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Title: A Novel, Photosynthesis-Associated Thioredoxin-Like Gene: Final Technical Report

Abstract

Many aspects of the biosynthesis and physiological regulation of the photosynthetic apparatus of plants, algae and cyanobacteria remain to be understood, and are likely to involve yet-unidentified proteins that carry out oxidation/reduction (redox) reactions. TxlA from Synechococcus sp. strain PCC 7942 and its homologues from other cyanobacteria and plants, including Sll1980 from the cyanobacterium Synechocystis sp. strain PCC 6803, are likely to be among these proteins. In fact, the homologue of TxlA in the plant Arabidopsis thaliana, HCF164, may be required for synthesis of the cytochrome b6f complex that transfers electrons between the two photosynthetic reaction centers. TxlAs share an N-terminal hydrophobic domain, a central thioredoxin-like domain, and a unique C-terminal hydrophilic domain. Plant and algal TxlAs are nuclear-encoded and have an additional N-terminal domain that targets them to the chloroplast. We have found that the common N-terminal domain of TxlA anchors it to a membrane, probably the thylakoid (photosynthetic) membrane (where HCF164 is also localized, with its thioredoxin-like domain in the thylakoid lumen). We have also found that the thioredoxin-like domain is likely to assume the conformation typical of thioredoxins and possesses thioredoxin-like redox activity in vitro, and that the C-terminal domain is important to the structure and functionmore » of the thioredoxin-like domain both in vivo and in vitro. These data show that TxlAs have the cellular location and enzymatic activity expected of a protein involved in the biosynthesis of redox components or redox regulation of the photosynthetic apparatus. We were unable to inactivate the thioredoxin-like domain of TxlA in either PCC 7942 or PCC 6803, under either photosynthetic or heterotrophic growth conditions. We also found that expression of antisense txlA mRNA from an IPTG-regulated promoter in PCC 7942 was lethal, most likely because it effectively inactivated txlA by ''RNA silencing''. These results are consistent with a role for TxlA in the synthesis of the cytochrome b6f complex, which is required for both photosynthetic and respiratory electron transport in cyanobacteria. In contrast, our PCC 7942 mutants in which the C-terminal domain of TxlA was removed are viable and appear to have normal cytochrome content, but have a subtle pigmentation phenotype (increased content of phycocyanin relative to chlorophyll) that depends on both light and CO2 availability. We have also found that PCC 6803 Sll1980 inactivation mutant merodiploids have a similar pigmentation phenotype to the PCC 7942 C-terminal truncation mutants when grown photoautotrophically. In addition, when grown heterotrophically the PCC 6803 Sll1980 inactivation mutant merodiploids remain green instead of turning a golden color like the wild-type, and they are more sensitive to the b6f complex inhibitor DBMIB than is wild type PCC 6803. That the PCC 6803 Sll1980 inactivation mutant merodiploids have these phenotypes despite the fact that they still contain normal copies of the sll1980 gene suggests that the presence of truncated Sll1980 protein interferes with the function of normal Sll1980 protein. Together, these physiological data suggest that TxlA has an essential redox role in cyanobacteria, perhaps a biosynthetic one, and may also have a nonessential regulatory role reflected in the phenotypes of the PCC 7942 C-terminal truncation mutants and the PCC 6803 Sll1980 inactivation mutant merodiploids.« less

Authors:
Publication Date:
Research Org.:
Stony Brook University, Stony Brook, NY; Rensselaer Polytechnic Institute, Troy, NY
Sponsoring Org.:
USDOE Office of Science (SC)
OSTI Identifier:
850272
Report Number(s):
DOE/ER/15329
TRN: US200707%%221
DOE Contract Number:  
FG02-02ER15329
Resource Type:
Technical Report
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; ALGAE; ARABIDOPSIS; BIOSYNTHESIS; CHLOROPHYLL; CYANOBACTERIA; CYTOCHROMES; ELECTRONS; GENES; IN VITRO; IN VIVO; INACTIVATION; MEMBRANES; MUTANTS; PHENOTYPE; PHOTOSYNTHETIC REACTION CENTERS; PHYCOCYANIN; PROTEINS; RNA; STRAINS; SYNTHESIS

Citation Formats

Collier, Jackie, L. A Novel, Photosynthesis-Associated Thioredoxin-Like Gene: Final Technical Report. United States: N. p., 2005. Web. doi:10.2172/850272.
Collier, Jackie, L. A Novel, Photosynthesis-Associated Thioredoxin-Like Gene: Final Technical Report. United States. doi:10.2172/850272.
Collier, Jackie, L. Tue . "A Novel, Photosynthesis-Associated Thioredoxin-Like Gene: Final Technical Report". United States. doi:10.2172/850272. https://www.osti.gov/servlets/purl/850272.
@article{osti_850272,
title = {A Novel, Photosynthesis-Associated Thioredoxin-Like Gene: Final Technical Report},
author = {Collier, Jackie, L},
abstractNote = {Many aspects of the biosynthesis and physiological regulation of the photosynthetic apparatus of plants, algae and cyanobacteria remain to be understood, and are likely to involve yet-unidentified proteins that carry out oxidation/reduction (redox) reactions. TxlA from Synechococcus sp. strain PCC 7942 and its homologues from other cyanobacteria and plants, including Sll1980 from the cyanobacterium Synechocystis sp. strain PCC 6803, are likely to be among these proteins. In fact, the homologue of TxlA in the plant Arabidopsis thaliana, HCF164, may be required for synthesis of the cytochrome b6f complex that transfers electrons between the two photosynthetic reaction centers. TxlAs share an N-terminal hydrophobic domain, a central thioredoxin-like domain, and a unique C-terminal hydrophilic domain. Plant and algal TxlAs are nuclear-encoded and have an additional N-terminal domain that targets them to the chloroplast. We have found that the common N-terminal domain of TxlA anchors it to a membrane, probably the thylakoid (photosynthetic) membrane (where HCF164 is also localized, with its thioredoxin-like domain in the thylakoid lumen). We have also found that the thioredoxin-like domain is likely to assume the conformation typical of thioredoxins and possesses thioredoxin-like redox activity in vitro, and that the C-terminal domain is important to the structure and function of the thioredoxin-like domain both in vivo and in vitro. These data show that TxlAs have the cellular location and enzymatic activity expected of a protein involved in the biosynthesis of redox components or redox regulation of the photosynthetic apparatus. We were unable to inactivate the thioredoxin-like domain of TxlA in either PCC 7942 or PCC 6803, under either photosynthetic or heterotrophic growth conditions. We also found that expression of antisense txlA mRNA from an IPTG-regulated promoter in PCC 7942 was lethal, most likely because it effectively inactivated txlA by ''RNA silencing''. These results are consistent with a role for TxlA in the synthesis of the cytochrome b6f complex, which is required for both photosynthetic and respiratory electron transport in cyanobacteria. In contrast, our PCC 7942 mutants in which the C-terminal domain of TxlA was removed are viable and appear to have normal cytochrome content, but have a subtle pigmentation phenotype (increased content of phycocyanin relative to chlorophyll) that depends on both light and CO2 availability. We have also found that PCC 6803 Sll1980 inactivation mutant merodiploids have a similar pigmentation phenotype to the PCC 7942 C-terminal truncation mutants when grown photoautotrophically. In addition, when grown heterotrophically the PCC 6803 Sll1980 inactivation mutant merodiploids remain green instead of turning a golden color like the wild-type, and they are more sensitive to the b6f complex inhibitor DBMIB than is wild type PCC 6803. That the PCC 6803 Sll1980 inactivation mutant merodiploids have these phenotypes despite the fact that they still contain normal copies of the sll1980 gene suggests that the presence of truncated Sll1980 protein interferes with the function of normal Sll1980 protein. Together, these physiological data suggest that TxlA has an essential redox role in cyanobacteria, perhaps a biosynthetic one, and may also have a nonessential regulatory role reflected in the phenotypes of the PCC 7942 C-terminal truncation mutants and the PCC 6803 Sll1980 inactivation mutant merodiploids.},
doi = {10.2172/850272},
journal = {},
number = ,
volume = ,
place = {United States},
year = {2005},
month = {9}
}