The role of cis-elements in transcriptional regulation of the phaseolin promoter /
β-phaseolin, the major seed storage protein of Phaseolus vulgaris, is under strict transcriptional regulation during development in transgenic tobacco. The stringency in spatial regulation and the quantitative levels of promoter activity are displayed by the lack of expression in vegetative tissues...
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| Format: | Thesis Book |
| Language: | English |
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[Place of publication not identified] :
[publisher not identified] ;
2001.
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| Subjects: | |
| Online Access: | http://proxy.library.tamu.edu/login?url=http://proquest.umi.com/pqdweb?did=725921381&sid=1&Fmt=2&clientId=2945&RQT=309&VName=PQD |
| Summary: | β-phaseolin, the major seed storage protein of Phaseolus vulgaris, is under strict transcriptional regulation during development in transgenic tobacco. The stringency in spatial regulation and the quantitative levels of promoter activity are displayed by the lack of expression in vegetative tissues and high levels of activity during embryogenesis. Dimethyl sulfate-in vivo footprinting analyses showed factor binding to twenty three cis-elements of the promoter in seed tissues. The complex footprinting obtained is probably a summation of all the different factor-DNA interaction in various regions of the seed. To determine the functional contributions of the cis-elements, two sets of experiments were undertaken. In the first set of experiments, an alternative plant model system, Arabidopsis thaliana, was evaluated for its ability to maintain the rigor of phas promoter regulation. Results showed that a majority of the transgenic Arabidopsis plants maintains the stringency of regulation. High levels of expression were obtained in seed tissues. However, 32% of the lines transformed with a vector bearing the phas promoter adjacent to the T-DNA right border showed reporter expression in vegetative tissues due to an 'enhancer-trap' phenomenon. Based on the observations acquired from a vast majority of transgenic lines, the major step has been accomplished in moving the phas system to the genetically and experimentally compliant Arabidopsis model. In the second set of experiments, the contributions of ten cis-elements to the regulation of the phas promoter were determined by incorporating site-directed substitution mutations into the -295phas promoter. The effect of these mutations on the spatial expression and quantitative levels of activity of the debilitated promoters were evaluated in Arabidopsis. Results showed that both redundancy and hierarchy exist in cis-element functions. The G-box and CCAAAT box have been identified as major positive elements. The E-box and RY sites have been shown to function in mediating high levels of expression. The RY elements have synergistic interactions to facilitate activation, some RY elements being required for radicle-specific repression. The CACA element has dual positive and negative regulatory roles and the vicilin box is a repressive element. Combinatorial interactions between cis-elements are important in conferring module-specific expression within the embryo. |
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| Item Description: | Vita. "Major Subject: Biology". |
| Physical Description: | xii, 157 leaves : illustrations ; 28 cm. Issued also on microfiche from University Microfilm Inc. |
| Bibliography: | Includes bibliographical references (leaves 138-155). |