Analysis of the VIT1 Promoter Activity in Developing Arabidopsis thaliana Plants

Iron (Fe) deficiency in plants is one of the widespread problems limiting agricultural production. Generating crops more tolerant to Fe deficiency by genetic engineering or breeding is of great interest but challenging due to the knowledge gaps in general plant Fe homeostasis. Although several genes involved in Fe homeostasis have been identified, characterization of their roles is mainly limited to specific organs at specific developmental stages of the plant, where their mutants show the most striking phenotype. Vacuolar Iron Transporter 1 (VIT1) is a well-known gene that has been characterized for its function in the mature seed of Arabidopsis thaliana. VIT1 is an Fe transporter that determines the correct distribution of Fe storage in this organ. The study aimed to explore new physiological functions for VIT1. As a first step, Arabidopsis thaliana plants that contain PromoterVIT1: GUS constructs were used to study the temporal and spatial expression of the gene throughout the plant’s lifecycle. GUS histochemical staining revealed the VIT1 promoter is active in the mature leaves and mature reproductive organs. VIT1 promoter activity in the stamen increased developmentally and was limited to tapetum and guard cells in the pollen sac. In the female organ, VIT1 promoter activity increased as the pistil developed into a silique. Although all the silique exhibited staining, staining density was higher in the peduncle, replum, and stigma regions. Inside the developing silique, funicles were heavily stained. Furthermore, in silico analyses of VT1 transcriptome and protein levels confirmed flower and the silique are hot spots for VT1 activity. Thus, the results may suggest a possible involvement of VT1 protein in several stages of the reproductive system, specifically in the flowering and in the fruit developmen

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Abadía J, Vázquez S, Rellán-Álvarez R, El-Jendoubi H, Abadía A, Álvarez-Fernández A, López-Millán AF. 2011. Towards a knowledge-based correction of iron chlorosis. Plant Physiology and Biochemistry, 49(5): 471–482. https://doi.org/10.1016/j.plaphy.2011.01.026
Briat JF, Lebrun M. 1999. Plant responses to metal toxicity. Comptes Rendus de l’Academie Des Sciences - Serie III, 322(1): 43–54. https://doi.org/10.1016/S0764-4469(99)800 16-X
Castaings L, Caquot A, Loubet S, Curie C. 2016. The high- affinity metal Transporters NRAMP1 and IRT1 Team up to Take up Iron under Sufficient Metal Provision. Scientific Reports, 6: 37222. https://doi.org/10.1038/srep37222
Connorton JM, Jones ER, Rodríguez-Ramiro I, Fairweather-Tait S, Uauy C, Balk J. 2017. Wheat vacuolar iron transporter TaVIT2 transports Fe and Mn and is effective for biofortification. Plant Physiology, 174(4): 2434–2444. https://doi.org/10.1104/pp.17.00672
Durrett TP, Gassmann W, Rogers EE. 2007. The FRD3-mediated efflux of citrate into the root vasculature is necessary for efficient iron translocation. Plant Physiology, 144(1): 197– 205. https://doi.org/10.1104/pp.107.097162
Eroglu S, Giehl RFH, Meier B, Takahashi M, Terada Y, Ignatyev K, Andresen E, Küpper H, Peiter E, Von Wirén N. 2017. Metal tolerance protein 8 mediates manganese homeostasis and iron reallocation during seed development and germination. Plant Physiology, 174(3): 1633–1647. https://doi.org/10.1104/pp.16.01646
Eroglu S, Karaca N, Vogel-Mikus K, Kavčič A, Filiz E,Tanyolac, B. 2019. The conservation of VT1-dependent iron distribution in seeds. Frontiers in Plant Science, 10. https://doi.org/10.3389/fpls.2019.00907
Eroglu S, Meier B, Von Wirén N, Peiter E. 2016. The vacuolar manganese transporter mtp8 determines tolerance to iron deficiency-induced chlorosis in arabidopsis. Plant Physiology, 170(2): 1030–1045. https://doi.org/10.1104/pp. 15.01194
Holloway RE, Bertrand I, Frischke AJ, Brace DM, Mclaughlin MJ, Shepperd W. 2001. Improving fertiliser efficiency on calcareous and alkaline soils with fluid sources of P, N and Zn. Plant and Soil, 236(2): 209–219. https://doi.org/10.1023/ A:1012720909293
Hruz T, Laule O, Szabo G, Wessendorp F, Bleuler S, Oertle L, Widmayer P, Gruissem W, Zimmermann P. 2008. Genevestigator V3: A Reference Expression Database for the Meta-Analysis of Transcriptomes. Advances in Bioinformatics, 2008, 1–5. https://doi.org/10.1155/2008/42 0747
Kato T, Kumazaki K, Wada M, Taniguchi R, Nakane T, Yamashita K, Hirata K, Ishitani R, Ito K, Nishizawa T, Nureki O. 2019. Crystal structure of plant vacuolar iron transporter VT1. Nature Plants, 5(3): 308–315. https://doi. org/10.1038/s41477-019-0367-2
Kim SA, Punshon T, Lanzirotti A, Li A, Alonso JM, Ecker JR, Kaplan J, Guerinot M. Lou. 2006. Localization of iron in Arabidopsis seed requires the vacuolar membrane transporter VT1. Science, 314(5803): 1295–1298. https://doi.org/10. 1126/science.1132563
Korshunova YO, Eide D, Clark WG, Guerinot M. Lou, Pakrasi, H. B. (1999). The IRT1 protein from Arabidopsis thaliana is a metal transporter with a broad substrate range. Plant Molecular Biology, 40(1): 37–44. https://doi.org/10.1023/ A:1026438615520
Mergner J, Frejno M, List M, Papacek M, Chen X, Chaudhary A, Samaras P, Richter S, Shikata H, Messerer M, Lang D, Altmann S, Cyprys P, Zolg DP, Mathieson T, Bantscheff M, Hazarika RR, Schmidt T, Dawid C, Kuster B. 2020. Mass- spectrometry-based draft of the Arabidopsis proteome. Nature, 579(7799): 409–414. https://doi.org/10.1038/s4158 6-020-2094-2
Narayanan N, Beyene G, Chauhan RD, Gaitán-Solís E, Gehan J, Butts P, Siritunga D, Okwuonu I, Woll A, Jiménez-Aguilar DM, Boy E, Grusak MA, Anderson P, Taylor NJ. 2019. Biofortification of field-grown cassava by engineering expression of an iron transporter and ferritin. Nature Biotechnology, 37(2): 144–151. https://doi.org/10.1038/s415 87-018-0002-1
Narayanan N, Beyene G, Chauhan RD, Gaitán-Solis E, Grusak MA, Taylor N, Anderson P. 2015. Overexpression of Arabidopsis VT1 increases accumulation of iron in cassava roots and stems. Plant Science, 240: 170–181. https://doi.org/ 10.1016/j.plantsci.2015.09.007
Schuler M, Rellán-Álvarez R, Fink-Straube C, Abadía J, Bauera P. 2012. Nicotianamine functions in the phloem-based transport of iron to sink organs, in pollen development and pollen tube growth in Arabidopsis. Plant Cell, 24(6): 2380– 2400. https://doi.org/10.1105/tpc.112.099077
Vigani G, Zocchi G, Bashir K, Philippar K, Briat JF. 2013. Signals from chloroplasts and mitochondria for iron homeostasis regulation. Trends in Plant Science, 18(6): 305– 311. https://doi.org/10.1016/j.tplants.2013.01.00