Identification of lead-regulated genes by suppression subtractive hybridization in the heavy metal accumulator Sesbania drummondii
Heavy metal contamination of soils is of widespread occurrence as a result of human, agricultural and industrial activities. Among heavy metals, lead is a potential pollutant that readily accumulates in soils and sediments. Although lead is not an essential element for plants, it gets easily absorbed and accumulated in Sesbania drummondii, which exhibits a significant level of tolerance to lead. The response of a metal tolerant plant to heavy metal stress involves a number of biochemical and physiological pathways. To investigate the overall molecular response of a metal-tolerant plant to lead exposure, suppression subtractive hybridization (SSH) was used to construct a cDNA library enriched in lead induced mRNA transcripts from lead-tolerant Sesbania. Screening the library by reverse Northern analysis revealed that between 20 and 25% of clones selected from the library were differentially regulated in lead treated plants. After differential screening, we isolated several differentially expressed cDNA clones, including a type 2 metallothionein (MT) gene which is involved in detoxification and homeostasis and shown to be differentially regulated in lead treated plants. The data from the reverse Northern analysis was further confirmed with conventional Northern analysis of a select group of genes including MT, ACC synthase/oxidase, cold-, water stress-, and other abiotic stress-induced genes, which are up-regulated rapidly in response to lead treatment. The mRNA levels of MT increased substantially after lead treatment indicating a potential role for it under lead stress in Sesbania. The present results show that SSH can serve as an effective tool for isolating genes induced in response to lead heavy metal tolerance in Sesbania. A better understanding of lead induced gene expression in Sesbania should help select candidates associated with remediation of heavy metal toxicity. The possible link between this result and the heavy-metal response of plants is discussed.
KeywordsDifferential gene expression Sesbania drummondii Lead Suppression subtractive hybridization Heavy metal stress
Suppression subtractive hybridization
Reactive oxygen species
Expressed sequence tags
Polymerase chain reaction
Financial supports from the Sponsored Programs, the Graduate School, and Biotechnology Center of Applied Research and Technology Program of Ogden College of Science and Engineering at Western Kentucky University are greatly acknowledged.
- Garbisu C, Hernandex-Allica J, Barrutia O, Alkorta I, Becerril JM (2002) Phytoremediation: a technology using green plants to remove contaminants from polluted areas. Rev Environ Health 17:75–90Google Scholar
- Langebartels C, Schraudner M, Heller W, Ernst D, Sandermann H (2000) Oxidative stress and defense reactions in plants exposed to air pollutants and UV-B radiation. In: Inze D, Van Montagu M (eds) Oxidative stress in plants. Harwood Academic Publishers, Amsterdam, pp 105–135Google Scholar
- Narusaka Y, Narusaka M, Seki M, Umezawa T, Ishida J, Nakajima M, Enju A, Shinozaki K (2004) Crosstalk in the response to abiotic and biotic stresses in Arabidopsis: analysis of gene expression in cytochrome P450 gene superfamily by cDNA microarray. Plant Mol Biol 55:327–342PubMedCrossRefGoogle Scholar
- Suh MC, Choi D, Liu JR (1998) Cadmium resistance in transgenic tobacco plants expressing the Nicotiana glutinosa L. metallothionein-like gene. Mol Cell 8:678–684Google Scholar
- Weber M, Trampczynska A, Clemens S (2005) Comparative transcriptome analysis of toxic metal responses in Arabidopsis thaliana and the Cd2+-hypertolerant facultative metallophyte Arabidopsis halleri. Plant Cell Eviron 1–14Google Scholar