UTEROGLOBULIN, CC10 KDA PROTEIN, PHOSPHOLIPASE A2, OSTEOPONTIN AND HIV-1 PROTEASE

子宫球蛋白、CC10 KDA 蛋白、磷脂酶 A2、骨桥蛋白和 HIV-1 蛋白酶

• 批准号: 3756687
• 负责人: A B MUKHERJEE
• 金额: --
• 依托单位: EUNICE KENNEDY SHRIVER NATIONAL INSTITUTE OF CHILD HEALTH & HUMAN DEVELOPMENT
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3756687
• 关键词: RNA splicing; X ray crystallography; enzyme activity; enzyme substrate; fibronectins; gene expression; globulins; human immunodeficiency virus 1; human tissue; neutralizing antibody; phospholipase A2; phospholipase inhibitor; phosphoproteins; posttranscriptional RNA processing; site directed mutagenesis; tissue /cell culture

We have:(a) demonstrated that Clara cell 10kDa (cc10kDa) protein is the counterpart of rabbit uteroglobin (rUG) by obtaining over expression of recombinant cc10kDa in E.coli and detailed characterization of this protein; (b) established that human UG (hUG) gene is expressed in many organs and tissues besides the pulmonary Clara cells suggesting that hUG may be a multifunctional cytokine-like protein; (c) delineated that hUG gene expression in the human endometrium is regulated by ovarian steroid hormones; (d) established and characterized two temperature-sensitive epithelial cell lines derived from the rabbit endometrium which differentiate in vitro and express UG gene in response to steroid hormones; (e) developed a highly improved expression vector which stabilizes the cDNA to be expressed in a bacterial host and requires no ampicillin for this stability; (f) crystallized recombinant hUG and carried out X-ray diffraction studies; (g) carried out multidimensional NMR studies on hUG which suggest a structure for this protein which is nearly identical to rUG; (h) site-directed mutagenesis of hUG suggests that mutation of Lys-43-> Asp 43 leads to inactivation of phospholipase A2-inhibitory property of hUG; (i) discovered a novel high affinity cell surface binding protein for hUG on NIH 3T3 and human trophoblast cells and when huG binds to this putative receptor it inhibits chemoinvasion of the artifiicial basement membrane by these cells. In addition, the choriocarcinoma cells were found to be lacking this receptor and their invasion is not affected by hUG; (j) established that invasion of prostatic carcinoma cells is suppressed by hUG; (k) identified a specific region of group I PLA2(residues 21-40) interaction of which with a neutralizing antibody leads to complete inhibition of its activity; (l) filed an invention report on the anti-invasive effects of hUG (m) established and standardized a radiometric assay for HIV-1 protease;(n) completed a preliminary study on the beneficial effects of hUG on a primate model of neonatal respiratory distress syndrome. The results suggest that recombinant hUG inhibits lung PLA2 activity and prevents migration of inflammatory cells into this organ. Long-term effects of this therapy are now being investigated (o) established that osteopontin (OP)-fibronectin interaction in bone and extracellular matrix (ECM) protein is mediated by transglutaminase (TG) and (p) demonstrated that disruption of cellular Src (c-Src) protooncogene causes dramatic suppression of OP gene expression. Since OP is essential for new bone formation by ostcoclas, our results may in part explain.

我们已经：(a) 证明 Clara 细胞 10kDa (cc10kDa) 蛋白是 通过获得兔子宫珠蛋白（rUG）的过度表达 大肠杆菌中的重组 cc10kDa 及其详细表征 蛋白质; (b) 确定人类 UG (hUG) 基因在许多细胞中表达 除肺 Clara 细胞外的器官和组织表明 hUG 可能是多功能细胞因子样蛋白； (c) 描述了 hUG 人类子宫内膜的基因表达受卵巢类固醇的调节 荷尔蒙； (d) 建立并表征了两种温度敏感的 来源于兔子宫内膜的上皮细胞系 体外分化并表达 UG 基因以响应类固醇 荷尔蒙； (e) 开发了一种高度改进的表达载体 稳定 cDNA 在细菌宿主中表达，不需要 氨苄西林用于这种稳定性； (f) 结晶重组 hUG 和 进行X射线衍射研究； (g) 多维开展 hUG 的 NMR 研究表明该蛋白质的结构是 与 rUG 几乎相同； (h) hUG 的定点诱变表明 Lys-43→Asp 43 的突变导致磷脂酶失活 hUG 的 A2 抑制特性； (i) 发现了一种新型高亲和力细胞 NIH 3T3 和人滋养层细胞上 hUG 的表面结合蛋白 当 huG 与这个假定的受体结合时，它会抑制化学侵袭 这些细胞形成人工基底膜。此外， 绒毛膜癌细胞被发现缺乏这种受体及其 侵袭不受hUG影响； (j) 确定入侵 hUG 抑制前列腺癌细胞； (k) 确定了具体的 I组PLA2（残基21-40）的区域，其中与 中和抗体导致其活性完全抑制； (l) 提交了hUG抗侵袭作用的发明报告(m) 建立并标准化了 HIV-1 蛋白酶的放射测定法；(n) 完成了关于 hUG 对 a 的有益影响的初步研究 新生儿呼吸窘迫综合征的灵长类动物模型。结果 表明重组 hUG 抑制肺 PLA2 活性并预防 炎症细胞迁移到该器官中。长期影响 目前正在研究这种疗法 (o) 确定骨桥蛋白 (OP)-纤连蛋白在骨和细胞外基质 (ECM) 中的相互作用 蛋白质由转谷氨酰胺酶 (TG) 介导，(p) 证明 细胞 Src (c-Src) 原癌基因的破坏会导致严重的后果 OP基因表达的抑制。由于 OP 对于新骨至关重要 ostcoclas 的形成，我们的结果可以部分解释。