MOLECULAR BIOLOGY OF P CARINII GP120

由卡氏肺孢子虫引起的肺炎是一种 获得性免疫缺陷患者死亡的主要原因 综合征和其他免疫功能低下的宿主。的长期目标 这项拟议的研究旨在鉴定卡氏肺孢子虫的抗原 参与对感染的后天抵抗力。患者或感染者 从卡氏肺炎(PCP)中康复的动物展示 对一种丰富的、免疫优势的表面糖蛋白的免疫应答 在PCP的雪貂模型中，它的MR为120 kDa (Gpl2O)。这个数据和其他数据表明，表面糖蛋白是 重要的寄主--P。隆突相互作用。当务之急是 本研究拟对雪貂P. Carinii GP 120。这项拟议的研究主要有三个方面。 几个重组的cDNA克隆，每个克隆编码一部分用于 卡氏雪貂P.carinii gpl2O是用特异性抗体分离出来的。 对本地雪貂P.carinii gpl2O的毒力。核苷酸序列分析 提示该克隆对应于编码基因的转录本 至少有两种不同的gpl2O亚型。这些结果支持 卡氏雪貂gpl2O基因多态的假说 分子中蛋白质核心的变异。为了测试这一点 假设，与预测的雪貂对应的全长cDNA 卡氏P.carinii gpl2O亚型将被克隆和测序。三、分析了 核苷酸序列将建立cDNAs的相互关系 克隆人。 最近建立的大鼠和人卡氏肺吸虫核型分析 这种遗传变异存在于卡氏肺孢子虫基因组的染色体上 水平。研究染色体异质性的可能性 雪貂gp120基因的组织结构和电泳核型 卡氏肺孢子虫将是 用脉冲场凝胶电泳法和GP L2O特异性 染色体将通过Southern杂交进行定位。 在真核和原核表达系统中表达后， 糖基化和非糖基化重组蛋白的抗原性 用免疫学方法将gpl2O与天然gpl2O进行比较。 卡氏雪貂P.carinii GP 120的分子特征将是必不可少的 了解其与宿主免疫系统相互作用的基础。 从拟议研究中获得的数据将有助于设计 PCP的诊断试剂和免疫疗法。

Pneumonia caused by the opportunistic pathogen Pneumocystis carinii is a primary cause of mortality in patients with acquired immunodeficiency syndrome and in other immunocompromised hosts. The long range goal of the proposed study is to identify antigens of P. carinii that are involved in acquired resistance to infection. Patients or infected animals that have recovered from P. carinii pneumonia (PCP) demonstrate an immune response to an abundant, immunodominant surface glycoprotein of P. carinii, which in the ferret model of PCP has an Mr of 120 kDa (gpl2O). This and other data indicate that the surface glycoprotein is important in the host-P. carinii interaction. The immediate focus of this proposal is to perform a molecular biological analysis of ferret P. carinii gp 120. There are three main aspects to the proposed study. Several recombinant cDNA clones, each encoding a portion of the cDNA for ferret P. carinii gpl2O, have been isolated using specific antibodies against native ferret P. carinii gpl2O. Nucleotide sequence analysis suggested that the cDNA clone's correspond to transcripts encoding at least two different isoforms of gpl2O. These results support the hypothesis that genetic polymorphism of ferret P. carinii gpl2O results in variation of the protein core of the molecule. To test this hypothesis, the full-length cDNAs corresponding to the predicted ferret P. carinii gpl2O isoforms will be cloned and sequenced. Analysis of the nucleotide sequences will establish the inter-relationship of the cDNA clones. Recent analyses of the karyotypes of rat and human P. carinii established that genetic variation exists in the P. carinii genome at the chromosomal level. To investigate the potential for heterogeneity in the chromosomal organization of the gp120 gene, the electrophoretic karyotype of ferret P. carinii will be determined by pulsed-field gel electrophoresis and the gp l2O-specific chromosomes will be mapped by Southern hybridization. After expression in eukaryotic and prokaryotic expression systems, the antigenic relatedness of glycosylated and non-glycosylated recombinant gpl2O to native gpl2O will be compared using immunological methods. Molecular characterization of ferret P. carinii gp 120 will be essential to understand the basis of its interaction with the host immune system. The data obtained from the proposed studies will contribute to the design of diagnostic reagents and immunotherapy for PCP.