Person:
Veramendi Charola, Jon

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Veramendi Charola

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Jon

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AgronomĆ­a, BiotecnologĆ­a y AlimentaciĆ³n

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IMAB. Research Institute for Multidisciplinary Applied Biology

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0000-0002-3214-213X

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539

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Now showing 1 - 3 of 3
  • PublicationOpen Access
    Overexpression of thioredoxin m in chloroplasts alters carbon and nitrogen partitioning in tobacco
    (Oxford University Press, 2021) AncĆ­n RĆ­podas, MarĆ­a; Larraya Reta, Luis MarĆ­a; Florez-Sarasa, Igor; BĆ©nard, Camille; FernĆ”ndez San MillĆ”n, Alicia; Veramendi Charola, Jon; Gibon, Yves; Fernie, Alisdair R.; Aranjuelo Michelena, Iker; FarrĆ”n Blanch, Inmaculada; Agronomia, Bioteknologia eta Elikadura; Institute for Multidisciplinary Research in Applied Biology - IMAB; AgronomĆ­a, BiotecnologĆ­a y AlimentaciĆ³n
    In plants, there is a complex interaction between carbon (C) and nitrogen (N) metabolism, and its coordination is fundamental for plant growth and development. Here, we studied the influence of thioredoxin (Trx) m on C and N partitioning using tobacco plants overexpressing Trx m from the chloroplast genome. The transgenic plants showed altered metabolism of C (lower leaf starch and soluble sugar accumulation) and N (with higher amounts of amino acids and soluble protein), which pointed to an activation of N metabolism at the expense of carbohydrates. To further delineate the effect of Trx m overexpression, metabolomic and enzymatic analyses were performed on these plants. These results showed an up-regulation of the glutamine synthetase-glutamate synthase pathway; specifically tobacco plants overexpressing Trx m displayed increased activity and stability of glutamine synthetase. Moreover, higher photorespiration and nitrate accumulation were observed in these plants relative to untransformed control plants, indicating that overexpression of Trx m favors the photorespiratory N cycle rather than primary nitrate assimilation. Taken together, our results reveal the importance of Trx m as a molecular mediator of N metabolism in plant chloroplasts.
  • PublicationOpen Access
    Functional improvement of human cardiotrophin 1 produced in tobacco chloroplasts by co-expression with plastid thioredoxin m
    (MDPI, 2020) Ancƭn Rƭpodas, Marƭa; Sanz Barrio, Ruth; Santamarƭa, Eva; FernƔndez San MillƔn, Alicia; Larraya Reta, Luis Marƭa; Veramendi Charola, Jon; FarrƔn Blanch, Inmaculada; Institute for Multidisciplinary Research in Applied Biology - IMAB
    Human cardiotrophin 1 (CT1), a cytokine with excellent therapeutic potential, was previously expressed in tobacco chloroplasts. However, the growth conditions required to reach the highest expression levels resulted in an impairment of its bioactivity. In the present study, we have examined new strategies to modulate the expression of this recombinant protein in chloroplasts so as to enhance its production and bioactivity. In particular, we assessed the effect of both the fusion and co-expression of Trx m with CT1 on the production of a functional CT1 by using plastid transformation. Our data revealed that the Trx m fusion strategy was useful to increase the expression levels of CT1 inside the chloroplasts, although CT1 bioactivity was significantly impaired, and this was likely due to steric hindrance between both proteins. By contrast, the expression of functional CT1 was increased when co-expressed with Trx m, because we demonstrated that recombinant CT1 was functionally active during an in vitro signaling assay. While Trx m/CT1 co-expression did not increase the amount of CT1 in young leaves, our results revealed an increase in CT1 protein stability as the leaves aged in this genotype, which also improved the recombinant proteinā€™s overall production. This strategy might be useful to produce other functional biopharmaceuticals in chloroplasts.
  • PublicationOpen Access
    New in vivo approach to broaden the thioredoxin family interactome in chloroplasts
    (MDPI, 2022) Ancƭn Rƭpodas, Marƭa; FernƔndez Irigoyen, Joaquƭn; Santamarƭa Martƭnez, Enrique; Larraya Reta, Luis Marƭa; FernƔndez San MillƔn, Alicia; Veramendi Charola, Jon; FarrƔn Blanch, Inmaculada; Ciencias de la Salud; Osasun Zientziak; Institute for Multidisciplinary Research in Applied Biology - IMAB
    Post-translational redox modifications provide an important mechanism for the control of major cellular processes. Thioredoxins (Trxs), which are key actors in this regulatory mechanism, are ubiquitous proteins that catalyse thiol-disulfide exchange reactions. In chloroplasts, Trx f, Trx m and NADPH-dependent Trx reductase C (NTRC) have been identified as transmitters of the redox signal by transferring electrons to downstream target enzymes. The number of characterised Trx targets has greatly increased in the last few years, but most of them were determined using in vitro procedures lacking isoform specificity. With this background, we have developed a new in vivo approach based on the overexpression of His-tagged single-cysteine mutants of Trx f, Trx m or NTRC into Nicotiana benthamiana plants. The over-expressed mutated Trxs, capable of forming a stable mixed disulfide bond with target proteins in plants, were immobilised on affinity columns packed with Ni-NTA agarose, and the covalently linked targets were eluted with dithiothreitol and identified by mass spectrometry-based proteomics. The in vivo approach allowed identification of 6, 9 and 42 new potential targets for Trx f, Trx m and NTRC, respectively, and an apparent specificity between NTRC and Trxs was achieved. Functional analysis showed that these targets are involved in several cellular processes.