Plant Physiology and Biochemistry 109(2016)561-570 Contents lists available at ScienceDirect PPB Plant Physiology and Biochemistry ELSEVIER journal homepage:www.elsevier.com/locate/plaphy Research article Comparative transcriptome profiling of chilling stress responsiveness CrossMark in grafted watermelon seedlings Jinhua Xu .5,Man Zhang Guang Liu b,Xingping Yang B,Xilin Hou" State Key Laboratory of Crop Genetics and Germplasm Enhancement.College of Horticulture.Nanjing Agricultural University.Nanjing 210095.Jiangsu. China Institute of Vegetable.Jiangsu Academy of Agricultural Sciences/Jiangsu Key Laboratory for Horticultural Crop Genetic Improvement,Nanjing 210014. Jiangsu,China ARTICLE INFO ABSTRACT Article history: Rootstock grafting may improve the resistance of watermelon plants to low temperatures.However Received 6 April 2016 information regarding the molecular responses of rootstock grafted plants to chilling stress is limited.To Received in revised form elucidate the molecular mechanisms of chilling tolerance in grafted plants,the transcriptomic responses 3 November 2016 Accepted 3 November 2016 of grafted watermelon under chilling stress were analyzed using RNA-seq analysis.Sequencing data were Available online 5 November 2016 used for digital gene expression(DGE)analysis to characterize the transcriptomic responses in grafted watermelon seedlings.A total of 702 differentially-expressed genes(DEGs)were found in rootstock grafted(RG)watermelon relative to self-grafted(SG)watermelon:among these genes,522 genes were Keywords: Watermelon up-regulated and 180 were down-regulated.Additionally.164 and 953 genes were found to specifically Rootstock grafting expressed in RG and SG seedlings under chilling stress,respectively.Functional annotations revealed that Differentially-expressed gene(DEG) up-regulated DEGs are involved in protein processing.plant-pathogen interaction and the spliceosome. Chilling stress whereas down-regulated DEGs are associated with photosynthesis.Moreover,13 DEGs were randomly selected for quantitative real time PCR(gRT-PCR)analysis.The expression profiles of these 13 DEGs were consistent with those detected by the DGE analysis,supporting the reliability of the DGE data.This work provides additional insight into the molecular basis of grafted watermelon responses to chilling stress. 2016 Published by Elsevier Masson SAS 1.Introduction 2010).The root system activity and SOD activity of grafted cu- cumber seedlings were found to be higher than those of ungrafted Low temperature is one of the major environmental factors that cucumbers under chilling stress (Li et al.,2008).Zhou et al.(2009) severely limits plant growth and development,especially for the reported that figleaf gourd grafting significantly alleviated cu- chilling sensitive cultivated watermelon (C.lanatus).Watermelon cumber seedling growth inhibition by chilling at 7 C while also grows best at temperatures ranging from 21 to 29C,with growth increasing light utilization and reducing the accumulation of ceasing at 10C death occurring at temperatures of 1 C(Noh et al., reactive oxygen species after chilling.Inhibition of the light- 2013).Because of its low-temperature sensitivity,it is very difficult saturated rate of CO2 assimilation,the maximum carboxylation to obtain good yields and fruit quality during the cold seasons.To activity,Rubisco content and initial Rubisco activity were all found avoid such difficulties and improve the growth performance. to be weaker in grafted cucumber plants after chilling at 7C(Zhou watermelon seedlings are usually grafted onto rootstocks to confer et al.,2007).The relative growth rate of shoots and root mass ratios resistance to low temperatures (Lee and Oda,2003) increased at 15C when tomato seedlings were grafted onto a cold- Grafting has been attempted in several crops to increase plant tolerant rootstock(Venema et al.,2008).Grafted watermelon(Ding tolerance to low temperatures.Rootstocks alleviated the negative et al,2011)and muskmelon (Justus and Kubota,2010)seedlings effects of low temperatures on scion performance by supplying the were found to have better storability under low-temperature scion with more water,nutrients and hormones (Schwarz et al., storage than non-grafted seedlings.These studies have helped us to understand the physiological basis of increased low-temperature tolerance of rootstock-grafted plants.In addition to the physiolog- Corresponding author. ical responses,novel proteins were identified in pumpkin rootstock E-mail address:hxl@njau.edu.cn (X.Hou). grafted cucumber plants using proteomics techniques,and these http://dx.doi.org/10.1016/j.plaphy.2016.11.002 0981-9428/2016 Published by Elsevier Masson SAS.Research article Comparative transcriptome profiling of chilling stress responsiveness in grafted watermelon seedlings Jinhua Xu a, b , Man Zhang b , Guang Liu b , Xingping Yang b , Xilin Hou a, * a State Key Laboratory of Crop Genetics and Germplasm Enhancement, College of Horticulture, Nanjing Agricultural University, Nanjing 210095, Jiangsu, China b Institute of Vegetable, Jiangsu Academy of Agricultural Sciences/Jiangsu Key Laboratory for Horticultural Crop Genetic Improvement, Nanjing 210014, Jiangsu, China article info Article history: Received 6 April 2016 Received in revised form 3 November 2016 Accepted 3 November 2016 Available online 5 November 2016 Keywords: Watermelon Rootstock grafting Differentially-expressed gene (DEG) Chilling stress abstract Rootstock grafting may improve the resistance of watermelon plants to low temperatures. However, information regarding the molecular responses of rootstock grafted plants to chilling stress is limited. To elucidate the molecular mechanisms of chilling tolerance in grafted plants, the transcriptomic responses of grafted watermelon under chilling stress were analyzed using RNA-seq analysis. Sequencing data were used for digital gene expression (DGE) analysis to characterize the transcriptomic responses in grafted watermelon seedlings. A total of 702 differentially-expressed genes (DEGs) were found in rootstock grafted (RG) watermelon relative to self-grafted (SG) watermelon; among these genes, 522 genes were up-regulated and 180 were down-regulated. Additionally, 164 and 953 genes were found to specifically expressed in RG and SG seedlings under chilling stress, respectively. Functional annotations revealed that up-regulated DEGs are involved in protein processing, plant-pathogen interaction and the spliceosome, whereas down-regulated DEGs are associated with photosynthesis. Moreover, 13 DEGs were randomly selected for quantitative real time PCR (qRT-PCR) analysis. The expression profiles of these 13 DEGs were consistent with those detected by the DGE analysis, supporting the reliability of the DGE data. This work provides additional insight into the molecular basis of grafted watermelon responses to chilling stress. © 2016 Published by Elsevier Masson SAS. 1. Introduction Low temperature is one of the major environmental factors that severely limits plant growth and development, especially for the chilling sensitive cultivated watermelon (C. lanatus). Watermelon grows best at temperatures ranging from 21 to 29
C, with growth ceasing at 10
C death occurring at temperatures of 1
C (Noh et al., 2013). Because of its low-temperature sensitivity, it is very difficult to obtain good yields and fruit quality during the cold seasons. To avoid such difficulties and improve the growth performance, watermelon seedlings are usually grafted onto rootstocks to confer resistance to low temperatures (Lee and Oda, 2003). Grafting has been attempted in several crops to increase plant tolerance to low temperatures. Rootstocks alleviated the negative effects of low temperatures on scion performance by supplying the scion with more water, nutrients and hormones (Schwarz et al., 2010). The root system activity and SOD activity of grafted cu￾cumber seedlings were found to be higher than those of ungrafted cucumbers under chilling stress (Li et al., 2008). Zhou et al. (2009) reported that figleaf gourd grafting significantly alleviated cu￾cumber seedling growth inhibition by chilling at 7
C while also increasing light utilization and reducing the accumulation of reactive oxygen species after chilling. Inhibition of the light￾saturated rate of CO2 assimilation, the maximum carboxylation activity, Rubisco content and initial Rubisco activity were all found to be weaker in grafted cucumber plants after chilling at 7
C (Zhou et al., 2007). The relative growth rate of shoots and root mass ratios increased at 15
C when tomato seedlings were grafted onto a cold￾tolerant rootstock (Venema et al., 2008). Grafted watermelon (Ding et al., 2011) and muskmelon (Justus and Kubota, 2010) seedlings were found to have better storability under low-temperature storage than non-grafted seedlings. These studies have helped us to understand the physiological basis of increased low-temperature tolerance of rootstock-grafted plants. In addition to the physiolog￾ical responses, novel proteins were identified in pumpkin rootstock grafted cucumber plants using proteomics techniques, and these * Corresponding author. E-mail address: hxl@njau.edu.cn (X. Hou). Contents lists available at ScienceDirect Plant Physiology and Biochemistry journal homepage: www.elsevier.com/locate/plaphy http://dx.doi.org/10.1016/j.plaphy.2016.11.002 0981-9428/© 2016 Published by Elsevier Masson SAS. Plant Physiology and Biochemistry 109 (2016) 561e570