Effect of high light conditions on the response of Arabidopsis thaliana plants with suppressed mitochondrial alternative oxidase

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Abstract

BACKGROUND: Plants as sessile organisms have developed biochemical pathways to protect themselves from the excess light energy. Mitochondrial alternative oxidase (AOX) participates in the oxidation of reductants exported from chloroplasts, thereby optimizing photosynthesis and protecting cells from photodamage.

AIM: The effect of high light on respiration and the relative transcripts content of a number of genes in Arabidopsis thaliana plants of the T-DNA insertional line for AOX1a (aox1a) was studied and compared with the response of the antisense silencing of AOX1a line (AS-12) and wild type line Col-0.

MATERIALS AND METHODS: Four-week-old A. thaliana plants of three lines grown at 90 µmol/m2 · s and then exposed to moderately high light conditions, 400 µmol/m2 · s, in a short-term experiment (8 h). Respiratory pathways activity, gene expression, and superoxide anion content were determined during experiment.

RESULTS: Plants of the aox1a line in response to high light were characterized by the absence of the total and alternative respiration reaction and the absence of the AOX1 protein in spite of the increased mRNA level of AOX1c, in contrast to the Col-0 and AS-12 lines. Also, an increased content of transcripts of only SAPX and CHS were found, while in the other lines a compensatory increase in the expression of many “defense” genes was revealed.

CONCLUSIONS: Thus, the aox1a line was characterized by a low compensatory effect at the level of defense systems activation. This is apparently caused by the absence of the AOX1 protein and, as a result, the weakening of the stress signal and stress response. The results obtained indicate the important role of AOX in the response of respiration to light stress; can be used to study the signaling pathways of regulation of AOX1a expression.

About the authors

Elena V. Garmash

Institute of Biology Komi Science Centre of the Ural Branch of the Russian Academy of Sciences

Author for correspondence.
Email: garmash@ib.komisc.ru
ORCID iD: 0000-0001-8104-5048
SPIN-code: 4512-8460

Dr. Sci. (Biol.), leading research associate

Russian Federation, Syktyvkar

Kirill V. Yadrikhinskiy

Pitirim Sorokin Syktyvkar State University

Email: kirill030442@gmail.com

student
Russian Federation, Syktyvkar

Mikhail A. Shelyakin

Institute of Biology Komi Science Centre of the Ural Branch of the Russian Academy of Sciences

Email: shelyakin@ib.komisc.ru
ORCID iD: 0000-0001-8537-6995
SPIN-code: 9526-1351

Cand. Sci. (Biol.), research associate

Russian Federation, Syktyvkar

Elena S. Belykh

Institute of Biology Komi Science Centre of the Ural Branch of the Russian Academy of Sciences

Email: belykh@ib.komisc.ru
ORCID iD: 0000-0002-0182-6475
SPIN-code: 1804-9569

Cand. Sci. (Biol.), research associate

Russian Federation, Syktyvkar

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2. Fig. 1. Total respiration (Vt) and activity of CN-resistant (alternative) respiration (RCN) in leaves of Arabidopsis thaliana plants with different expression of AOX1a grown at 90 μmol/(m2 · s) (0 h) and exposed for 8 h to increased illumination of 400 μmol/(m2 · s). Col-0 is a wild ecotype, aox1a is a line with a knockout of AOX1a, AS-12 is a line transformed by an antisense copy of AOX1a. The mean values from three experiments are presented (in each experiment n = 5) and their standard errors. The significance of parameter changes among all the presented average values is marked with different lowercase Latin letters (ANOVA, Duncan’s test, p < 0.05)

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3. Fig. 2. The relative content of alternative oxidase (AOX) gene transcripts in the leaves of Arabidopsis thaliana plants with different AOX1a expression grown at 90 μmol/(m2 · s) (0 h) and exposed to increased illumination for 8 h — 400 μmol/(m2 · s). Col-0 — wild ecotype, aox1a is a line with a knockout of AOX1a, AS-12 is a line transformed by an antisense copy of AOX1a. The mean values from three experiments (in each experiment n = 9) and their standard errors are presented. The significance of parameter changes for each gene is marked with different lowercase Latin letters (ANOVA, Duncan’s test, p < 0.05)

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4. Fig. 3. Alternative oxidase (AOX) immunoblots in a protein extract obtained from the leaves of Arabidopsis thaliana plants of lines with different levels of AOX1a expression grown at 90 μmol/(m2 · s) (0 h) and exposed for 8 h to high light at 400 μmol/(m2 · s). Col-0 — a wild ecotype, aox1a — T-DNA insertional line for AOX1a, AS-12 — antisense silencing of AOX1a line. The arrows on the left indicate the molecular weights of marker proteins

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5. 24 Fig. 4. The content of superoxide radical in the leaves of Arabidopsis thaliana plants of lines with different levels of AOX1a expression, grown at 90 μmol/(m2 · s) (0 h) and exposed for 8 h to high light at 400 μmol/(m2 · s). Col-0 — wild ecotype, aox1a — T-DNA insertional line for AOX1a, AS-12 — antisense silencing of AOX1a line. The mean values from three experiments (in each experiment n = 4–6) and their standard errors are presented. The reliability of parameter changes is marked with different lowercase Latin letters (ANOVA, Duncan test, p < 0.05)

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6. Fig. 5. Heat maps of genes encoding components of the respiratory pathways and antioxidant enzymes relative to their level in the control variant (0 h) in Arabidopsis thaliana plants with different expression of AOX1a grown at 90 μmol/(m2 · s) (0 h) and exposed for 8 h to high light at 400 μmol/(m2 · s). Col-0 — wild ecotype, aox1a — T-DNA insertional line for AOX1a, AS-12 — antisense silencing of AOX1a line. Black indicates an increase, white indicates a decrease in the relative content of gene transcripts

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