Abstract
Allatostatins (ASTs) are insect neuropeptide hormones that regulate diverse physiological functions, including feeding, growth and development, and reproduction. Therefore, regulation of allatostatin receptor (AstR) activity can be an effective tool for controlling insect growth and proliferation. Here, we describe a novel screening system using a mammalian cell line in which AstR is ectopically expressed, combined with fluorescence-based measurements of the membrane potential. HEK293T cells that do not express cognate receptors for AST became responsive to AST upon transfection with AstR. The response of the membrane potential to AST could be reliably detected by measuring the fluorescence of DiBAC4(3), a voltage-sensitive dye. We also discovered that overexpressing GIRK1/2 in this cell line could augment the magnitude of hyperpolarization by AST. Our screening system produces a fast and reliable readout for the efficient screening of AstR agonists.
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Acknowledgments
We thank Prof. Young-sik Lee for kindly providing the AstR cDNA clone. This work was supported by the Basic Science Research Program through the National Research Foundation of Korea (NRF) (NRF-2009-0075797) and a Korea university Grant.
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Na, J., Kim, W. & Yoon, BJ. Development of a novel screening system for allatostatin receptor agonists in search of new candidate insect growth regulators. Biotechnol Lett 36, 1439–1446 (2014). https://doi.org/10.1007/s10529-014-1505-6
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DOI: https://doi.org/10.1007/s10529-014-1505-6