Characterization of Calcium Binding Peptide Derived from Calreticulin and Its Effect on Increasing Bioavailable Calcium in Plants

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Title: Characterization of Calcium Binding Peptide Derived from Calreticulin and Its Effect on Increasing Bioavailable Calcium in Plants
Author: Tsou, Pei-Lan
Advisors: Dr. Ross Whetten, Committee Member
Dr. Eric Davies, Committee Member
Dr. Dominique Robertson, Committee Chair
Dr. Nina S. Allen, Committee Member
Abstract: Modulation of cytosolic calcium levels in both plants and animals is achieved by a system of Ca2+ -transport and storage pathways that include Ca2+ buffering proteins in the lumen of intracellular compartments. We used a transgenic approach to modulate calcium stores in the lumen of the endoplasmic reticulum in plants. We were able to use the low affinity, high capacity Ca2+ binding characteristics of the C-domain of calreticulin (CRT) to selectively increase Ca2+ storage in the endoplasmic reticulum, and to determine if those alterations affected plant physiological responses to stress. I fused the CRT C-domain to the C-terminal portion of the green fluorescent protein (GFP) and obtained transgenic plants that expressed high levels of the GFP-CRT C-domain fusion protein in the ER. I showed that this fusion protein was able to bind Ca2+ in vitro. These plants had increased Ca2+ stores in the ER and the ER calcium stores were used by the plant in times of calcium stress. The transformed seedlings also showed delayed loss of chlorophyll after transfer to high salt medium. The data suggest that expression of this calcium binding protein not only has the ability to increase bioavailable calcium stores but that the ER Ca2+ stores are important for plants to relieve the effect of salt stress. In the process of developing the transgenic plants, I also developed an array of other tools for subcellular localization, and targeting of the C-domain to various subcellular organelles. With two other colleagues, we also developed a GFP imaging system using the onion epidermal cells for visualizing subcellular localization of GFP and GFP fusion proteins.
Date: 2002-08-19
Degree: PhD
Discipline: Botany
URI: http://www.lib.ncsu.edu/resolver/1840.16/5202


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