Functional characterization of RNA polymerase III subunit RPC4, a non plastidic ADP-glucose pyrophosphorylase interacting protein in Arabidopsis
plastidic ADP-glucose pyrophosphorylase interacting protein in Arabidopsis
- 발행기관 Pohang University of Science and Technology
- 발행년도 2013
- 학위수여년월 2013. 2
- 학위명 석사
- 학과 및 전공 일반대학원 융합생명공학부
- 실제URI http://www.dcollection.net/handler/postech/000001556295
- 본문언어 영어
- 저작권 포항공과대학교 논문은 저작권에 의해 보호받습니다.
초록/요약
We previously have studied ADP-glucose pyrophosphorylase (ATP: alpha-glucose-1-phosphate adenylyl transferase, ADGase), a key regulatory enzyme in the starch biosynthetic pathway of plants. Surprisingly, I found that ADP-glucose pyrophosphorylase small subunit APS1 (ADG1) is not only localized in chloroplast but also in non plastidic region, especially, small proportion in nucleus. To unravel function of non plastidic ADG1, by applying yeast two hybrid screens, we have identified one specific ADG1 unconventional partner: RNA polymerase III subunit 4 (RPC4). In Arabidopsis, RNA polymerase III subunit 4 was encoded by two genes At5g09380 (RPC4) and At4g25180 (RPC4 homolog). RPC4 is widely expressed in Arabidopsis and green fluorescent protein-RPC4 fusion protein localizes specifically to the cell nucleus, forms two different patterns. I have characterized Arabidopsis lines harboring T-DNA insertions in the coding sequence of RPC4. Remarkably, rpc4 loss of function mutant displayed similar phenotype with adg1 mutant in growth defects, observed in smaller leaf size, delayed flowering and delayed senescence. Moreover, rpc4 homolog loss of function mutant showed similar longer petiole phenotypes with adg1 mutant under low light and dark grown seedling. Together with evidence for direct interactions of ADG1 and RPC4, these genetic and phenotypic analyses supported the idea that RPC4 might act together with ADG1 in the same pathway. A BLAST search of the Arabidopsis thaliana database using the amino acid sequence of the 30-kDa Arabidopsis RPC4 as query revealed one striking match with a 42-kDa protein encoded Rpc53p [Saccharomyces cerevisiae], which is well studied in regulating tRNA gene transcription in vivo in S. cerevisiae. Finding of RPC4 protein as one of unconventional ADG1 interacting partners supported the first characterization of RPC4 protein in Arabidopsis development. As the result, it is suggested the function of ADG1 in regulation of RNA polymerase III through interacting with RNA pol III subunit so that plants can adapt to the specific environmental conditions
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ABSTRACT i
LIST OF FIGURES AND TABLES v
ABBREVIATIONS vi
1. INTRODUCTION 1
1.1 Moonlighting proteins 1
1.2 ADP-glucose pyrophosphorylase is a key regulatory enzyme in the starch biosynthetic pathway 1
1.3 ADG1 can localize in non plastidic region 2
2. MATERIALS AND METHODS 3
3. RESULTS 5
3.1 Identification of ADG1 Unconventional Partners 5
3.2 Confirmation of interaction between ADG1 and candidates 6
3.3 Functional RNA polymerase III subunit 4 in Arabidopsis development 13
3.3.1 Characterization of RNA polymerase III subunit 4 protein in Arabidopsis 13
3.3.2 T-DNA insertion knockout mutant selection 16
3.3.3 Phenotypic analysis of rpc4 mutants 17
3.3.4 Arabidopsis RPC4 encodes a protein with striking similarity to RPC53 in yeast 17
4 DISCUSSION 25
5 CONCLUSION 27
6 REFERENCES 28
7 ACKNOWLEDGMENT 33
8 CURRICULUM VITAE 34