NOVEL PAIR OF IMMUNOGLOBULIN LIKE RECEPTORS IN MICE

小鼠体内的一对新型免疫球蛋白样受体

• 批准号: 6510761
• 负责人: Hiromi Kubagawa
• 金额: $ 27.22万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-08-01 至 2003-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6510761
• 关键词: B lymphocyte; antibody receptor; antireceptor antibody; chimeric proteins; crosslink; gene targeting; laboratory mouse; laboratory rat; ligands; macrophage; monoclonal antibody; nucleic acid sequence; protein localization; protein structure function; receptor expression; tissue /cell culture; transfection

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.

一对新的小鼠免疫球蛋白样受体(PIR)基因家族 是这些研究的重点。原型全长序列 CDNA克隆PIR-A和PIR-B可预测I型跨膜蛋白 75 kD和87 kD具有相似的胞外结构域，但不同的跨膜和 细胞质区域。预测的PIR-A蛋白具有相对较短的 细胞质尾巴和跨膜区的带电Arg残基， 提示与另一种跨膜蛋白(S)有关 形成一个信号转换单元。相比之下，PIR-B蛋白具有 非极性跨膜区和相对较长的细胞质尾巴 具有两个共同的免疫受体酪氨酸抑制基序 (ITIM)。预测的多肽序列在五个中是不变的 连续的PIR-B cDNA克隆，但所有七个PIR-A型cDNAs不同 克隆人。PIR-A和PIR-B的表达似乎仅限于B 淋巴细胞和髓系细胞，其中这两个基因都有表达 同时。目前的研究旨在检验这一假设。 PIR-A和PIR-B的cDNA编码受体蛋白与相关 配体结合特异性，但细胞内信号不同 性质使PIR-A分子络合物具有活化性 而PIR-B可能通过其ITIM样蛋白具有抑制潜力 图案。第一批实验是用来生产单抗的。 针对PIR-A和PIR-A上共同和独特表位的抗体 B分子，并使用这些抗体来确定它们的细胞 分布和相对表达水平及其结构和单位 组成，以及PIR-交联剂对B细胞和 巨噬细胞反应。第二个实验将检验 PIR-B分子中类ITIM基序的功能潜能。 第三组实验的目的是分离和测序基因组 PIR-A和PIR-B克隆(I) PIR-A基因的序列多样性和(Ii)破坏单个 在胚胎干细胞中复制PIR-B基因以产生PIR-B基因敲除小鼠 具有不平衡的PIR-A活性。第四组实验将 在具有代表性的巨噬细胞和B细胞系中检测PIR-A转录本 以确定它们是表达单一还是多个受体 会员。最后的实验是为了确定配体(S) 对于这些PIR分子。这些研究可能揭示一个重要的 体液中PIR-A和PIR-B受体的调节作用 炎症和过敏反应。