Characterization of the "Gs-like" activity of Xlas

描述（由申请人提供）： 刺激性G蛋白（Gsalpha）的α亚基及其大变体XLalphas通过使用替代启动子和前mRNA剪接而衍生自GNAS 1。与Gsalpha不同，XLalphas显示有限的组织分布，包括在肾脏中的表达，并且在所有研究的组织中父系表达。尽管有不同的氨基末端结构域，这两种蛋白质是相同的，在很长一段羧基末端的氨基酸，包括几乎所有的结构特征Gsalpha。因此，XLalphas和Gsalpha在细胞中可能具有相似的功能特性，我们最近发表的结果与这一假设一致。鉴于Gsalpha在许多不同的生物反应中的重要性，XLalphas的“GS样”活性也可能在体内发挥重要作用。GNASI的Galpha编码外显子内的杂合失活突变与显示父母特异性遗传的多种表型相关，包括假性甲状旁腺功能减退症（PHP）、进行性骨发育不全（POH）和奥尔布赖特遗传性骨营养不良（AHO）。父系传播后，这些突变中的大多数被预测为破坏Gsalpha和XLalphas，这表明这些疾病中的一些，如POH的发病机制，可能涉及不仅缺乏Gsalpha，但也XLalphas。此外，在母系遗传后发生的疾病（例如PHP-Ia）中，从完整的父本等位基因表达的XLalphas活性可能在某些细胞中提供补偿性“GS样”信号传导，从而有助于某些类型的激素抵抗的选择性PHP。因此，我的主要目标是进一步探索XLalphas的“GS样”活性在这些疾病的分子和遗传机制中的作用。我的第一个具体目标涉及更详细的体外表征XLas，并将其功能特性与Gsalpha的功能特性进行比较，特别是关于可能的受体选择性和天然存在的GNASI突变的差异效应。我还将确定其信号活性的重要的独特的XLovias特定功能。此外，我将检查XLalphas在肾脏中的空间和时间表达，从而进一步了解XLas可能参与该组织中甲状旁腺激素介导的作用。最后，我将确定是否XLalphas可以有“GS样”的活动在体内，通过产生一个动物模型与靶向表达XLalphas在肾近曲小管。这些研究将有助于阐明XLalpha的生物学作用，并可能进一步提高目前对GNAS 1相关疾病的理解。

DESCRIPTION (provided by applicant): The alpha subunit of the stimulatory G protein (Gsalpha), and its large variant XLalphas, are derived from GNAS1 through the use of alternative promoters and pre-mRNA splicing. XLalphas, unlike Gsalpha, shows limited tissue distribution, including expression in kidney, and is expressed paternally in all investigated tissues. Despite having distinct amino-terminal domains, the two proteins are identical over a long stretch of carboxyl-terminal amino acids comprising almost all the structural features characterized for Gsalpha. Thus, XLalphas and Gsalpha may have similar functional properties in the cell, and our recently published results are consistent with this hypothesis. Given the importance of Gsalpha in numerous different biological responses, the "Gs-like" activity of XLalphas may also have significant roles in the body. Heterozygous inactivating mutations within Gsalpha-encoding exons of GNASI are associated with multiple phenotypes that show parent-specific inheritance, including pseudohypoparathyroidism (PHP), progressive osseous heteroplasia (POH), and Albright's hereditary osteodystrophy (AHO). After paternal transmission, most of these mutations are predicted to disrupt both Gsalpha and XLalphas, suggesting that pathogenesis of some of these disorders, such as POH, may involve not only the deficiency of Gsalpha but also of XLalphas. Moreover, in disorders that develop after maternal inheritance, such as PHP-Ia, the activity of XLalphas expressed from the intact paternal allele may provide compensatory "Gs-like" signaling in certain cells, thereby contributing to the selectivity of hormone resistance in certain types of PHP. My main objective is thus to further explore the role of the "Gs-like" activity of XLalphas in the molecular and genetic mechanisms underlying these diseases. My first specific aim involves a more detailed in vitro characterization of XLas, and to compare its functional properties with those of Gsalpha, particularly regarding possible receptor selectivity and differential effects of naturally-occurring GNASI mutations. I will also identify the unique XLalphas-specific features that are important for its signaling activity. In addition, I will examine spatial and temporal expression of XLalphas in the kidney to therefore provide further insights into possible involvement of XLas in actions mediated by parathyroid hormone in this tissue. Finally, I will determine whether XLalphas can have "Gs-like" activity in vivo, by generating an animal model with targeted expression of XLalphas in the renal proximal tubule. These studies will be helpful in clarifying the biological roles of XLalphas, and may furthermore improve the current understanding of the GNASl-related disorders.