A novel fluorescent assay for sucrose transporters

Abstract

Sucrose transporters (SUTs or SUCs) play a critical role in the long distance transport of carbohydrates in plants. Products of photosynthesis must have an efficient means of being distributed to cells in the plant that depend on the net import of fixed carbon such as roots, flowers, and seeds. In many plants, this is achieved by active loading of the phloem using H+-coupled sucrose transporters. The SuSy7 growth assay is one of the traditional methods used to identify sucrose transporters but it tends to have high background and it is not quantitative. Here we introduce a novel assay for SUT activity which is based on the ability of type I sucrose transporters to transport the fluorescent coumarin β-glucoside, esculin. The esculin uptake assay is rapid and sensitive and should be useful for preliminary tests of sucrose transporter function by heterologous expression in yeast. This assay is also suitable for selection of yeast showing esculin uptake activity using FACS, making possible the rapid scanning of cDNA libraries for type I SUTs.