Limits...
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.

Show MeSH

Related in: MedlinePlus

Fluorescence measurements as test for PSII activity in Δycf3 plants. Wild-type and mutant plants  grown under low light conditions were dark adapted, and  leaf samples were illuminated with white actinic light.  As a control, a wild-type  sample treated with the plastoquinone-reducing herbicide 3-(3,4-dichlorphenyl)- 1,1-dimethylurea (DCMU)  was included. PSII activity is  clearly detectable in Δycf3  plants. However, comparison  of the variable fluorescent  yields indicates that the mutant accumulates fewer functional PSII reaction centers  than the wild type, which  is most likely the result of  photooxidative damage as  caused by the lack of functional electron acceptors  downstream of PSII. The  course of the fluorescence  curve recorded for the mutant is virtually identical with the one  of the wild-type sample treated with DCMU demonstrating that  in both cases electrons accumulate in PSII and are not transferred to downstream components of the photosynthetic electron  transport chain.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2139824&req=5

Figure 5: Fluorescence measurements as test for PSII activity in Δycf3 plants. Wild-type and mutant plants grown under low light conditions were dark adapted, and leaf samples were illuminated with white actinic light. As a control, a wild-type sample treated with the plastoquinone-reducing herbicide 3-(3,4-dichlorphenyl)- 1,1-dimethylurea (DCMU) was included. PSII activity is clearly detectable in Δycf3 plants. However, comparison of the variable fluorescent yields indicates that the mutant accumulates fewer functional PSII reaction centers than the wild type, which is most likely the result of photooxidative damage as caused by the lack of functional electron acceptors downstream of PSII. The course of the fluorescence curve recorded for the mutant is virtually identical with the one of the wild-type sample treated with DCMU demonstrating that in both cases electrons accumulate in PSII and are not transferred to downstream components of the photosynthetic electron transport chain.

Mentions: Presence of functional PSII units in the mutant plants was further confirmed by measurements of PSII-dependent chlorophyll fluorescence at room temperature (Fig. 5). Even a moderate light flux of as little as 80 μE/m2s (corresponding to ∼6% of normal sunlight) resulted in a completely reduced pool of the primary quinone-type acceptor QA. This finding indicates that the electrons generated by PSII are not efficiently accepted by one of the downstream components of the electron transfer chain.


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)

Fluorescence measurements as test for PSII activity in Δycf3 plants. Wild-type and mutant plants  grown under low light conditions were dark adapted, and  leaf samples were illuminated with white actinic light.  As a control, a wild-type  sample treated with the plastoquinone-reducing herbicide 3-(3,4-dichlorphenyl)- 1,1-dimethylurea (DCMU)  was included. PSII activity is  clearly detectable in Δycf3  plants. However, comparison  of the variable fluorescent  yields indicates that the mutant accumulates fewer functional PSII reaction centers  than the wild type, which  is most likely the result of  photooxidative damage as  caused by the lack of functional electron acceptors  downstream of PSII. The  course of the fluorescence  curve recorded for the mutant is virtually identical with the one  of the wild-type sample treated with DCMU demonstrating that  in both cases electrons accumulate in PSII and are not transferred to downstream components of the photosynthetic electron  transport chain.
© Copyright Policy
Related In: Results  -  Collection

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

Figure 5: Fluorescence measurements as test for PSII activity in Δycf3 plants. Wild-type and mutant plants grown under low light conditions were dark adapted, and leaf samples were illuminated with white actinic light. As a control, a wild-type sample treated with the plastoquinone-reducing herbicide 3-(3,4-dichlorphenyl)- 1,1-dimethylurea (DCMU) was included. PSII activity is clearly detectable in Δycf3 plants. However, comparison of the variable fluorescent yields indicates that the mutant accumulates fewer functional PSII reaction centers than the wild type, which is most likely the result of photooxidative damage as caused by the lack of functional electron acceptors downstream of PSII. The course of the fluorescence curve recorded for the mutant is virtually identical with the one of the wild-type sample treated with DCMU demonstrating that in both cases electrons accumulate in PSII and are not transferred to downstream components of the photosynthetic electron transport chain.
Mentions: Presence of functional PSII units in the mutant plants was further confirmed by measurements of PSII-dependent chlorophyll fluorescence at room temperature (Fig. 5). Even a moderate light flux of as little as 80 μE/m2s (corresponding to ∼6% of normal sunlight) resulted in a completely reduced pool of the primary quinone-type acceptor QA. This finding indicates that the electrons generated by PSII are not efficiently accepted by one of the downstream components of the electron transfer chain.

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