Bleaching Phenotype Research Articles

Tetrapyrrole Metabolism Is Involved in Lesion Formation, Cell Death, in theArabidopsis lesion initiation 1Mutant

The Arabidopsis lesion initiation 1 (len1) mutant develops lesions on leaves without pathogen attack. The len1 plants display lesion formation as they grow under short-day conditions (SD), but not under long-day conditions (LD). This study was conducted to examine how lesion formation, viz., cell death, in len1 plants occurs under SD. I present genetic and physiological data to show that tetrapyrrole metobolism is necessary for lesion formation in len1 plants. Lesion formation was suppressed in the len1lin2 double mutant under SD. lesion initiation 2 (lin2) is another lesion mimic mutant with a defect in tetrapyrrole biosynthesis. Suppression of lesion formation in len1 plants was also observed when they were crossed with the mutants that had defects in other steps in tetrapyrrole metabolism. Suppression was correlated with reduced chlorophyll (Chl) levels in the double mutants. Furthermore, I found that dark-to-light transition caused a bleached phenotype in len1 plants, as in the case of antisense ACD1 (acd, accelerated cell death) plants. ACD1 encodes pheophorbide a oxygenase (PaO), which is involved in Chl catabolism in Arabidopsis. These results suggest that tetrapyrrole metabolism, especially Chl breakdown, might be involved in lesion formation in len1 plants.

Involvement of the SppA1 peptidase in acclimation to saturating light intensities in Synechocystis sp. strain PCC 6803.

The sll1703 gene, encoding an Arabidopsis homologue of the thylakoid membrane-associated SppA peptidase, was inactivated by interposon mutagenesis in Synechocystis sp. strain PCC 6803. Upon acclimation from a light intensity of 50 to 150 microE m(-2) s(-1), the mutant preserved most of its phycobilisome content, whereas the wild-type strain developed a bleaching phenotype due to the loss of about 40% of its phycobiliproteins. Using in vivo and in vitro experiments, we demonstrate that the DeltasppA1 strain does not undergo the cleavage of the L(R)(33) and L(CM)(99) linker proteins that develops in the wild type exposed to increasing light intensities. We conclude that a major contribution to light acclimation under a moderate light regime in cyanobacteria originates from an SppA1-mediated cleavage of phycobilisome linker proteins. Together with changes in gene expression of the major phycobiliproteins, it contributes an additional mechanism aimed at reducing the content in phycobilisome antennae upon acclimation to a higher light intensity.

Photo-oxidative Stress in a Xanthophyll-deficient Mutant of Chlamydomonas

When there is an imbalance between the light energy absorbed by a photosynthetic organism and that which can be utilized in photosynthesis, photo-oxidative stress can damage pigments, proteins, lipids, and nucleic acids. In this work we compared the wild type and a xanthophyll-deficient mutant of Chlamydomonas reinhardtii in their response to high amounts of light. Wild-type Chlamydomonas cells were able to acclimate to high amounts of light following transfer from low light conditions. In contrast, the npq1 lor1 double mutant, which lacks protective xanthophylls (zeaxanthin and lutein) in the chloroplast, progressively lost viability and photosynthetic capacity along with destruction of thylakoid membrane protein-pigment complexes and accumulation of reactive oxygen species and membrane lipid peroxides. Loss of viability was partially rescued by lowered oxygen tension, suggesting that the high sensitivity of the mutant to light stress is caused by the production of reactive oxygen species in the chloroplast. Cell death was not prevented by the addition of an organic carbon source to the growth medium, demonstrating that the photo-oxidative damage can target other essential chloroplast processes besides photosynthesis. From the differential sensitivity of the mutant to exogenously added pro-oxidants, we infer that the reactive oxygen species produced during light stress in npq1 lor1 may be singlet oxygen and/or superoxide but not hydrogen peroxide. The bleaching phenotype of npq1 lor1 was not due to enhanced photodamage to photosystem II but rather to a less localized phenomenon of accumulation of photo-oxidation products in chloroplast membranes.

Bansformation of tobacco with a mutated cyanobacterial phytoene desaturase gene confers resistance to bleaching herbicides.

Carotenoids are constituents of the photosynthetic apparatus and essential for plant survival because of their involvement in protection of chlorophylls against photooxidation. Certain classes of herbicides are interfering with carotenoid biosynthesis leading to pigment destruction and a bleached plant phenotype. One important target site for bleaching herbicides is the enzyme phytoene desaturase catalysing the desaturation of phytoene in zeta-carotene. This enzymatic reaction can be inhibited by norflurazon or fluridone. We have transformed tobacco with a mutated cyanobacterial phytoene desaturase gene (pds) derived from the Synechococcus PCC 7942 mutant NFZ4. Characterization of the resulting transformants revealed an up to 58 fold higher norflurazon resistance in comparison to wild type controls. The tolerance for fluridone was also increased 3 fold in the transgenics. Furthermore, the transformed tobacco maintained a higher level of D1 protein of photosystem II indicating a lower susceptibility to photooxidative damage in the presence of norflurazon. In contrast, the genetic manipulation did not confer herbicide resistance against zeta-carotene desaturase inhibitors.

Virus-induced silencing of FtsH gene in Nicotiana benthmiana causes a striking bleached leaf phenotype.

A recombinant Potato virus X (PVX) vector, pTXS.FtsH, harboring partial sequence of FtsH gene of Nicotiana benthamiana was constructed to silence the expression of endogenous FtsH homologous gene in N. benthamiana. Inoculation with in vitro runoff transcript of pTXS.FtsH to N. benthamiana plants allowed silencing of FtsH, causing striking bleaching of upper leaves reminiscent of var2 mutant phenotype of Arabidopsis thaliana. FtsH-silenced plants exhibited no resistance against Tobacco mosaic virus (TMV) and a phytopathogenic fungus Botrytis cinerea. Virus-induced gene silencing (VIGS) of N. benthamiana with PVX as demonstrated here would be an efficient and rapid method to study the function of other elements of photosystem II (PSII) in planta.

Composition of photosystem II antenna in light-harvesting complex II antisense tobacco plants at varying irradiances.

Plants with genes coding for chlorophyll a/b-binding proteins of light-harvesting complex II (LHCII) in antisense orientation (Lhcb) that are characterized by severely reduced Lhcb transcript levels (below 10% of wild type) do not show a bleached phenotype due to a specific loss of the polypeptide. To produce such a phenotype, a conceptually different antisense approach was tested with a dual-functional transcript encoding the gene for hygromycin phosphotransferase and the transit sequence of Lhcb1-2 in the antisense orientation. Using increasing concentrations of hygromycin, transformants with Lhcb steady-state levels as low as 9% of wild type were regenerated and grown in a growth chamber. Together with Lhcb antisense plants obtained in an earlier study, these antisense plants were analyzed biochemically for their photosystem II (PSII) antenna composition under varying light conditions. All antisense plants showed a characteristic low-irradiance-induced increase of their PSII antenna size as determined by higher chlorophyll concentrations, an increased content of LHCII, and a constant chlorophyll b-to-lutein ratio in comparison with control plants. One to 5% of the total Lhcb transcript amount was sufficient to allow unrestricted formation of the PSII antenna at low irradiance, suggesting that LHCH biogenesis is not controlled primarily by transcription.

Molecular lesions associated with white gene mutations induced by I-R hybrid dysgenesis in Drosophila melanogaster

We have identified molecular lesions associated with six mutations, w and w, of the white gene of Drosophila melanogaster. These mutations arose in flies subject to I-R hybrid dysgenesis. Four of the mutations give rise to coloured eyes and are associated with insertions of 5.4-kb elements indistinguishable from the I factor controlling I-R dysgenesis. The insertion associated with w is at a site which, within the resolution of these experiments, is identical to that of two previously studied I factors. This appears to be a hot-spot for I factor insertion. We have compared the sites of these insertions with sequences complementary to white gene mRNA identified by Pirrotta and Bröckl. The hot-spot is in the fourth intron. The insertion carried by w is either within, or just beyond, the last exon. The insertion carried by w is near the junction of the first exon and first intron. The w mutation is a derivative of w. It contains an insertion of I factor DNA within, or immediately adjacent to, the F-like element associated with w, and results in restoration of some eye colour. This insertion is just upstream of the start of the white mRNA. Mutations w and w are deletions removing mRNA coding sequences. Both determine a bleached white phenotype.