Homepage: elsevier/locate/febsopenbioPotential use of potassium efflux-deficient yeast for studying trafficking signals and potassium channel functionsJoshua D. Bernstein, Yukari Okamoto, Minjee Kim, Sojin Shikano*Department of Biochemistry and Molecular Genetics, University of Illinois at Chicago, Chicago, IL 60607, USAa r t i c l ei n f oa b s t r a c tThe activity of potassium (K + ) channels critically depends upon their density around the cell surface membrane, that is regulated by dynamic protein rotein interactions that typically involve distinct trafficking signals around the cargo proteins. Within this paper we explored the possibility of utilizing the Saccharomyces cerevisiae strain B31 for identification with the signal motifs that regulate surface expression of membrane proteins and for studying structure unction relationships of K + channels. B31 cells lack the K + efflux technique and have been reported to show overloaded K + -mediated development inhibition in high K + media upon heterologous expression of a mammalian inwardly rectifying K + channel (Kir2.1). We show that when the expression of wild-type Kir2.1 channel inhibits the growth of B31 cells in higher K + media, the human disease-causing mutations of Kir2.1 that abolish K + conduction (V302M) or surface trafficking ( 314/315) totally restores the growth. The expression of two-pore-domain K + channel KCNK3 or KCNK9 also inhibited the growth of B31 in higher K + media even though C-terminal mutations that cut down their 14-3-3 protein-dependent cell surface trafficking restored the growth of B31. Ultimately, the expression of Kir2.1 channels that had been C-terminally fused with recognized sequence motifs such as ER retention/retrieval signals and an endocytosis signal permitted the development of B31 in high K + media.5-Hydroxypicolinaldehyde Chemscene These results demonstrate the potential of B31 yeast strain as a exclusive biological tool to screen the random peptide libraries for novel sequence signals that down-regulate surface expression of membrane proteins, too as to systematically determine the structural determinants for cell surface trafficking and/or ion conductance of K + channels.Formula of 77500-04-0 C 2013 The Authors.PMID:33406813 Published by Elsevier B.V. on behalf of Federation of European Biochemical Societies. All rights reserved.Report history: Received 18 February 2013 Received in revised form 2 April 2013 Accepted 11 April 2013 Keyword phrases: Potassium channel Yeast Trafficking Surface membrane Screening1. Introduction Ion channels play critical roles inside the cellular response to signals and changes within the extracellular environment by controlling ion homeostasis and membrane excitability. Their right functioning critically is dependent upon the fine tuning of their localization at cell surface. This course of action is dictated by the secretory and endocytic vesicular trafficking pathways that generally involve dynamic interactions involving distinct sequence signals on the cargos and cellular transport machineries. Quite a few crucial discoveries on trafficking signals have come in the studies on ion channels which includes K + channels. As an illustration, the Arg-based (`RXR’) endoplasmic reticulum (ER) retention/This is an open-access article distributed under the terms on the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, offered the original author and supply are credited. Abbreviations: ER, endoplasmic reticulum; FCM, flow cytometry; COP, coatomer protein; IP, immunoprecipitation; Ab, antibody; HA, hemagglutinine; SD, regular dev.