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Targeted inactivation of a tobacco intron-containing open reading frame reveals a novel chloroplast-encoded photosystem I-related gene.

Ruf S, Kössel H, Bock R - J. Cell Biol. (1997)

Bottom Line: Faithful transcription of photosystem I genes as well as correct mRNA processing and efficient transcript loading with ribosomes in the Deltaycf3 plants suggest a posttranslational cause of the PSI-defective phenotype.We therefore propose that ycf3 encodes an essential protein for the assembly and/or stability of functional PSI units.This study provides a first example for the suitability of reverse genetics approaches to complete our picture of the coding capacity of higher plant chloroplast genomes.

View Article: PubMed Central - PubMed

Affiliation: Institut für Biologie III, Universität Freiburg, Germany.

ABSTRACT
The chloroplast genome of all higher plants encodes, in its large single-copy region, a conserved open reading frame of unknown function (ycf3), which is split by two group II introns and undergoes RNA editing in monocotyledonous plants. To elucidate the function of ycf3 we have deleted the reading frame from the tobacco plastid genome by biolistic transformation. We show here that homoplasmic Deltaycf3 plants display a photosynthetically incompetent phenotype. Molecular analyses indicate that this phenotype is not due to a defect in any of the general functions of the plastid genetic apparatus. Instead, the mutant plants specifically lack detectable amounts of all photosystem I (PSI) subunits analyzed. In contrast, at least under low light conditions, photosystem II subunits are still present and assemble into a physiologically active complex. Faithful transcription of photosystem I genes as well as correct mRNA processing and efficient transcript loading with ribosomes in the Deltaycf3 plants suggest a posttranslational cause of the PSI-defective phenotype. We therefore propose that ycf3 encodes an essential protein for the assembly and/or stability of functional PSI units. This study provides a first example for the suitability of reverse genetics approaches to complete our picture of the coding capacity of higher plant chloroplast genomes.

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Phenotype of homoplasmic Δycf3 plants. (A) A mutant  plant kept under standard light conditions (3.5–4 W/m2). Massive  photooxidative damage in mutant chloroplasts results in completely white plants. (B) A mutant plant grown under low light  conditions (0.4–0.5 W/m2). Bars, 1 cm.
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Figure 3: Phenotype of homoplasmic Δycf3 plants. (A) A mutant plant kept under standard light conditions (3.5–4 W/m2). Massive photooxidative damage in mutant chloroplasts results in completely white plants. (B) A mutant plant grown under low light conditions (0.4–0.5 W/m2). Bars, 1 cm.

Mentions: Shoots from homoplasmic Δycf3 lines displayed a pale-green phenotype upon regeneration on spectinomycin-containing medium under standard light conditions (3.5–4 W/m2). When transferred to boxes (for rooting on drug- and phytohormone-free medium), the plants bleached out completely within a few days (Fig. 3 A). The phenotype was much less severe under low light conditions (0.4–0.5 W/m2). The plants were now light green (Fig. 3 B), and nearly indistinguishable from wild-type plants kept under identical conditions. However, the mutant plants grew very slowly, and after maintenance for more than 6 wk the lower leaves began to turn white. Only young leaves (up to 3-wk-old) from plants grown under low light conditions were used for the following molecular analyses.


Targeted inactivation of a tobacco intron-containing open reading frame reveals a novel chloroplast-encoded photosystem I-related gene.

Ruf S, Kössel H, Bock R - J. Cell Biol. (1997)

Phenotype of homoplasmic Δycf3 plants. (A) A mutant  plant kept under standard light conditions (3.5–4 W/m2). Massive  photooxidative damage in mutant chloroplasts results in completely white plants. (B) A mutant plant grown under low light  conditions (0.4–0.5 W/m2). Bars, 1 cm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2139824&req=5

Figure 3: Phenotype of homoplasmic Δycf3 plants. (A) A mutant plant kept under standard light conditions (3.5–4 W/m2). Massive photooxidative damage in mutant chloroplasts results in completely white plants. (B) A mutant plant grown under low light conditions (0.4–0.5 W/m2). Bars, 1 cm.
Mentions: Shoots from homoplasmic Δycf3 lines displayed a pale-green phenotype upon regeneration on spectinomycin-containing medium under standard light conditions (3.5–4 W/m2). When transferred to boxes (for rooting on drug- and phytohormone-free medium), the plants bleached out completely within a few days (Fig. 3 A). The phenotype was much less severe under low light conditions (0.4–0.5 W/m2). The plants were now light green (Fig. 3 B), and nearly indistinguishable from wild-type plants kept under identical conditions. However, the mutant plants grew very slowly, and after maintenance for more than 6 wk the lower leaves began to turn white. Only young leaves (up to 3-wk-old) from plants grown under low light conditions were used for the following molecular analyses.

Bottom Line: Faithful transcription of photosystem I genes as well as correct mRNA processing and efficient transcript loading with ribosomes in the Deltaycf3 plants suggest a posttranslational cause of the PSI-defective phenotype.We therefore propose that ycf3 encodes an essential protein for the assembly and/or stability of functional PSI units.This study provides a first example for the suitability of reverse genetics approaches to complete our picture of the coding capacity of higher plant chloroplast genomes.

View Article: PubMed Central - PubMed

Affiliation: Institut für Biologie III, Universität Freiburg, Germany.

ABSTRACT
The chloroplast genome of all higher plants encodes, in its large single-copy region, a conserved open reading frame of unknown function (ycf3), which is split by two group II introns and undergoes RNA editing in monocotyledonous plants. To elucidate the function of ycf3 we have deleted the reading frame from the tobacco plastid genome by biolistic transformation. We show here that homoplasmic Deltaycf3 plants display a photosynthetically incompetent phenotype. Molecular analyses indicate that this phenotype is not due to a defect in any of the general functions of the plastid genetic apparatus. Instead, the mutant plants specifically lack detectable amounts of all photosystem I (PSI) subunits analyzed. In contrast, at least under low light conditions, photosystem II subunits are still present and assemble into a physiologically active complex. Faithful transcription of photosystem I genes as well as correct mRNA processing and efficient transcript loading with ribosomes in the Deltaycf3 plants suggest a posttranslational cause of the PSI-defective phenotype. We therefore propose that ycf3 encodes an essential protein for the assembly and/or stability of functional PSI units. This study provides a first example for the suitability of reverse genetics approaches to complete our picture of the coding capacity of higher plant chloroplast genomes.

Show MeSH
Related in: MedlinePlus