RH PROTEIN AND THE RH BLOOD GROUP SYSTEM

RH家族由多种多孔膜蛋白组成，广泛存在于从最原始的单细胞黏液霉菌到人类的各种物种中。两个成员，可变的RHCED多肽和不变的RHAG糖蛋白，被限制在红细胞上，定义了临床上重要的RH血型系统。这些蛋白质具有重要但未知的功能，其突变基因的红细胞形态和生理学产物的变化证明了这一点。根据我们的初步研究，我们假设RHAG同系物在红系和非红系细胞中都具有保守的转运功能，而RHCED系列仅在红系细胞中具有相关或修饰的功能。我们将使用生物化学实验和基因操作的模型系统来检验这一假说。我们的长期目标是确定RH表达的分子细胞需求，阐明RH家族蛋白可能作为膜转运蛋白的功能，并揭示RHAG/RHCED在红细胞中单独和共同的功能作用。为实现这些目标，我们的具体目标如下。1)为了确定RH表达的分子细胞需求，我们将使用Rh零突变作为一个自然模型，并确定它们对细胞路由、膜插入和蛋白质相互作用的机制影响。2)为了研究可能的传输功能，将粘菌直系物RhgA作为一个简单的模型系统。我们将通过插入基因失活来评估RhgA的功能，并通过野生型RhgA和人RH蛋白的表达拯救来研究缺失突变体。候选配体将被寻找并在运输测试中进行测试。3)为了揭示RHAG/RHCED的个体和集体功能，我们将在小鼠中通过同源重组靶向干扰RHAG和RHAG同源同源物，并确定突变的红细胞表型。建议的研究将有助于确定RH蛋白家族长期寻求的功能作用，并为结构/功能关系提供新的见解。所获得的知识还将加深我们对与RH血型系统相关的临床问题的理解，使临床服务部门能够探索新的管理方法。

The RH family is composed of multiple polytopic membrane proteins present in a wide variety of species ranging from the most primitive unicellular slime mold to man. Two members, the variable RHCED polypeptides and the invariant RHAG glycoprotein, are restricted to red cells, defining the clinically important RH blood group system. These proteins have an important but unidentified function, as evidenced by changes in red cell morphology and physiology by products of their mutated genes. Based on our preliminary studies, we hypothesize that the RHAG homologs possess a conserved transport function in both erythroid and nonerythroid cells, while the RHCED series perform a related or modified function confined only to erythroid cells. We will examine this hypothesis by using model systems amenable to biochemical experimentation and genetic manipulation. Our long-term objectives are to define the molecular cellular requirement for RH expression, elucidate the function of the RH family proteins possibly as membrane transporters and unravel the individual and collective functional roles of RHAG/RHCED in red cells. To meet these goals, our specific aims are as follows. 1) To define the molecular cellular requirement for RH expression, we will use Rh-null mutations as a natural model and determine their mechanistic effects on cellular routing, membrane insertion and protein interaction. 2) To study the possible transport function, the slime mold ortholog RhgA will be employed as a simple model system. We will assess the function of RhgA by insertional gene inactivation and study the null mutants by expression rescue with wild type RhgA and human RH proteins. Candidate ligands will be sought and tested in transport assays. 3) To unravel the individual and collective functions of RHAG/RHCED, we will target-disrupt by homologous recombination the Rhced and Rhag orthologs in mice and determine the mutant red cell phenotypes. The proposed studies should lead to identification of the long-sought functional role of the RH family of proteins, and provide new insight into the structure/function relationships. The knowledge gained will also deepen our understanding of clinical problems associated with the RH blood group system, allowing the clinical service to explore new approaches for their management.