Cloning and functional analysis of taste receptors for sourness and saltiness

我们通过基因同源性和功能表达相结合的方法，从大鼠环状乳头的cDNA文库中鉴定了编码味觉酸味受体(MDEG1)的cDNA。当MDEG1在非洲爪哇卵母细胞中表达时，表现出质子门控阿米洛利敏感阳离子通道的特征。有趣的是，在相同pH条件下，食醋的主要成分--醋酸的酸刺激产生的内向电流比盐酸刺激的大。原位杂交显示MDEG1的mRNA仅定位于味蕾。MDEG1免疫反应阳性细胞位于味蕾细胞的细胞表面和顶端，尤其是被归类为味觉受体细胞的III型细胞。这是第一个被克隆的味觉受体基因。为了分离酸味受体的另一个亚基，我们筛选了一个大鼠环状乳头cDNA文库，并将其命名为MDEG2。免疫组织化学和电生理研究表明，MDEG2在单个味觉细胞顶端与MDIEG1形成异源低聚通道，并改变了MDEG1通道的脱敏作用。在不改变其他通道性质的情况下，MDEG2中Gly481的单一氨基酸取代显著改变了杂寡体的减敏速率。这些发现表明，MDEG2调节酸味受体的脱敏，有助于酸味适应，MDEG2中的Gly481是控制酸味感觉持续时间的关键氨基酸残基。

We have identifieda cDNA encoding a taste receptor for sourness (MDEG1) from a rat circumvallate papilla cDNA library by combined cDNA homology and functional expression approaches. When expressed in Xenopus laevis oocytes, MDEG1 showed characteristics of a proton-gated amiloride-sensitive cation channel. Interestingly, acidic stimulation by acetic acid, the chief ingredient of vinegar, generatedlarger inwardcurrents than that by hydrochloric acidat equal pH.In situ hybridization revealed that the mRNA of MDEG1 was exclusively localized in taste buds. MDEG1 immunoreactivity was found on the cell surface and in the apical portion of taste bud cells, especially in type III cells which are classifiedas gustatory receptor cells. This is the first taste receptor cDNA to be cloned. To isolate another subunit of the sour-taste receptor, we screened a rat circumvallate papilla cDNA library and iden tified MDEG2. Immunohistochemical and electrophysiological studies demonstrated that MDEG2 formed heterooligomeric channels with MDIEG1 at the apical portion of single taste-cells and modified the desensitization of MDEG1 channel. The desensitization rate of the heterooligomer was dramatically altered by a single amino acid substitution of Gly481 in MDEG2 without changing other channel properties. These findings suggest that MDEG2 regulates the sour-taste receptor desensitization contributing to sour-taste adaptation and that Gly481 in MDEG2 is the key amino acidresidue that controls the duration of sourtaste sensation.