Solanum lycopersicum AUXIN RESPONSE FACTOR 9 regulates cell division activity during early tomato fruit development

Journal of Experimental Botany Solanum lycopersicum AUXIN RESPONSE FACTOR 9 regulates cell division activity during early tomato fruit development Maaike de Jong 0 4 Mieke Wolters-Arts 4 Bernardus C. J. Schimmel 2 4 Catharina L. M. Stultiens 4 Peter F. M. de Groot 4 Stephen J. Powers 3 Yury M. Tikunov 1 Arnoud G. Bovy 1 Celestina Mariani 4 Wim H. Vriezen 4 5 Ivo Rieu 4 0 Present address: The Sainsbury Laboratory, University of Cambridge , Bateman Street, Cambridge, CB2 1LR , UK 1 Plant Research International, Wageningen University & Research Plant Breeding , Droevendaalsesteeg 1, 6708PB Wageningen , The Netherlands 2 Present address: Department of Population Biology, Institute for Biodiversity and Ecosystem Dynamics, University of Amsterdam , Science Park 904, 1098 XH Amsterdam , The Netherlands 3 Computational and Systems Biology, Rothamsted Research , West Common, Harpenden, Hertfordshire, AL5 2JQ , UK 4 Department of Molecular Plant Physiology, Institute for Water and Wetland Research, Radboud University Nijmegen , Heyendaalseweg 135, 6525 AJ Nijmegen , The Netherlands 5 Present address: Bayer CropScience Vegetable Seeds , Voort 6, 6083 AC Nunhem , The Netherlands The transformation of the ovary into a fruit after successful completion of pollination and fertilization has been associated with many changes at transcriptomic level. These changes are part of a dynamic and complex regulatory network that is controlled by phytohormones, with a major role for auxin. One of the auxin-related genes differentially expressed upon fruit set and early fruit development in tomato is Solanum lycopersicum AUXIN RESPONSE FACTOR 9 (SlARF9). Here, the functional analysis of this ARF is described. SlARF9 expression was found to be auxin-responsive and SlARF9 mRNA levels were high in the ovules, placenta, and pericarp of pollinated ovaries, but also in other plant tissues with high cell division activity, such as the axillary meristems and root meristems. Transgenic plants with increased SlARF9 mRNA levels formed fruits that were smaller than wild-type fruits because of reduced cell division activity, whereas transgenic lines in which SlARF9 mRNA levels were reduced showed the opposite phenotype. The expression analysis, together with the phenotype of the transgenic lines, suggests that, in tomato, ARF9 negatively controls cell division during early fruit development. Auxin; AUXIN RESPONSE FACTOR 9 (ARF9); cell division; fruit development; fruit size; tomato (Solanum lycopersicum L; ) - The development of the closed carpel is thought to be one of the features that contributed to the evolutionary success of the angiosperms (Scutt et al., 2006). The carpel is the female reproductive organ that differentiates into stigma, style, and the ovary, the latter of which encloses the ovules. After successful completion of pollination and fertilization, the ovary Abbreviations: AuxRE, auxin response elements; BR, brassinosteroid; CaMV, cauliflower mosaic virus; c-DNA-AFLP, cDNA-amplified fragment length polymorphism–based transcript profiling; CTD, C-terminal homo- and heterodimerization domains; DAP, days after pollination; FDR, false discovery rate; GA, gibberellin; GSEA, genome set enrichment analysis; GUS, β-glucuronidase; IAA, indole-3-acetic acid; kb, kilobase; MR, middle region; OE, overexpression line; PA, polyamine; PCA principal component analysis; SED, standard error of the difference. © The Author 2015. Published by Oxford University Press on behalf of the Society for Experimental Biology. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited. develops into a fruit, with the ovary wall becoming the pericarp and the ovules developing into seeds. The fruit creates a protected environment for the seeds to mature and may mediate dispersal of the mature seeds (Gillaspy et al., 1993). It is widely assumed that reproductive development occurs at the expense of vegetative growth (Snow and Whigham, 1989). Accordingly, the wild progenitors of many fruit crop species produce smaller fruits compared to the domesticated species (Tanksley, 2004; Doebley et al., 2006). A good example is the tomato, with wild relatives such as Solanum pimpinellifolium bearing small fruit, and cultivated tomato species (Solanum lycopersicum L.) producing large fruit with a more than 100-fold increase in weight compared to wild species (Grandillo et  al., 1999; Tanksley, 2004). The tomato has been extensively used as a plant model species to study fruit development. Although research has mainly focussed on the later stages of fruit growth, processes occurring during fruit set and early stages of fruit development also have implications on the traits of the mature fruit, such as fruit size and shape (Paran and van der Knaap, 2007). During flower development, cells at the floral meristems proliferate and differentiate to form the floral organs. The rate, duration, and direction of cell divisions in the developing ovary may already substantially impact final fruit size and shape (Bohner and Bangerth, 1988; van der Knaap et al., 2014). When the ovary has reached its mature size, cell division activity stops. After a few days, the flower may abscise or, upon successful completion of pollination and fertilization, set fruit by resuming further cell division (Gillaspy et al., 1993). This period continues for 10–14 days, and largely determines the final number of cells in the fruit (Bohner and Bangerth, 1988). In the next stage of development, fruit growth essentially depends on cell expansion, with cells increasing up to a 100-fold in volume (Tanksley, 2004). After this 6–7-week period the fruit has reached its final size and will start to ripen (Mapelli et al., 1978; Bünger-Kibler and Bangerth, 1982; Gillaspy et al., 1993). Auxin plays an important role in tomato fruit set and fruit development. The auxin concentration in the ovary rapidly increases within 2  days after pollination (Mariotti et  al., 2011). The application of auxin on unpollinated ovaries leads to the formation of fruits without the need for pollination and fertilization (Gustafson, 1936; Bünger-Kibler and Bangerth, 1982). Similarly, affecting auxin synthesis or responsiveness by the ovary-specific expression of the iaaM or rolB genes from Agrobacterium spp. (Ficcadenti et  al., 1999; Carmi et al., 2003) or the overexpression of the auxin receptor TRANSPORT INHIBITOR RESPONSE 1 (TIR1) (Ren et al., 2011) resulted in the formation of seedless tomato fruits. Down-regulation of transcription factors involved in the regulation of auxin-mediated gene expression, like Aux/indole-3-acetic acid (IAA) 9 and AUXIN RESPONSE FACTOR 7 (ARF7) also results in fruit development without the need for pollination and fertilization (Wang et  al., 2005a; (...truncated)