Omics approaches for identification of stress responsive genes
Author: Dr. Amolkumar U. Solanke and Dr. SV Amitha Mithra
NRC on Plant Biotechnology, LBS Building, IARI, Pusa Campus, New Delhi
Comparative study of transcripts, proteins and metabolites in stressed and un-stressed crop plants help to pick important stress associated genes, products which can be useful in providing various levels of tolerance. These three omics approaches have a fundamental difference from genomics in that these are dynamic in nature where as genome is static across the tissues and lifetime of an individual.
Microarray analysis, EST (Expressed Sequence Tag) collection, SAGE (Serial Analysis of Gene Expression) and NGS (Next Generation Sequencing) profiling are the methods to analyse genes expressed in stressed conditions. The ready to use microarray chips are available for many plants like Arabidopsis, rice, tomato, cotton which can be directly used for analysis. For transcriptome profiling of other plants, customized chips can be designed. Many studies are available in specific stresses using these chips. Comparative transcriptome changes in response to salt, osmotic, and cold stress had been studied using microarray in Arabidopsis (Kreps et al., 2002). Unlike microarray, EST collection can be used in any crop in which previous sequencing information is not available. A large amount of EST data is available for many crop species on NCBI and Plant Genome Index databases. However, the numbers of stress related libraries in these EST databases are very few. Recently, transcript profiling for salt tolerant Festuca rubra ssp. litoralis has been done to reveal regulatory network controlling salt acclimatization (Diedhiou et al., 2008). Transcript profiling of water-limited roots of hexaploid wheat (Mohammadi et al., 2008), drought exposed maize and chickpea (Marino et al., 2008; Gao et al., 2008), chilling and salt stress exposed sunflower (Fernandez et al., 2008) and salt tolerant model halophyte, Thellungiella halophila (Taji et al., 2004; 2008) has been carried out. SAGE is also emerging as technique to find out genes related to stress condition though it requires whole genome information to analyse the obtained gene fragments. This technique has been applied to compare cold related genes in Arabidopsis (Robinson and Parkin, 2008) and to find out drought stress-responsive transcripts in roots in chickpea (Molina et al., 2008). Recent advancement in sequencing technology i.e. Next generation Sequencing technologies can be directly used to identify genes expressed in stress condition by comparing it with control counterpart. Advantage of this technique is that it can identify novel genes.
The protein profiling or proteomics is another approach which has been used for study of related changes during stress. Analysis of proteins from control and stressed samples on 2D gel electrophoresis or through mass-spectrometry can provide information about new proteins as well as increased or decreased amount of proteins in stress condition. Proteome analysis in rice seedlings for cold stress (Cui et al., 2005) and in wheat for drought stress (Hajheidari et al., 2007) provides information about proteins specific to these stresses. Proteome analysis of sugar beet (Hajheidari et al., 2005) and chickpea (Bhushan et al., 2007) exposed to dehydration stress has been carried out. Similar work have been done to identify effect of salt stress on Suaeda aegyptiaca (Askari et al., 2006) and Physcomitrella patens (Wang et al., 2008). Proteomics of poplar exposed to chilling stress has also been done by Renaut et al. (2004). Reports are also available for root proteomic responses of two Agrostis species contrasting in heat tolerance (Xu and Huang, 2008).
Recently, researchers have tried to find out metabolites protecting plants in stress conditions, along with genes and proteins responsible for their generation. This type of analysis is referred as metabolite profiling or metabolome analysis. Generally, sophisticated mass spectrometry instruments like, gas chromatography mass spectrometry (GC-MS), liquid chromatography mass spectrometry (LC-MS), gas chromatography-time-of-flight mass spectrometry (GC-TOF-MS), capillary electrophoresis mass spectrometry (CE-MS), high-performance liquid chromatography-ultraviolet-electrospray ionization mass spectrometry (HPLC-UV-ESI-MS), fourier-transformed infrared spectroscopy or NMR-based methods and statistical methods like principal component analysis (PCA) and batch-learning self-organizing mapping analysis (BL-SOM) are used to understand metabolome profiles. This is a new aspect and therefore few reports related to stress metabolites are available. In 2004, Kaplan et al. explored the temperature stress metabolome of Arabidopsis, and found that cold shock influenced metabolome is far more profound than heat shock. Analysis of transcript and metabolite profiles of cold acclimated Arabidopsis by same group revealed an intricate relationship of cold-regulated gene expression with changes in metabolite content (Kaplan et al., 2007). These studies also show the prominent role of carbohydrate metabolism, which seems to be a major feature of the reprogramming of the metabolome during temperature stress, especially trehalose biosynthesis (Guy et al., 2008; Iordachescu and Imai, 2008). Metabolome studies on drought stress showed the role of ABA in regulating metabolic network (Seki et al., 2007; Urano et al., 2008). Metabolome studies of salt stressed Arabidopsis, lotus and rice revealed some conserved and some divergent metabolic responses in these plants (Sanchez et al., 2008). A time course of metabolic profiling of Arabidopsis cell culture suggests that methylation cycle for the supply of methyl groups, the phenylpropanoid pathway for lignin production and glycinebetaine biosynthesis are the short term responses whereas co-induction of glycolysis and sucrose metabolism and co-reduction of the methylation cycle are long term responses to salt stress (Kim et al., 2007). The role of secondary metabolites like genistin and group B saponins for salt tolerance has been revealed by comparative metabolic profiling of soybean in salt stress (Wu et al., 2008).
Thus transcriptomics, proteomics and metabolomics studies with system biology approach are expected to provide holistic view of stress responses and identification of few key genes which help plants to mitigate stress conditions.
Kreps, J.A., Wu, Y., Chang, H.S., Zhu, T., Wang, X., and Harper, J.F. (2002). Transcriptome changes for Arabidopsis in response to salt, osmotic, and cold stress. Plant Physiol 130, 2129-2141.
Diedhiou, C.J., Popova, O.V., and Golldack, D. (2009). Transcript profiling of the salt-tolerant Festuca rubra ssp. litoralis reveals a regulatory network controlling salt acclimatization. J Plant Physiol 166, 697-711.
Mohammadi, M., Kav, N.N., and Deyholos, M.K. (2008). Transcript expression profile of water-limited roots of hexaploid wheat (Triticum aestivum 'Opata'). Genome 51, 357-367.
Molina, C., Rotter, B., Horres, R., Udupa, S.M., Besser, B., Bellarmino, L., Baum, M., Matsumura, H., Terauchi, R., Kahl, G., and Winter, P. (2008). SuperSAGE: the drought stress-responsive transcriptome of chickpea roots. BMC Genomics 9, 553.
Marino, R., Ponnaiah, M., Krajewski, P., Frova, C., Gianfranceschi, L., Pe, M.E., and Sari-Gorla, M. (2009). Addressing drought tolerance in maize by transcriptional profiling and mapping. Mol Genet Genomics 281, 163-179.
Gao, W.R., Wang, X.S., Liu, Q.Y., Peng, H., Chen, C., Li, J.G., Zhang, J.S., Hu, S.N., and Ma, H. (2008). Comparative analysis of ESTs in response to drought stress in chickpea (C. arietinum L.). Biochem Biophys Res Commun 376, 578-583.
Fernandez, P., Di Rienzo, J., Fernandez, L., Hopp, H.E., Paniego, N., and Heinz, R.A. (2008). Transcriptomic identification of candidate genes involved in sunflower responses to chilling and salt stresses based on cDNA microarray analysis. BMC Plant Biol 8, 11.
Taji, T., Seki, M., Satou, M., Sakurai, T., Kobayashi, M., Ishiyama, K., Narusaka, Y., Narusaka, M., Zhu, J.K., and Shinozaki, K. (2004). Comparative genomics in salt tolerance between Arabidopsis and Arabidopsis-related halophyte salt cress using Arabidopsis microarray. Plant Physiol 135, 1697-1709.
Taji, T., Sakurai, T., Mochida, K., Ishiwata, A., Kurotani, A., Totoki, Y., Toyoda, A., Sakaki, Y., Seki, M., Ono, H., Sakata, Y., Tanaka, S., and Shinozaki, K. (2008). Large-scale collection and annotation of full-length enriched cDNAs from a model halophyte, Thellungiella halophila. BMC Plant Biol 8, 115.
Robinson, S.J., and Parkin, I.A. (2008). Differential SAGE analysis in Arabidopsis uncovers increased transcriptome complexity in response to low temperature. BMC Genomics 9, 434.
Cui, S., Huang, F., Wang, J., Ma, X., Cheng, Y., and Liu, J. (2005). A proteomic analysis of cold stress responses in rice seedlings. Proteomics 5, 3162-3172.
Hajheidari, M., Abdollahian-Noghabi, M., Askari, H., Heidari, M., Sadeghian, S.Y., Ober, E.S., and Salekdeh, G.H. (2005). Proteome analysis of sugar beet leaves under drought stress. Proteomics 5, 950-960.
Hajheidari, M., Eivazi, A., Buchanan, B.B., Wong, J.H., Majidi, I., and Salekdeh, G.H. (2007). Proteomics uncovers a role for redox in drought tolerance in wheat. J Proteome Res 6, 1451-1460.
Bhushan, D., Pandey, A., Choudhary, M.K., Datta, A., Chakraborty, S., and Chakraborty, N. (2007). Comparative proteomics analysis of differentially expressed proteins in chickpea extracellular matrix during dehydration stress. Mol Cell Proteomics 6, 1868-1884.
Askari, H., Edqvist, J., Hajheidari, M., Kafi, M., and Salekdeh, G.H. (2006). Effects of salinity levels on proteome of Suaeda aegyptiaca leaves. Proteomics 6, 2542-2554.
Wang, X., Yang, P., Gao, Q., Liu, X., Kuang, T., Shen, S., and He, Y. (2008). Proteomic analysis of the response to high-salinity stress in Physcomitrella patens. Planta 228, 167-177.
Renaut, J., Lutts, S., Hoffmann, L., and Hausman, J.F. (2004). Responses of poplar to chilling temperatures: proteomic and physiological aspects. Plant Biol (Stuttg) 6, 81-90.
Xu, C., and Huang, B. (2008). Root proteomic responses to heat stress in two Agrostis grass species contrasting in heat tolerance. J Exp Bot 59, 4183-4194.
Kaplan, F., Kopka, J., Haskell, D.W., Zhao, W., Schiller, K.C., Gatzke, N., Sung, D.Y., and Guy, C.L. (2004). Exploring the temperature-stress metabolome of Arabidopsis. Plant Physiol 136, 4159-4168.
Kaplan, F., Kopka, J., Sung, D.Y., Zhao, W., Popp, M., Porat, R., and Guy, C.L. (2007). Transcript and metabolite profiling during cold acclimation of Arabidopsis reveals an intricate relationship of cold-regulated gene expression with modifications in metabolite content. Plant J 50, 967-981.
Guy, C., Kaplan, F., Kopka, J., Selbig, J., and Hincha, D.K. (2008). Metabolomics of temperature stress. Physiol Plant 132, 220-235.
Iordachescu, M., and Imai, R. (2008). Trehalose biosynthesis in response to abiotic stresses. J Integr Plant Biol 50, 1223-1229.
Seki, M., Umezawa, T., Urano, K., and Shinozaki, K. (2007). Regulatory metabolic networks in drought stress responses. Curr Opin Plant Biol 10, 296-302.
Urano, K., Maruyama, K., Ogata, Y., Morishita, Y., Takeda, M., Sakurai, N., Suzuki, H., Saito, K., Shibata, D., Kobayashi, M., Yamaguchi-Shinozaki, K., and Shinozaki, K. (2009). Characterization of the ABA-regulated global responses to dehydration in Arabidopsis by metabolomics. Plant J 57, 1065-1078.
Sanchez, D.H., Siahpoosh, M.R., Roessner, U., Udvardi, M., and Kopka, J. (2008). Plant metabolomics reveals conserved and divergent metabolic responses to salinity. Physiol Plant 132, 209-219.
Kim, J.K., Bamba, T., Harada, K., Fukusaki, E., and Kobayashi, A. (2007). Time-course metabolic profiling in Arabidopsis thaliana cell cultures after salt stress treatment. J Exp Bot 58, 415-424.
Wu, W., Zhang, Q., Zhu, Y., Lam, H.M., Cai, Z., and Guo, D. (2008). Comparative metabolic profiling reveals secondary metabolites correlated with soybean salt tolerance. J Agric Food Chem 56, 11132-11138.
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