Publicação
Functional characterisation of HZF and LZF genes in heat stress response of Arabidopsis thaliana
| Resumo: | Superior plants are well adapted to the environmental perturbations they are inevitably exposed to. Heat stressful conditions are often responsible for the reallocation of the metabolic processes in tolerant plants. Understanding how plants perceive high temperatures and coordinate cellular growth in response to heat stress is essential to surpass the potential negative effects and significant crops yield reduction. HZF is gene encoding a C3H2C3-type RING (Really Interesting New Gene) zinc finger protein, proposed to be a heat response determinant in Arabidopsis thaliana. HZF loss-of-function causes a delay in the germination process under heat stress conditions, suggesting that this gene might be important for the extension of the dormancy period or the inhibition of the germination triggering. RING finger proteins are sometimes found in dimmers of redundant pairs of RING-type E3 ubiquitin ligases. A highly homologous gene to HZF (LZF), also encoding for a RING finger protein, has been proposed to have a redundant function to HZF in standard environmental conditions. In silico analysis of both HZF and LZF revealed that the expression of both genes might be influenced by abscisic acid (ABA), which suggests that HZF-LZF regulation might play a role in ABA signalling during germination. The present work gives new perspectives on how the acquisition of thermotolerance might depend on HZF expression. Following the bioinformatic analyses, which strongly suggested similar regulation profiles for HZF and LZF, the functional characterisation of loss-of-function and overexpression mutant lines for these genes was initiated. HZF and LZF appear to be both heat-responsive but only HZF seems to be important at the onset of seed germination under heat stress conditions. However, in what concerns to ABA hormonal regulation, both genes seem to play an important role. HZF and LZF gene expression analyses revealed positive feedback loops between one another, further supporting the coregulation hypothesis. The examination of transgenic plants containing the GUS reporter gene under the control of HZF native promoter has corroborated its involvement in heat responses and implicated HZF in ABA and auxin-dependent root development events. The subcellular localisation by fluorescent imaging of the HZF product suggests that it could be directed to structures that control the plant cell expansion. To better understand the mechanisms underlying HZF role in the processes that it could be involved, the HZF recombinant protein has been expressed in the E. coli heterologous systems. The recombinant HZF protein could be used in future biochemical assays, for assessing the putative the ubiquitin-ligase activity. Although far from being elucidated, the results obtained in this work contributed for a better understanding of HZF and LZF role on heat stress responses and other physiological processes. |
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| Autores principais: | Oliveira, Rute Patrícia Martins Barbosa de |
| Ano: | 2010 |
| País: | Portugal |
| Tipo de documento: | dissertação de mestrado |
| Tipo de acesso: | acesso restrito |
| Instituição associada: | Universidade do Minho |
| Idioma: | inglês |
| Origem: | RepositóriUM - Universidade do Minho |
| Resumo: | Superior plants are well adapted to the environmental perturbations they are inevitably exposed to. Heat stressful conditions are often responsible for the reallocation of the metabolic processes in tolerant plants. Understanding how plants perceive high temperatures and coordinate cellular growth in response to heat stress is essential to surpass the potential negative effects and significant crops yield reduction. HZF is gene encoding a C3H2C3-type RING (Really Interesting New Gene) zinc finger protein, proposed to be a heat response determinant in Arabidopsis thaliana. HZF loss-of-function causes a delay in the germination process under heat stress conditions, suggesting that this gene might be important for the extension of the dormancy period or the inhibition of the germination triggering. RING finger proteins are sometimes found in dimmers of redundant pairs of RING-type E3 ubiquitin ligases. A highly homologous gene to HZF (LZF), also encoding for a RING finger protein, has been proposed to have a redundant function to HZF in standard environmental conditions. In silico analysis of both HZF and LZF revealed that the expression of both genes might be influenced by abscisic acid (ABA), which suggests that HZF-LZF regulation might play a role in ABA signalling during germination. The present work gives new perspectives on how the acquisition of thermotolerance might depend on HZF expression. Following the bioinformatic analyses, which strongly suggested similar regulation profiles for HZF and LZF, the functional characterisation of loss-of-function and overexpression mutant lines for these genes was initiated. HZF and LZF appear to be both heat-responsive but only HZF seems to be important at the onset of seed germination under heat stress conditions. However, in what concerns to ABA hormonal regulation, both genes seem to play an important role. HZF and LZF gene expression analyses revealed positive feedback loops between one another, further supporting the coregulation hypothesis. The examination of transgenic plants containing the GUS reporter gene under the control of HZF native promoter has corroborated its involvement in heat responses and implicated HZF in ABA and auxin-dependent root development events. The subcellular localisation by fluorescent imaging of the HZF product suggests that it could be directed to structures that control the plant cell expansion. To better understand the mechanisms underlying HZF role in the processes that it could be involved, the HZF recombinant protein has been expressed in the E. coli heterologous systems. The recombinant HZF protein could be used in future biochemical assays, for assessing the putative the ubiquitin-ligase activity. Although far from being elucidated, the results obtained in this work contributed for a better understanding of HZF and LZF role on heat stress responses and other physiological processes. |
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