Influence of knockout of At4g20990 gene encoding α-CA4 on photosystem II light-harvesting antenna in plants grown under different light intensities and day lengths

Abstract

Effect of knockout of the At4g20990 gene encoding α-carbonic anhydrase 4 (α-CA4) in Arabidopsis thaliana in plants grown in low light (LL, 80 μmol photons m−2 s−1) or in high light (HL, 400 μmol photons m−2 s−1) under long (LD, 16 h) or short (SD, 8 h) day length was studied. In α-CA4 knockout plants, under all studied conditions, the non-photochemical quenching was lower; the decrease was more pronounced under HL. This pointed to α-CA4 implication in the processes leading to energy dissipation in PSII antenna. In this context the content of major antenna proteins Lhcb1 and Lhcb2 was lower in α-CA4 knockouts than in wild-type (WT) plants under all growth conditions. The expression level of lhcb2 gene was also lower in mutants grown under LD, LL and HL in comparison to WT. At the same time, this level was higher in mutants grown under SD, LL and it was the same under SD, HL. Overall, the data showed that the knockout of the At4g20990 gene affected both the contents of proteins of PSII light-harvesting complex and the expression level of genes encoding these proteins, with peculiarities dependent on day length. These data together with the fact of a decrease of non-photochemical quenching of leaf chlorophyll a fluorescence in α-CA4-mut as compared with that in WT plants implied that α-CA4 participates in acclimation of photosynthetic apparatus to light intensity, possibly playing important role in the photoprotection. The role of this CA can be especially important in plants growing under high illumination conditions.

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Acknowledgements

The authors express their gratitude to Dr. V.A. Mudrik for providing plant growth chamber functioning, to Dr. I.A. Naydov for help with densitometry, and to Dr. J.V. Moroney of Louisiana State University for providing the seeds of homozygous lines of mutants with knocked out genes of CA produced in his laboratory, the possibility to conduct primer design and PCR. This work was supported by Russian Foundation for Basic Research No. 15-04-03883.

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Correspondence to Natalia N. Rudenko.

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Supplementary Figure 1
figure4

Scheme of Arabidopsis plant growth of SD-plants, 8 h day/16 h night (dark gray), and LD-plants, 16 h day/8 h night (light gray) at low (LL, 80 μmol photons m−2 s−1) and high (HL, 400 μmol photons m−2 s−1) light intensity. (GIF 51 kb)

Supplementary Figure 2
figure5

Result of electrophoresis in agarose gel of PCR products obtained with specific primer pairs designed in different exon sequences of the At4g20990 gene with WT cDNA (A), α-CA4-mut (line 8–8) (B), α-CA4-mut (line 9–12) (C). D – PCR products obtained with specific primer pair designed in exon sequences of actin gene with WT cDNA. E – DNA molecular weight ladder. The length of PCR products in gels A and D correspond to predicted ones. (GIF 66 kb)

Supplementary Figure 3
figure6

Fragments of gels after SDS electrophoresis of thylakoid proteins from leaves of WT and the mutant lines, 8–8 and 9–12, of SD (8 h day/16 h night) (A) and LD (16 h day/8 h night) (B). The plants were grown either under low light (LL, 80 μmol photons m−2 s−1) or high light (HL, 400 μmol photons m−2 s−1). Numbers near the right side of the gel represent the molecular masses of marker proteins in kDa. The amounts of Lhcb proteins were determined by densitometric analysis. The data for α-CA4-mut are in % of those of WT grown under the same light intensity. In parentheses are data for WT grown at HL in % of those of WT grown in LL. Data are shown as mean ± the SE. An adaptation to an increased light intensity in both WT and mutant was manifested in the decrease in the contents of Lhch3, and Lhcb6 proteins, especially in the mutant in LD-plants; the contents of Lhcb3 and Lhcb6 decreased in WT to 30% and in mutant to 6% as compared with their contents in LL. The contents of Lhcb4 and Lhcb5 proteins under increase in light intensity during growth decreased in both WT and α-CA4-mut at LD; at SD, the contents of these proteins in WT did not change, while they did notably increase in the mutant. (GIF 99 kb)

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Rudenko, N.N., Fedorchuk, T.P., Vetoshkina, D.V. et al. Influence of knockout of At4g20990 gene encoding α-CA4 on photosystem II light-harvesting antenna in plants grown under different light intensities and day lengths. Protoplasma 255, 69–78 (2018). https://doi.org/10.1007/s00709-017-1133-9

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Keywords

  • Arabidopsis thaliana
  • Carbonic anhydrase
  • Thylakoid
  • Non-photochemical quenching
  • Light-harvesting complex of photosystem II
  • Lhcb proteins