MECHANISMS INVOLVED IN THE INTRACELLULAR RETENTION OF A2C ADRENERGIC RECEPTOR

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得主要资金， 因此可以在其他CRISP条目中表示。所列机构为 中心，不一定是研究者的机构。 最近的实验证据表明，α 2C肾上腺素能受体（AR）在雷诺现象（RP）中的重要作用。这种受体亚型在37 ℃时在质膜上表达很少，在内质网（ER）和高尔基体中积累。冷暴露后，α 2C-AR向细胞表面的运输大大增强。结构分析表明，α 2C-AR在第三胞内环和C-末端具有不寻常的高数量的精氨酸（R）残基，组织在11个推定的精氨酸分选基序（RXR）中。当嵌入其他蛋白质中时，该RXR基序已显示诱导ER滞留。我们的初步实验表明，从第三个细胞内环中删除8个假定的RXR基序大大增强了30 ℃时α 2C-AR的转运。基于这一观察，我们的总体假设是，独特的结构基序在α 2C-AR调节其运输。该实验计划旨在区分ER阻滞和高尔基体至质膜转运受阻的贡献，并通过鉴定赋予α 2C-AR转运温度敏感性的RXR基序并确定这些效应中涉及的分子机制，阐明α 2C运输调制中涉及的机制。此外，将研究Rab 8和Rab 14 GTP酶在温度敏感性α 2C-AR受体质膜表达中的作用。这些研究将产生关于α 2C-AR细胞内蓄积的分子决定因素的新的和重要的信息，并可能为设计雷诺现象的更有效的治疗策略提供基础。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Recent experimental evidence indicates an important role of alpha2C adrenergic receptor (AR) in Raynaud Phenomenon (RP). This receptor subtype is poorly expressed at the plasma membrane at 37oC, accumulating in endoplasmic reticulum (ER) and Golgi apparatus. Alpha2C-AR traffic to the cell surface is greatly enhanced after cold exposure. Structural analysis indicate that alpha2C-AR has an unusual high number of arginine (R) residues in the third intracellular loop and in the C-terminus, organized in eleven putative arginine sorting motifs (RXR). When embedded in other proteins, this RXR motif has been shown to induce ER retention. Our preliminary experiments demonstrated that deletion of the eight putative RXR motifs from the third intracellular loop greatly enhanced alpha2C-AR transport at 30oC. Based on this observation, our overall hypothesis is that unique structural motifs in alpha2C-AR regulate its trafficking. The experimental plan aims to distinguish between the contributions of ER arrest and obstruction of transport from Golgi to the plasma membrane and it will elucidate the mechanisms involved in the alpha2C traffic modulation by identifying the RXR motifs conferring temperature sensitivity to alpha2C-AR transport and determining the molecular mechanisms involved in these effects. Further, the role of Rab8 and Rab14 GTPases in the temperature sensitive alpha2C-AR receptor plasma membrane expression will be studied. These studies will produce novel and important information regarding the molecular determinants of alpha2C-AR intracellular accumulation and may provide foundation for designing more effective therapeutic strategies in Raynaud Phenomenon.