Circadian Regulation of Expression of Carotenoid Metabolism Genes (PSY2, LCYE, CRTRB1, NCED1) in Leaves of Tomato Solanum lycopersicum L.

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Abstract

The circadian dynamics of the expression of key genes of carotenoid metabolism (PSY2, LCYE, CrtRB1, and NCED1) in the photosynthetic tissue of tomato Solanum lycopersicum L. (cultivar Korneevsky) plants was characterized. An in silico analysis of the gene expression pattern was carried out and a high level of their transcripts was detected in the leaf tissue. qRT-PCR analysis of gene expression was performed at six time points during the day and found the highest levels of PSY2, LCYE and NCED1 transcripts in the second half of the light phase, and CrtRB1 – at the end of the dark phase. The content and composition of carotenoids in leaf tissue in the middle of the day was determined and it was shown that the leaf accumulates 1.5 times more compounds of the ɛ/β-branch of carotenoid biosynthesis pathway than compounds of the β/β-branch.

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About the authors

M. A. Filyushin

Institute of Bioengineering, Federal Research Center “Fundamentals of Biotechnology” of the Russian Academy of Sciences

Author for correspondence.
Email: michel7753@mail.ru
Russian Federation, Moscow

A. V. Shchennikova

Institute of Bioengineering, Federal Research Center “Fundamentals of Biotechnology” of the Russian Academy of Sciences

Email: michel7753@mail.ru
Russian Federation, Moscow

E. Z. Kochieva

Institute of Bioengineering, Federal Research Center “Fundamentals of Biotechnology” of the Russian Academy of Sciences

Email: michel7753@mail.ru
Russian Federation, Moscow

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Supplementary files

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2. Fig. 1. Heatmap of gene expression of PSY2 (Solyc02g081330; NCBI gene ID 543964), LCYE (Solyc12g008980; 544129), NCED1 (Solyc07g056570; 544163), and CrtRB1 (Solyc06g036260; 544133) in tomato (S. lycopersicum) cv. Heinz, constructed from TomExpress transcriptomic data (http://tomexpress.toulouse.inra.fr/). Organs analyzed: root (1); leaf (2); bud (3); fruit with diameter 1 (4), 2 (5), and 3 (6) cm; immature fruit of final size (7); fruit at the stage of color change from green to red (8). Data visualization (http://www2.heatmapper.ca/expression/) was performed based on FPKM values (Fragments per kilo base of transcript per million mapped fragments) indicated in the boxes (rounded to two decimal places).

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3. Fig. 2. (a) Daily dynamics of gene expression of PSY2 (Solyc02g081330; NCBI gene ID 543964), LCYE (Solyc12g008980; 544129), NCED1 (Solyc07g056570; 544163), and CrtRB1 (Solyc06g036260; 544133) in leaf tissue of seedlings of tomato cv. Korneevsky. Day and night periods are indicated below the graphs by white and gray boxes, respectively. Primers: PSY2 (5´-aactgagctctgctagtagatg-3´, 5´-gcactagagatcttgcataagca-3´); LCYE (5´-tatgtcagacacgacgctcaat-3´, 5´-catgctagcagcagcacca-3´); NCED1 (5´-tgctcttagctacgatgtgat-3´, 5´-gcgaaatcatgcatcattgttgg-3´); CrtRB1 (5´-cacttcttatcaatgaggaaggg-3´, 5´-cttactgaataagaagagcctac-3´). Data were normalized against two reference genes Expressed (gene ID 101263039; 5´-gctaagaacgctggacctaatg-3´, 5´-tgggtgtgcctttctgaatg-3´) and actin-7 (101262163; 5´-cattgtgctcagtggtggttc-3´, 5´-tctgctggaaggtgctaagtg-3´). According to statistical analysis (Two-way ANOVA; GraphPad Prism v. 8, GraphPad Software Inc., USA; https://www.graphpad.com/scientific-software/prism/), expression levels of each gene differ significantly (p < 0.05) between time points, except PSY2 (15.00 vs. 22.00; 6.00 vs. 8.00), LCYE (6.00 vs. 8.00), and NCED1 (3.00 vs. 8.00, 24.00; 6.00 vs. 15.00, 22.00). (b) Carotenoid composition (%) in leaves of tomato cv. Korneevsky seedlings collected in the middle of the daylight photosynthetic phase (15.00). The total carotenoid content was 307.3 μg/g fresh weight. The total carotenoid content (mg/g fresh weight) and its composition (% of total carotenoids of individual compounds) were determined according to [15] in two biological and three technical replicates.

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