A novel WRKY transcriptional factor from Thlaspi caerulescens negatively regulates the osmotic stress tolerance of transgenic tobacco

A novel member of the WRKY gene family, designated TcWRKY53, was isolated from a cadmium (Cd)-treated Thlaspi caerulescens cDNA library by differential screening. WRKY proteins specifically bind to W-boxes, which are found in the promoters of many genes involved in defense and response to environmental stress. TcWRKY53 contains a 975-bp open reading frame encoding a putative protein of 324 amino acids. Homology searches showed that TcWRKY53 resembles similar WRKY domain-containing proteins from rice, parsley and tobacco, especially AtWRKY53 from Arabidopsis thaliana. Semi-quantitative RT-PCR showed that the expression of TcWRKY53 was strongly induced by various environmental stresses, including an excess of NaCl, drought, cold and the signal molecule salicylic acid (SA). The expression of TcWRKY53 in response to NaCl, drought and cold suggested a possible role of TcWRKY53 in abiotic stress response. However, physiological tests indicated that the expression of TcWRKY53 in tobaccos decreases tolerance to sorbitol during seedling root development. This was consistent with PEG6000 treatment of tobacco seedlings, and together these results indicate a negative modulation of TcWRKY53 in response to osmotic stress. Furthermore, two ethylene responsive factor (ERF) family genes, NtERF5 and NtEREBP-1, were negatively induced in TcWRKY53-overexpressing transgenic plants. In contrast, a LEA family gene, NtLEA5, showed no change, suggesting that TcWRKY53 might regulate the plant osmotic stress response by interacting with an ERF-type transcription factor rather than by regulating function genes directly.