Structural and functional studies on molecular evolution of calcium signaling tool kit

钙生物学领域的一个基本和具有挑战性的问题是，在单细胞和多细胞生物的进化中，钙信号机制和遗产可以追溯到多大程度？前些年，我们从克氏锥虫的寄生原生生物中鉴定了一个三磷酸肌醇受体(IP3R)的同源物，并证明了克氏锥虫IP3R(TcIP3R)是调节克氏锥虫复制、分化和侵染性的一种新的毒力因子。我们未来的研究将重点放在钙信号工具包的关键成员上，包括来自原始多细胞生物体-线虫的那些。在这个方案中，我们将研究人类基质相互作用蛋白(STIMs)和ORAI钙通道的原始同源物，这两个通道最近被证明在哺乳动物各种细胞类型的存储操作钙内流(SOCE)功能中发挥着关键作用。STIMs的两个亚型(STIM1和STIM2)作为内质网(ER)腔内钙感受器，激活ORAI钙通道，有三个亚型(ORAI1，2 Lcium信号已被用于各种生物体的生理功能)。我们最近对线虫STIM(CeSTIM)的结构研究表明，暴露在内质网管腔中的N-末端结构域在N-末端具有一个先前未发现的扭曲的螺旋，这可能解释了ceSTIM特异的基础ER钙水平下在点状细胞外围的结构性定位。我们也一直在研究线虫ORAI(CeORAI)和人类ORAI亚型，并成功地表达和纯化了ceORAI通道的多聚体功能形式。我们计划对ceSTIM和ceORAI进行进一步的结构和功能研究，以阐明线虫和哺乳动物之间与SOCE相关的共同和不同的机制。我们相信，我们对祖先IP3R、STIM和ORAI同源物的研究将为理解人类和其他哺乳动物中个体异构体的特定异构体角色提供重要线索，这些角色是作为基本生物功能从更简单的生物体继承到人类的，以及在高等生物体中哪些角色已经进化以适应更复杂的生存策略。

One of the fundamental and challenging questions in the calcium biology field is how far the calcium signaling mechanisms and legacy go back in the evolution of unicellular and multicellular organisms? In previous years, we identified an inositol 1,4,5-trisphosphate receptor (IP3R) homologue from a parasitic protist, Trypanosoma cruzi and demonstrated that T. cruzi IP3R (TcIP3R) is a novel virulence factor regulating replication, differentiation and infectivity of T. cruzi. Our future studies focus on key members of the calcium signaling toolkit, including those from a primitive multicellular organism, C. elegans. In this proposal we will study the primitive homologues of human stromal interacting proteins (STIMs) and ORAI Ca2+ channels, which were recently shown to play a crucial role in the store-operated calcium entry (SOCE) function in various cell types in mammals. Two isoforms of STIMs (STIM1 and STIM2) function as endoplasmic reticulum (ER) luminal Ca2+ sensors and activate ORAI Ca2+ channels, which have three isoforms (ORAI1, 2 lcium signals have been utilized for physiological functions of various organisms. Our recent structural studies on C.elegans STIM (ceSTIM) have shown that the N-terminal domain, which is exposed to the ER lumen, possesses a previously unidentified kinked helix at the N-terminus, which may explain the constitutive localization at the cell periphery in puncta under basal ER Ca2+ levels that is specific to ceSTIMs. We have also been studying C. elegans ORAI (ceORAI) as well as human ORAI isoforms and succeeded in expression and purification of the multimeric functional form of the ceORAI channel. We plan to perform further structural and functional studies on ceSTIM and ceORAI in order to elucidate common and diverged mechanisms associated with SOCE between C. elegans and mammals. We believe that our studies of ancestral IP3R, STIM and ORAI homologues will provide important clues to understanding the isoform-specific roles of the individual isoforms in humans and other mammals, which roles have been inherited from simpler organisms to humans as essential biological functions, and which roles have evolved in higher organisms to adapt to more complicated survival strategies.