NOVEL PAIR OF IMMUNOGLOBULIN LIKE RECEPTORS IN MICE

一个新的小鼠成对免疫球蛋白样受体（PIR）基因家族 是这些研究的重点。 原型全长的序列 cDNA克隆，PIR-A和PIR-B，预测I型跨膜蛋白， 75和87 kD，具有相似的胞外域，但不同的跨膜和 胞质区 预测的PIR-A蛋白具有相对短的 胞质尾区和跨膜区中的带电Arg残基， 表明与另外的跨膜蛋白结合， 形成信号转换单元。 相比之下，PIR-B蛋白具有 非极性跨膜区和相对较长的胞质尾区 具有两个基于酪氨酸的共有免疫受体抑制基序 （ITIM）。 预测的肽序列在五个方面是不变的 连续的PIR-B cDNA克隆，但所有7个PIR-A型cDNA不同 克隆 PIR-A和PIR-B的表达似乎仅限于B 淋巴细胞和骨髓细胞，其中两种基因均表达 同步 本研究旨在验证这一假设 PIR-A和PIR-B cDNA编码的受体蛋白与 配体结合特异性，但不同的细胞内信号传导 性质，使得PIR-A分子复合物可以具有活化性质， 潜在的，而PIR-B可能通过其ITIM样抑制潜力 图案 第一个实验旨在生产单克隆抗体， 针对PIR-A和PIR-B上的共同和独特表位的抗体， B分子，并使用这些抗体来确定其细胞 分布和相对表达水平，它们的结构和单位 PIR-交联对B细胞的作用， 巨噬细胞反应。 第二个实验将检查 PIR-B分子中ITIM样基序的功能潜力。 第三组实验旨在分离和测序基因组DNA， PIR-A和PIR-B克隆（i）以确定PIR-A和PIR-B克隆的遗传基础。 PIR-A基因的序列多样性，以及（ii）破坏PIR-A基因的单一序列多样性。 在胚胎干细胞中复制PIR-B基因以产生PIR-B敲除小鼠 不平衡的PIR-A活性 第四组实验将 检查代表性巨噬细胞和B细胞系中的PIR-A转录物 以确定它们是否表达单个或多个受体 成员 最后的实验旨在鉴定配体 对于这些PIR分子。 这些研究可能揭示了一个重要的 PIR-A和PIR-B受体在体液中的调节作用， 炎症和过敏反应。

A novel paired immunoglobulin-like receptor (PIR) gene family in mice is the focus of these studies. Sequences of the prototypic full length cDNA clones, PIR-A and PIR-B, predict type I transmembrane proteins of 75 and 87 kD with similar ectodomains, but distinctive transmembrane and cytoplasmic regions. The predicted PIR-A protein has a relatively short cytoplasmic tail and a charged Arg residue in the transmembrane region, suggesting association with an additional transmembrane protein(s) to form a signal transducing unit. In contrast, the PIR-B protein has a nonpolar transmembrane region and a relatively long cytoplasmic tail with two consensus immunoreceptor tyrosine-based inhibitory motifs (ITIM). The predicted peptide sequences are invariant in five consecutive PIR-B cDNA clones, but differ for all seven PIR-A type cDNA clones. PIR-A and PIR-B expression appears to be restricted to B lymphocytes and myeloid cells, wherein both genes are expressed simultaneously. The present studies are designed to test the hypothesis that the PIR-A and PIR-B cDNAs encode receptor proteins with related ligand-binding specificity, but different intracellular signaling properties so that the PIR-A molecular complex may have activating potential, while PIR-B may have inhibitory potential via its ITIM-like motifs. The first experiments are designed to produce monoclonal antibodies against common and distinctive epitopes on the PIR-A and PIR- B molecules and to use these antibodies to determine their cellular distribution and relative expression levels, their structure and unit composition, and the effects of PIR-crosslinkage on B cell and macrophage responses. The second experiments will examine the functional potential of the ITIM-like motifs in the PIR-B molecules. The third set of experiments is designed to isolate and sequence genomic PIR-A and PIR-B clones (i) to determine the genetic basis for the sequence diversity of the PIR-A genes and (ii) to disrupt the single copy PIR-B gene in embryonic stem cells to generate PIR-B knockout mice with unbalanced PIR-A activity. The fourth set of experiments will examine PIR-A transcripts in representative macrophage and B cell lines to determine whether they express single or multiple receptor members. The final experiments are designed to identify the ligand(s) for these PIR molecules. These studies may reveal an important regulatory role for the PIR-A and PIR-B receptors in humoral, inflammatory and allergic responses.