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Citrus carotenoid isomerase gene characterization by complementation of the “Micro-Tom” tangerine mutant

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Abstract

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Complementation of the “Micro-Tom” tomato tangerine mutant with a Citrus CRTISO allele restores the wild-type fruit carotenoid profile, indicating that the Citrus allele encodes an authentic functional carotenoid isomerase.

Abstract

Citrus fruits are rich in carotenoids; the genus offers a large diversity in composition, yet to be fully explored to improve fruit nutritional quality. As perennial tree species, Citrus lack the resources for functional genetic studies, requiring the use of model plant systems. Here, we used the “Micro-Tom” (MT) tomato carrying the tangerine mutation (t), deficient for the carotenoid isomerase (CRTISO) gene, to functionally characterize the homologous C. sinensis genes. We identified four putative loci in the C. sinensis genome, named CsCRTISO, CsCRTISO-Like 1, CsCRTISO-Like 2, and CsCRTISO-Like 2B, with the latter as a presumed duplication of CRTISO-Like 2. In general, all the Citrus paralogs showed less expression specialization than the tomato ones, with CsCRTISO being the most expressed gene in all tissues analyzed. MT-t plants were successfully complemented with the CsCRTISO, and fruits showed a carotenoid profile similar to the control, indicating that the Citrus allele indeed encodes an authentic functional carotenoid isomerase and that the signal peptide is functional in tomato. MT was silenced using an inverted repeat of a fragment from the Citrus CRTISO resulting in a stronger phenotype than MT-t. MT-t and MT silenced for CRTISO presented an overall decrease in transcript accumulation of all genes from the biosynthesis pathway. The expression of the Citrus CRTISO gene is able to restore the biosynthesis of carotenoids with the appropriate regulation in MT-t. The decrease in transcript accumulation in MT-t and MT-CRTISO-suppressed lines reinforces previous suggestions that transcriptional regulation of the carotenoid biosynthesis involves regulatory loops by intermediate products.

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Acknowledgements

We thank Maiara Curtolo for assistance in identifying the location of CsCRTISO-L1 in the Citrus genome. TTP, LEPP, EP, RAM, MMM, and AF were recipients of fellowships from the Brazilian National Research Council (CNPq), whose support was greatly appreciated. The funder had no role in the study design, data collection, analysis and interpretation, decision to publish, or preparation of the manuscript.

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Authors and Affiliations

  1. Centro de Energia Nuclear na Agricultura, Universidade de São Paulo, Av. Centenário 303, CP 96, Piracicaba, SP, 13400-970, Brazil

    Thaísa T. Pinheiro, Rodolfo A. Maniero, Mônica M. Martins & Antonio Figueira

  2. Escola Superior de Agricultura “Luiz de Queiroz”, Departamento de Ciências Biológicas, Universidade de São Paulo, Av. Pádua Dias 11, CP 09, Piracicaba, SP, 13418-900, Brazil

    Lázaro E. P. Peres

  3. Departamento de Alimentos e Nutrição Experimental, Food Research Center (FoRC), Faculdade de Ciências Farmacêuticas, Universidade de São Paulo, Av. Professor Lineu Prestes 580, Bloco 14, São Paulo, SP, 05508-000, Brazil

    Eduardo Purgatto

  4. Centro APTA Citros “Sylvio Moreira”, Instituto Agronômico, Rodovia Anhanguera, km 158, CP 04, Cordeirópolis, SP, 13490-970, Brazil

    Rodrigo R. Latado

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  1. Thaísa T. Pinheiro
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Pinheiro, T.T., Peres, L.E.P., Purgatto, E. et al. Citrus carotenoid isomerase gene characterization by complementation of the “Micro-Tom” tangerine mutant. Plant Cell Rep 38, 623–636 (2019). https://doi.org/10.1007/s00299-019-02393-2

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