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Accueil du site → Doctorat → Allemagne → 2001 → Analysis of ABA and drought stress mediated gene expression in the desiccation tolerant resurrection plant Craterostigma plantagineum

Köln Universität (2001)

Analysis of ABA and drought stress mediated gene expression in the desiccation tolerant resurrection plant Craterostigma plantagineum

Smith-Espinoza, Claudia J.

Titre : Analysis of ABA and drought stress mediated gene expression in the desiccation tolerant resurrection plant Craterostigma plantagineum

Auteur : Smith-Espinoza, Claudia J.

Université de soutenance : Köln, Univ., Diss.

Grade : PhD thesis 2001

The aim of the present study has been to get a better understanding of the molecular mechanisms leading to desiccation tolerance using the desiccation tolerant resurrection plant Craterostigma plantagineum as a model. To achieve this, two methods have been pursued ; (1) a transcript profiling approach to determine specific patterns of gene expression that lead to stress tolerance and (2) a mutational approach to dissect signalling networks that modulate plant responses to environmental cues. Molecular responses to either dehydration or salt treatment were compared in C. plantagineum . Although desiccation tolerant, C. plantagineum was shown to be sensitive to sodium chloride, thus offering an ideal system for the detection of specific dehydration responsive mRNAs. The analysis consisted of comparing the temporal sequence of gene expression in response to dehydration and sodium chloride treatment. For this purpose, a collection of 258 dehydration responsive genes were studied at the RNA level using a macroarray technique. Classification of a large number of genes based on temporal transcript accumulation patterns following sodium chloride and dehydration treatments allowed insight into the genetic programmes employed by C. plantagineum during the acquisition of desiccation tolerance. Of the transcripts that showed differential accumulation in response to dehydration or sodium chloride treatment, 23% were upregulated in response to both dehydration and sodium chloride. 19% of the gene collection were up-regulated by dehydration, but down-regulated by sodium chloride. This class of transcripts represent good candidates for further studies of specific gene responses to dehydration. A gene member of an aquaporin gene family was found to group in this class, and represents a good candidate for further analysis. Closer inspection revealed specific transcript profiles for groups of genes in response to dehydration treatment. One gene family, the so-called LEA genes, was studied in detail. This family displayed a synchronised transcript accumulation pattern in response to dehydration that differed from the pattern observed in response to sodium chloride stress. Therefore, transcript profiles observed only in the dehydration response, that differ from a more general stress response such as sodium chloride treatment, can be considered as molecular finger prints for the acquisition of desiccation tolerance. Most desiccation responses are mediated by abscisic acid (ABA). In order to dissect ABA/dehydration mediated signalling pathways, dominant C. plantagineum mutants were created via a T-DNA activation tagging approach. The screening of mutants was based on the observation that wild type C. plantagineum callus tissues do not survive desiccation without ABA treatment. Through the insertion of transcriptional enhancers, ectopic expression of genes is induced. The application of this approach has permitted the isolation of five independent desiccation tolerant callus lines that do not require ABA pre-treatment. Two of the transgenic lines were selected for further characterisation, lines 10 and H. The T-DNA flanking regions were rescued and the corresponding cDNAs isolated. None of the cDNA sequences had significant homology to reported genes present in public databases. However, the cDNA isolated from mutant line 10 showed limited homology to a previously characterised C. plantagineum gene and a mouse gene encoding a hypothetical protein. Expression analysis revealed that desiccation responsive genes were highly accumulated in fully hydrated callus and leaf tissues from mutant line 10, suggesting that the tagged locus is involved in the ABA/dehydration signalling network.


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