MOLECULAR CHARACTERIZATION OF CR1 AND RELATED PROTEINS

拟议研究的总体目标将是表征， 在分子和生物学的方式中，编码 小鼠和人类(CR1)基因家族的成员。这个基因 家族包括CR1和CR2基因，以及之前的两个新的 未被描述的基因。此外，染色体的定位 将检查人类CR1与其他相关基因的关系 与CR1的氨基酸和核酸同源性。 小鼠基因组似乎至少包含四个CR1/CR1- 以mRNA转录本形式表达的相关基因 各种不同的组织。主要目标将是 克隆并至少部分测序，如果不是完全测序的话 四个基因，并从生化和生物学两个方面进行表征 由这些不同的基因编码的蛋白质。这些基因中的一个 由于其组织特异性表达，很可能是小鼠CR1， 人CR1的核酸同源性及其大小 文字记录。另外两个基因似乎不编码CR1或 CR2，而是在组织中表达的两种蛋白质，这表明 它们被DAF或gp45-70或其他未表征 蛋白质。CR1基因家族编码的蛋白质将是 通过以下方式进行鉴定：(I)使用所述cDNA序列来产生 在大肠杆菌中的重组蛋白，(Ii)使用该重组蛋白 制备抗肽序列的多克隆抗血清，(Iii) 分析由这些抗血清鉴定的小鼠蛋白，以及 假设CR1相关基因具有类似CR1/CR2的活性， (4)抗体阻断小鼠蛋白的功能 结合，以及(V)通过以下方式鉴定这些基因的蛋白质产物 利用整个基因在噬菌体或 余弦向量。一旦小鼠CR1相关基因被 人类同源基因也将被分离出来并 特色化的。小鼠CR1基因与小鼠的连锁 1号染色体上的SLE基因(GLD和NZB- 连锁SLE基因)也将被评估。 人CR1、C4BP和H的分子连锁，可能 小鼠CR1相关基因Y，On的人类同源物 人类1号染色体将识别第二个主要补体 蛋白质位点(MHC中的C4位点将是第一个) 基因组。与CR1相关的CR1基因的结构 染色体区域的遗传不稳定性 CR1基因图谱提示染色体不稳定的一种机制 我们将对此进行调查。

The overall goal of the proposed research will be to characterize, in a molecular and biological fashion, the genes encoding the members of the murine and human (CR1) gene family. This gene family include the CR1 and CR2 genes, and two new, previously undescribed gene. Additionally, the chromosomal location of human CR1 will be examined in relation to other linked genes sharing amino acid and nucleic acid homology to CR1. The mouse genome appears to contain at least four CR1/CR1- related genes which are expressed as mRNA transcripts in a variety of different tissues. The primary objective will be to clone and at least partially, if not completely, sequence these four genes and to characterize both biochemically and biologically the proteins encoded by these distinct genes. One of these genes is likely to be murine CR1 due to its tissue specific expression, nucleic acid homology to human CR1, and the size of it transcript. The other two genes appear not to encode CR1 or CR2, but instead two proteins expressed in tissues which suggest that they mat by either DAF or gp45-70, or other uncharacterized proteins. The proteins encoded by the CR1 gene family will be identified by: (i) using the cDNA sequences to produce recombinant protein in E. coli., (ii) using the recombinant protein to generate polyclonal antisera against the peptide sequence, (iii) analyzing the murine proteins identified by these antisera, and presuming the CR1-related genes share CR1/CR2-like activities, (iv) blocking the functions of the murine proteins by antibody binding, and (v) identifying the protein products of these genes by transfecting studies using the entire gene in either phage or cosmid vectors. Once the murine CR1-related genes have been identified, the human homologues will also be isolated and characterized. The linkage of the murine CR1 gene on murine chromosome 1 to SLE genes on chromosome 1 (gld and the NZB- linked SLE gene) will also be evaluated. The molecular linkage of human CR1, C4bp, and H, and perhaps the human homologue of the murine CR1-related gene Y, on human chromosome 1 will identify the second major complement protein locus (the C4 locus in the MHC would be the first) in the genome. The structure of the CR1 gene associated with the genetic instability of the region of the chromosome to which the CR1 gene maps suggests a mechanism for chromosomal instability which will be investigated.