2+ Sarco/Endoplasmic Reticulum Ca -ATPases (SERCA) Contribute to GPCR-Mediated Taste Perception Naoko Iguchi1, Tadahiro Ohkuri1, Jay P. Slack2, Ping Zhong2, Liquan Huang 1 * 1 Monell Chemical Senses Center, Philadelphia, Pennsylvania, United States of America, 2 Givaudan Flavors Corporation, Cincinnati, Ohio, United States of America Abstract The sense of taste is important for providing animals with valuable information about the qualities of food, such as nutritional or harmful nature. Mammals, including humans, can recognize at least five primary taste qualities: sweet, umami (savory), bitter, sour, and salty. Recent studies have identified molecules and mechanisms underlying the initial steps of tastant-triggered molecular events in taste bud cells, particularly the requirement of increased cytosolic free Ca2+ concentration ([Ca2+] ) for normal taste signal transduction and transmission. Little, however, is known about the c mechanisms controlling the removal of elevated [Ca2+]c from the cytosol of taste receptor cells (TRCs) and how the disruption of these mechanisms affects taste perception. To investigate the molecular mechanism of Ca2+ clearance in TRCs, we sought the molecules involved in [Ca2+]c regulation using a single-taste-cell transcriptome approach. We found that Serca3, a member of the sarco/endoplasmic reticulum Ca2+-ATPase (SERCA) family that sequesters cytosolic Ca2+ into endoplasmic reticulum, is exclusively expressed in sweet/umami/bitter TRCs, which rely on intracellular Ca2+ release for signaling. Serca3-knockout (KO) mice displayed significantly increased aversive behavioral responses and greater gustatory nerve responses to bitter taste substances but not to sweet or umami taste substances. Further studies showed that Serca2 was mainly expressed in the T1R3-expressing sweet and umami TRCs, suggesting that the loss of function of Serca3 was possibly compensated by Serca2 in these TRCs in the mutant mice. Our data demonstrate that the SERCA family members play an important role in the Ca2+ clearance in TRCs and that mutation of these proteins may alter bitter and perhaps sweet and umami taste perception. Citation: Iguchi N, Ohkuri T, Slack JP, Zhong P, Huang L (2011) Sarco/Endoplasmic Reticulum Ca2+-ATPases (SERCA) Contribute to GPCR-Mediated Taste Perception. PLoS ONE 6(8): e23165. doi:10.1371/journal.pone.0023165 Editor: Hiroaki Matsunami, Duke University, United States of America Received January 25, 2011; Accepted July 13, 2011; Published August 2, 2011 Copyright: 2011 Iguchi et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was in part supported by Givaudan Flavors Corporation (http://www.givaudan.com/) and by grants from National Institutes of Health/ National Institute on Deafness and Other Communication Disorders (DC007487 to L.H.)(http://www.nidcd.nih.gov/) and from the National Science Foundation (DBJ-0216310 to N. Rawson)(http://www.nsf.gov/). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. No additional external funding received for this study. Competing Interests: NI and LH have declared that no competing interests exist. JPS and PZ are employees of Givaudan Flavors Corp. This does not alter the authors’ adherence to all the PLoS ONE policies on sharing data and materials. * E-mail: lhuang@monell.org Introduction The sense of taste is important to animal survival and growth; it not only plays a critical role in determining what should be ingested but also generates hedonic valence that can significantly affect an animal’s mood and mental activity [1]. Vertebrates, including humans, can recognize at least five primary taste qualities: sweet, umami, bitter, sour, and salty [2]. The interaction between the taste substances and receptors on the apical surface of taste bud cells initiates taste sensation, and the output signal from taste bud cells is relayed to the central nervous system via afferent gustatory nerves. Each taste bud consists of a small number of basal cells, which are presumed to be progenitor cells, and a heterogeneous population of 50–100 elongated cells that are classified as type I, II, or III based on their morphological, biochemical, and physiological properties [3]. Type I cells are suggested to be salt- taste-sensing cells, in addition to the long-standing presumption of a supportive function in the taste bud similar to that of glial cells in the nervous system [4,5,6,7]. Type II cells, also called taste ‘‘receptor’’ cells (TRCs), express molecular machinery for taste transduction for sweet, umami, and bitter, including G-protein-coupled taste receptors (GPCRs) and downstream effectors. Sweet and umami tastes are mediated by TRCs expressing GPCRs belonging to the T1R family: T1R2+T1R3 for sweet and T1R1+T1R3 for umami [8,9,10,11,12]. Bitter taste is mediated by another set of GPCRs, the T2R family [13,14,15,16]. Activation of T1R or T2R receptors stimulates G-proteins, which in turn activate phospholipase C (PLC) b2, leading to the production of the second messengers inositol-1,4,5-triphosphate (IP3) and diacyl- glycerol [17,18,19]. IP3 binds to its receptor, IP3R3, allowing the endoplasmic reticulum (ER) to release Ca2+, which increases cytosolic free Ca2+ concentrations ([Ca2+]c), triggering the opening of Trpm5 (transient receptor potential cation channel, subfamily M, member 5) [17,20,21]. Na+ influx through Trpm5 depolarizes the membrane potential, which leads to release of ATP onto gustatory afferent nerve fibers via hemichannels [22,23,24]. Type III cells, also called ‘‘presynaptic’’ cells, form prominent synapses with afferent nerve fibers and exhibit depolarization- dependent secretion of serotonin and norepinephrine; these cells are also responsible for sour taste sensation [25,26,27,28,29,30]. Although the signal transduction mechanisms for sour and salty taste qualities are not fully understood, studies have demonstrated that sour and salty taste stimuli also lead to increases in [Ca2+]c in responding taste cells [5,27,31]. Therefore, regardless of the modality of a taste substance, the activation of different taste signal PLoS ONE | www.plosone.org 1 August 2011 | Volume 6 | Issue 8 | e23165