genes. Plant Growth Regul. 2017;83: 55–68. doi: 10.1007/s10725-017-0283-2

59. Liu W, Han X, Zhan G, Zhao Z, Feng Y, Wu C. A Novel Sucrose-Regulatory MADS-Box Transcription Factor GmNMHC5 Promotes Root Development and Nodulation in Soybean (Glycine max [L.] Merr.). Int J Mol Sci. 2015;16: 20657–20673. doi: 10.3390/ijms160920657 26404246

60. Sun C-H, Yu J-Q, Wen L-Z, Guo Y-H, Sun X, Hao Y-J, et al. Chrysanthemum MADS-box transcription factor CmANR1 modulates lateral root development via homo-/heterodimerization to influence auxin accumulation in Arabidopsis. Plant Sci. 2018;266: 27–36. doi: 10.1016/j.plantsci.2017.09.017 29241564

61. Tanaka Y, Iwaki S, Tsukazaki T. Crystal structure of a plant multidrug and toxic compound extrusion family protein. Structure. 2017;25: 1455–1460.e2. doi: 10.1016/j.str.2017.07.009 28877507

62. Saier MH, Paulsen IT. Phylogeny of multidrug transporters. Semin Cell Dev Biol. 2001;12: 205–213. doi: 10.1006/scdb.2000.0246 11428913

63. Moriyama Y, Hiasa M, Matsumoto T, Omote H. Multidrug and toxic compound extrusion (MATE)-type proteins as anchor transporters for the excretion of metabolic waste products and xenobiotics. Xenobiotica. 2008;38: 1107–1118. doi: 10.1080/00498250701883753 18668441

64. Min X, Jin X, Liu W, Wei X, Zhang Z, Ndayambaza B, et al. Transcriptome-wide characterization and functional analysis of MATE transporters in response to aluminum toxicity in Medicago sativa L. PeerJ. 2019;7: e6302. doi: 10.7717/peerj.6302 30723620

65. Diener AC, Gaxiola RA, Fink GR. Arabidopsis ALF5, a multidrug efflux transporter gene family member, confers resistance to toxins. Plant Cell. 2001;13: 1625–1638. doi: 10.1105/TPC.010035 11449055

66. Nawrath C. EDS5, an Essential Component of Salicylic Acid-Dependent Signaling for Disease Resistance in Arabidopsis, Is a Member of the MATE Transporter Family. THE PLANT CELL ONLINE. 2002;14: 275–286. doi: 10.1105/tpc.010376 11826312

67. Tiwari M, Sharma D, Singh M, Tripathi RD, Trivedi PK. Expression of OsMATE1 and OsMATE2 alters development, stress responses and pathogen susceptibility in Arabidopsis. Sci Rep. 2014;4: 3964. doi: 10.1038/srep03964 24492654

68. Wang J, Hou Q, Li P, Yang L, Sun X, Benedito VA, et al. Diverse functions of multidrug and toxin extrusion (MATE) transporters in citric acid efflux and metal homeostasis in Medicago truncatula. Plant J. 2017;90: 79–95. doi: 10.1111/tpj.13471 28052433

69. Wu X-Y, Zhou G-C, Chen Y-X, Wu P, Liu L-W, Ma F-F, et al. Soybean cyst nematode resistance emerged via artificial selection of duplicated serine hydroxymethyltransferase genes. Front Plant Sci. 2016;7: 998. doi: 10.3389/fpls.2016.00998 27458476

70. Moreno JI, Martín R, Castresana C. Arabidopsis SHMT1, a serine hydroxymethyltransferase that functions in the photorespiratory pathway influences resistance to biotic and abiotic stress. Plant J. 2005;41: 451–463. doi: 10.1111/j.1365-313X.2004.02311.x 15659103

71. Voll LM, Jamai A, Renné P, Voll H, McClung CR, Weber APM. The photorespiratory Arabidopsis shm1 mutant is deficient in SHM1. Plant Physiol. 2006;140: 59–66. doi: 10.1104/pp.105.071399 16339799

72. Zhang Y, Sun K, Sandoval FJ, Santiago K, Roje S. One-carbon metabolism in plants: characterization of a plastid serine hydroxymethyltransferase. Biochem J. 2010;430: 97–105. doi: 10.1042/BJ20100566 20518745

73. Engel N, Ewald R, Gupta KJ, Zrenner R, Hagemann M, Bauwe H. The presequence of Arabidopsis serine hydroxymethyltransferase SHM2 selectively prevents import into mesophyll mitochondria. Plant Physiol. 2011;157: 1711–1720. doi: 10.1104/pp.111.184564 21976482

74. Wei Z, Sun K, Sandoval FJ, Cross JM, Gordon C, Kang C, et al. Folate polyglutamylation eliminates dependence of activity on enzyme concentration in mitochondrial serine hydroxymethyltransferases from Arabidopsis thaliana. Arch Biochem Biophys. 2013;536: 87–96. doi: 10.1016/ 23800877

75. Liu S, Kandoth PK, Warren SD, Yeckel G, Heinz R, Alden J, et al. A soybean cyst nematode resistance gene points to a new mechanism of plant resistance to pathogens. Nature. 2012;492: 256–260. doi: 10.1038/nature11651 23235880

76. Rutherford SL. Between genotype and phenotype: protein chaperones and evolvability. Nat Rev Genet. 2003;4: 263–274. doi: 10.1038/nrg1041 12671657

77. Phukan UJ, Jeena GS, Shukla RK. WRKY transcription factors: molecular regulation and stress responses in plants. Front Plant Sci. 2016;7: 760. doi: 10.3389/fpls.2016.00760 27375634

78. Song H, Sun W, Yang G, Sun J. WRKY transcription factors in legumes. BMC Plant Biol. 2018;18: 243. doi: 10.1186/s12870-018-1467-2 30332991

79. Zuriaga E, Romero C, Blanca JM, Badenes ML. Resistance to Plum Pox Virus (PPV) in apricot (Prunus armeniaca L.) is associated with down-regulation of two MATHd genes. BMC Plant Biol. 2018;18: 25. doi: 10.1186/s12870-018-1237-1 29374454

Článek vyšel v časopise


2019 Číslo 10