Promoter regulation of the Craterostigma plantagineum Cp LEA-like 11-24 gene and functional characterization of its transcription factor (CpbZIP1)
Promoter regulation of the Craterostigma plantagineum Cp LEA-like 11-24 gene and functional characterization of its transcription factor (CpbZIP1)
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DissertationDate of Exam
18.08.2014Date of Publication
26.08.2014Advisor
Bartels, DorotheaCo-Referee
Kirch, Hans-HubertMetadata
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Abstract
The resurrection plant Craterostigma plantagineum, a member of Linderniaceae family, is distributed in areas with variable water availability in Southern Africa. Two closely related species, L. brevidens which is endemic to montan rain forest of Africa and L. subracemosa which is found in central and Southern Africa, are used for comparative analyses for unravelling the molecular mechanisms of desiccation tolerance. While C. plantagineum and L. brevidens are desiccation tolerant, L. subracemosa represents a desiccation sensitive species. Desiccation tolerance is a very complex trait. Many genes that are involved in desiccation tolerance and the proteins that are encoded by these genes have been characterized in C. plantagineum. The Cp LEA-like 11-24 is one of these stress inducible genes in C. plantagineum that has been extensively studied both at the transcriptional and posttranscriptional level. Studies have been performed on the promoter regulation and transcription factor binding along with comparative promoter analyses between two desiccation tolerant and a desiccation sensitive species. Since promoter regions are the main regulatory elements for enhancing the transcriptional efficiency, developing an efficient transient transformation system was required to analyze promoter functions in response to different abiotic stresses. In this study, an efficient Agrobacterium-mediated transient transformation method was developed in three closely related Linderniaceae species. Different parameters that are important for enhancing the transformation efficiency were considered. These parameters include leaf size, Silwet L-77 concentration, bacterial density and duration of co-cultivation. After optimization, the activities of the minimal stress inducible promoter fragments of the LEA-like 11-24 from three species were examined under ABA and osmotic stress using optimized Agrobacterium co-cultivation method.
The transformation efficiency was found to be 100% in all three species. In addition, the induction pattern of each promoter fragment under ABA and osmotic stress was comparable with that of stably transformed plants or particle bombardment, suggesting this as the most suitable method for quantitatively analyzing promoter activity in the three species. The optimized method was used for cross comparison of the LEA-like 11-24 promoter fragments among two desiccation tolerant and a sensitive species to understand the effect of trans-regulatory factors in activating the LEA-like 11-24 promoter fragments. The results demonstrated that both desiccation tolerant and sensitive species contain essential trans-regulatory factors for the activity of LEA-like 11-24 promoter fragments. Using a yeast-one-hybrid system, it was previously shown the Cp LEA-like 11-24 promoter fragment interacts with CpbZIP1 protein. The CpbZIP1 belongs to the S1-class of AtbZIP transcription factors from Arabidopsis and has been shown to have 47% protein sequence identity with AtbZIP53. Transcript expression analysis of CpbZIP1 in response to ABA, dehydration and salt stress demonstrated the stress responsiveness of this gene. S1-class members of AtbZIP from Arabidopsis have complex expression patterns in different stress responses. The CpbZIP1 from C. plantagineum also showed similar complex expression pattern in response to stress. The function of CpbZIP1 transcription is completely unknown in C. plantagineum. In a functional study of the CpbZIP1 transcription factor, transgenic Arabidopsis plants overexpressing CpbZIP1 exhibited dwarf phenotype, abnormality in flower structure and small siliques suggesting that CpbZIP1 interferes with the developmental processes. Physiological and molecular data from transgenic plants overexpressing CpbZIP1 revealed the contribution of CpbZIP1 in remobilization of carbon/nitrogen or maintaining homeostasis of energy, as obtained for the bZIP homologue in Arabidopsis (AtbZIP53). Time dependent expression of CpbZIP1 revealed that apart from the function of CpbZIP1 in abiotic stress, most likely CpbZIP1 is regulated by circadian clock rhythm. The results from this study suggests that CpbZIP1 is stress inducible and involved in multiple functions like growth, flower development, accumulation of proline, remobilizing carbon/nitrogen and maintaining homeostasis of energy in low energy conditions.
The transformation efficiency was found to be 100% in all three species. In addition, the induction pattern of each promoter fragment under ABA and osmotic stress was comparable with that of stably transformed plants or particle bombardment, suggesting this as the most suitable method for quantitatively analyzing promoter activity in the three species. The optimized method was used for cross comparison of the LEA-like 11-24 promoter fragments among two desiccation tolerant and a sensitive species to understand the effect of trans-regulatory factors in activating the LEA-like 11-24 promoter fragments. The results demonstrated that both desiccation tolerant and sensitive species contain essential trans-regulatory factors for the activity of LEA-like 11-24 promoter fragments. Using a yeast-one-hybrid system, it was previously shown the Cp LEA-like 11-24 promoter fragment interacts with CpbZIP1 protein. The CpbZIP1 belongs to the S1-class of AtbZIP transcription factors from Arabidopsis and has been shown to have 47% protein sequence identity with AtbZIP53. Transcript expression analysis of CpbZIP1 in response to ABA, dehydration and salt stress demonstrated the stress responsiveness of this gene. S1-class members of AtbZIP from Arabidopsis have complex expression patterns in different stress responses. The CpbZIP1 from C. plantagineum also showed similar complex expression pattern in response to stress. The function of CpbZIP1 transcription is completely unknown in C. plantagineum. In a functional study of the CpbZIP1 transcription factor, transgenic Arabidopsis plants overexpressing CpbZIP1 exhibited dwarf phenotype, abnormality in flower structure and small siliques suggesting that CpbZIP1 interferes with the developmental processes. Physiological and molecular data from transgenic plants overexpressing CpbZIP1 revealed the contribution of CpbZIP1 in remobilization of carbon/nitrogen or maintaining homeostasis of energy, as obtained for the bZIP homologue in Arabidopsis (AtbZIP53). Time dependent expression of CpbZIP1 revealed that apart from the function of CpbZIP1 in abiotic stress, most likely CpbZIP1 is regulated by circadian clock rhythm. The results from this study suggests that CpbZIP1 is stress inducible and involved in multiple functions like growth, flower development, accumulation of proline, remobilizing carbon/nitrogen and maintaining homeostasis of energy in low energy conditions.
Classification (DDC)
570 Biowissenschaften, Biologie 580 Pflanzen (Botanik)Ataei, Saeedeh: Promoter regulation of the Craterostigma plantagineum Cp LEA-like 11-24 gene and functional characterization of its transcription factor (CpbZIP1). - Bonn, 2014. - Dissertation, Rheinische Friedrich-Wilhelms-Universität Bonn.
Online-Ausgabe in bonndoc: https://nbn-resolving.org/urn:nbn:de:hbz:5n-37279
Online-Ausgabe in bonndoc: https://nbn-resolving.org/urn:nbn:de:hbz:5n-37279
@phdthesis{handle:20.500.11811/6161,
urn: https://nbn-resolving.org/urn:nbn:de:hbz:5n-37279,
author = {{Saeedeh Ataei}},
title = {Promoter regulation of the Craterostigma plantagineum Cp LEA-like 11-24 gene and functional characterization of its transcription factor (CpbZIP1)},
school = {Rheinische Friedrich-Wilhelms-Universität Bonn},
year = 2014,
month = aug,
note = {The resurrection plant Craterostigma plantagineum, a member of Linderniaceae family, is distributed in areas with variable water availability in Southern Africa. Two closely related species, L. brevidens which is endemic to montan rain forest of Africa and L. subracemosa which is found in central and Southern Africa, are used for comparative analyses for unravelling the molecular mechanisms of desiccation tolerance. While C. plantagineum and L. brevidens are desiccation tolerant, L. subracemosa represents a desiccation sensitive species. Desiccation tolerance is a very complex trait. Many genes that are involved in desiccation tolerance and the proteins that are encoded by these genes have been characterized in C. plantagineum. The Cp LEA-like 11-24 is one of these stress inducible genes in C. plantagineum that has been extensively studied both at the transcriptional and posttranscriptional level. Studies have been performed on the promoter regulation and transcription factor binding along with comparative promoter analyses between two desiccation tolerant and a desiccation sensitive species. Since promoter regions are the main regulatory elements for enhancing the transcriptional efficiency, developing an efficient transient transformation system was required to analyze promoter functions in response to different abiotic stresses. In this study, an efficient Agrobacterium-mediated transient transformation method was developed in three closely related Linderniaceae species. Different parameters that are important for enhancing the transformation efficiency were considered. These parameters include leaf size, Silwet L-77 concentration, bacterial density and duration of co-cultivation. After optimization, the activities of the minimal stress inducible promoter fragments of the LEA-like 11-24 from three species were examined under ABA and osmotic stress using optimized Agrobacterium co-cultivation method.
The transformation efficiency was found to be 100% in all three species. In addition, the induction pattern of each promoter fragment under ABA and osmotic stress was comparable with that of stably transformed plants or particle bombardment, suggesting this as the most suitable method for quantitatively analyzing promoter activity in the three species. The optimized method was used for cross comparison of the LEA-like 11-24 promoter fragments among two desiccation tolerant and a sensitive species to understand the effect of trans-regulatory factors in activating the LEA-like 11-24 promoter fragments. The results demonstrated that both desiccation tolerant and sensitive species contain essential trans-regulatory factors for the activity of LEA-like 11-24 promoter fragments. Using a yeast-one-hybrid system, it was previously shown the Cp LEA-like 11-24 promoter fragment interacts with CpbZIP1 protein. The CpbZIP1 belongs to the S1-class of AtbZIP transcription factors from Arabidopsis and has been shown to have 47% protein sequence identity with AtbZIP53. Transcript expression analysis of CpbZIP1 in response to ABA, dehydration and salt stress demonstrated the stress responsiveness of this gene. S1-class members of AtbZIP from Arabidopsis have complex expression patterns in different stress responses. The CpbZIP1 from C. plantagineum also showed similar complex expression pattern in response to stress. The function of CpbZIP1 transcription is completely unknown in C. plantagineum. In a functional study of the CpbZIP1 transcription factor, transgenic Arabidopsis plants overexpressing CpbZIP1 exhibited dwarf phenotype, abnormality in flower structure and small siliques suggesting that CpbZIP1 interferes with the developmental processes. Physiological and molecular data from transgenic plants overexpressing CpbZIP1 revealed the contribution of CpbZIP1 in remobilization of carbon/nitrogen or maintaining homeostasis of energy, as obtained for the bZIP homologue in Arabidopsis (AtbZIP53). Time dependent expression of CpbZIP1 revealed that apart from the function of CpbZIP1 in abiotic stress, most likely CpbZIP1 is regulated by circadian clock rhythm. The results from this study suggests that CpbZIP1 is stress inducible and involved in multiple functions like growth, flower development, accumulation of proline, remobilizing carbon/nitrogen and maintaining homeostasis of energy in low energy conditions.},
url = {https://hdl.handle.net/20.500.11811/6161}
}
urn: https://nbn-resolving.org/urn:nbn:de:hbz:5n-37279,
author = {{Saeedeh Ataei}},
title = {Promoter regulation of the Craterostigma plantagineum Cp LEA-like 11-24 gene and functional characterization of its transcription factor (CpbZIP1)},
school = {Rheinische Friedrich-Wilhelms-Universität Bonn},
year = 2014,
month = aug,
note = {The resurrection plant Craterostigma plantagineum, a member of Linderniaceae family, is distributed in areas with variable water availability in Southern Africa. Two closely related species, L. brevidens which is endemic to montan rain forest of Africa and L. subracemosa which is found in central and Southern Africa, are used for comparative analyses for unravelling the molecular mechanisms of desiccation tolerance. While C. plantagineum and L. brevidens are desiccation tolerant, L. subracemosa represents a desiccation sensitive species. Desiccation tolerance is a very complex trait. Many genes that are involved in desiccation tolerance and the proteins that are encoded by these genes have been characterized in C. plantagineum. The Cp LEA-like 11-24 is one of these stress inducible genes in C. plantagineum that has been extensively studied both at the transcriptional and posttranscriptional level. Studies have been performed on the promoter regulation and transcription factor binding along with comparative promoter analyses between two desiccation tolerant and a desiccation sensitive species. Since promoter regions are the main regulatory elements for enhancing the transcriptional efficiency, developing an efficient transient transformation system was required to analyze promoter functions in response to different abiotic stresses. In this study, an efficient Agrobacterium-mediated transient transformation method was developed in three closely related Linderniaceae species. Different parameters that are important for enhancing the transformation efficiency were considered. These parameters include leaf size, Silwet L-77 concentration, bacterial density and duration of co-cultivation. After optimization, the activities of the minimal stress inducible promoter fragments of the LEA-like 11-24 from three species were examined under ABA and osmotic stress using optimized Agrobacterium co-cultivation method.
The transformation efficiency was found to be 100% in all three species. In addition, the induction pattern of each promoter fragment under ABA and osmotic stress was comparable with that of stably transformed plants or particle bombardment, suggesting this as the most suitable method for quantitatively analyzing promoter activity in the three species. The optimized method was used for cross comparison of the LEA-like 11-24 promoter fragments among two desiccation tolerant and a sensitive species to understand the effect of trans-regulatory factors in activating the LEA-like 11-24 promoter fragments. The results demonstrated that both desiccation tolerant and sensitive species contain essential trans-regulatory factors for the activity of LEA-like 11-24 promoter fragments. Using a yeast-one-hybrid system, it was previously shown the Cp LEA-like 11-24 promoter fragment interacts with CpbZIP1 protein. The CpbZIP1 belongs to the S1-class of AtbZIP transcription factors from Arabidopsis and has been shown to have 47% protein sequence identity with AtbZIP53. Transcript expression analysis of CpbZIP1 in response to ABA, dehydration and salt stress demonstrated the stress responsiveness of this gene. S1-class members of AtbZIP from Arabidopsis have complex expression patterns in different stress responses. The CpbZIP1 from C. plantagineum also showed similar complex expression pattern in response to stress. The function of CpbZIP1 transcription is completely unknown in C. plantagineum. In a functional study of the CpbZIP1 transcription factor, transgenic Arabidopsis plants overexpressing CpbZIP1 exhibited dwarf phenotype, abnormality in flower structure and small siliques suggesting that CpbZIP1 interferes with the developmental processes. Physiological and molecular data from transgenic plants overexpressing CpbZIP1 revealed the contribution of CpbZIP1 in remobilization of carbon/nitrogen or maintaining homeostasis of energy, as obtained for the bZIP homologue in Arabidopsis (AtbZIP53). Time dependent expression of CpbZIP1 revealed that apart from the function of CpbZIP1 in abiotic stress, most likely CpbZIP1 is regulated by circadian clock rhythm. The results from this study suggests that CpbZIP1 is stress inducible and involved in multiple functions like growth, flower development, accumulation of proline, remobilizing carbon/nitrogen and maintaining homeostasis of energy in low energy conditions.},
url = {https://hdl.handle.net/20.500.11811/6161}
}
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