Tomato ATP-Binding Cassette Transporter SlABCB4 Is Involved in Auxin Transport in the Developing Fruit

Abstract

Plant ATP binding cassette (ABC) transporters are membrane proteins that are important for transporting a wide range of compounds, including secondary metabolites and phytohormones. In Arabidopsis, some members of the ABCB subfamily of ABC transporter, also known as Multi-Drug Resistance proteins (MDRs), have been implicated in auxin transport. However, reports on the roles of the auxin-mediated ABCBs in fleshy fruit development are rare. Here, we present that SlABCB4, a member of the tomato ABCB subfamily, transports auxin in the developing fruit of tomato. Transient expression of SlABCB4-GFP fusion proteins in tobacco cells showed plasma membrane localization. The transport activity of SlABCB4, expressed in Nicotiana benthamiana protoplasts, revealed substrate specificity for indole-3-acetic acid export. Gene expression analysis of SlABCB4 revealed high expression levels at the early stages of fruit development. Therefore, SlABCB4 is considered to facilitate auxin distribution in tomato fruit, which is important for tomato fruit development.

Keywords: ABC transporter; auxin; fruit development.

Figure 1

Phylogenetic tree of full-size ABCBs of tomato and Arabidopsis. Tomato ABCBs are shown in red. Arabidopsis ABCBs, characterized as auxin transporters, are indicated in blue and the others in black. The scale bar shows a 5% divergence between protein sequences [25].

Figure 2

Subcellular localization of SlABCB4 in Nicotiana benthamiana epidermal cells. (A) Localization of SlABCB4-GFP fusion proteins with FM4-64 dye at the plasma membrane (PM). (B) Localization of SlABCB4-GFP fusion proteins and PIP1;4-mCherry fusion proteins, a PM marker. Top to down: Green fluorescence of SlABCB4-GFP fusion proteins; red fluorescence of FM4-64 dye or PIP1;4-mCherry fusion proteins, merged image of top and second pictures; and bright-field microscope images.

Figure 3

Transport assay of SlABCB4 in Nicotiana benthamiana protoplast. Export assay of (A) indole-3-acetic acid (IAA) and (B) benzoic acid (BA). IAA and BA transport activities in Nicotiana benthamiana protoplast, expressing SlABCB4, SlABCB4-GFP or GFP-SlABCB4 and vector control, were measured. Error bars show standard error for at least four experimental repeats. The asterisk is significant at p < 0.05.

Figure 4

Gene expression analysis of SlABCB4 in various tomato organs. RNAs were extracted from indicated organs of ‘MicroTom’. Transcript levels of SlABCB4 were detected by RT-qPCR. The ubiquitin gene was used as a constitutively expressed control gene. Error bars are standard errors of 3 independent experiments. DAP: days after pollination.