Tissue distribution of products of the mouse decay-accelerating factor (DAF) genes.: Exploitation of a Daf1 knock-out mouse and site-specific monoclonal antibodies
Tissue distribution of products of the mouse decay-accelerating factor (DAF) genes.: Exploitation of a Daf1 knock-out mouse and site-specific monoclonal antibodies
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DOI：
10.1046/j.1365-2567.2001.01287.x
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发表时间：
2001-10-01
期刊：
影响因子：
6.4
通讯作者：
Medof, ME
中科院分区：
文献类型：
--
作者：
Lin, F;Fukuoka, Y;Medof, ME
衰变加速因子（Decay-Accelerating Factor，缩写为ERF）是一种C3活化的膜调节因子，可保护自身细胞免受自体补体攻击。在人类中，β-淀粉样蛋白以糖基磷脂酰肌醇（GPI）锚定分子的形式均匀表达。在小鼠中，产生GPI锚定和跨膜锚定的GPI蛋白，每种蛋白都可以来自两种不同的基因（Daf 1和Daf 2）。在这份报告中，我们描述了一个Daf 1基因敲除小鼠产生的第一个产品的战略，针对一个或两个Daf基因。作为工作的一部分，我们最近描述的单克隆抗体对小鼠的DAF蛋白使用在酵母中合成的缺失突变体的特征，然后采用单克隆抗体与野生型和DAF 1基因敲除小鼠，以确定小鼠DAF 1和DAF 2基因产物的组织分布。为了提高免疫组化检测鼠抗人CD 4蛋白的灵敏度，我们采用了灵敏的酪胺荧光法。在野生型小鼠中，我们发现肾小球、气道和肠上皮、脾脏、所有组织中的血管内皮和睾丸的生精小管都有强烈的荧光标记。在Daf 1基因敲除小鼠中，在大多数组织中，标记被消融，但睾丸和脾脏树突状细胞的强标记仍然存在。在这两个网站，逆转录-聚合酶链反应分析确定了GPI和跨膜形式的Daf 2基因衍生的蛋白质。该结果与体内鼠β功能和鼠β结构的研究有关。
Decay-accelerating factor (DAF) is a membrane regulator of C3 activation that protects self cells from autologous complement attack. In humans, DAF is uniformly expressed as a glycosylphosphatidylinositol (GPI)-anchored molecule. In mice, both GPI-anchored and transmembrane-anchored DAF proteins are produced, each of which can be derived from two different genes (Daf1 and Daf2). In this report, we describe a Daf1 gene knock-out mouse arising as the first product of a strategy for targeting one or both Daf genes. As part of the work, we characterize recently described monoclonal antibodies against murine DAF protein using deletion mutants synthesized in yeast, and then employ the monoclonal antibodies in conjunction with wild-type and the Daf1 knock-out mice to determine the tissue distribution of the mouse Daf1 and Daf2 gene products. To enhance the immunohistochemical detection of murine DAF protein, we utilized the sensitive tyramide fluorescence method. In wild-type mice, we found strong DAF labelling of glomeruli, airway and gut epithelium, the spleen, vascular endothelium throughout all tissues, and seminiferous tubules of the testis. In Daf1 knock-out mice, DAF labelling was ablated in most tissues, but strong labelling of the testis and splenic dendritic cells remained. In both sites, reverse transcription-polymerase chain reaction analyses identified both GPI and transmembrane forms of Daf2 gene-derived protein. The results have relevance for studies of in vivo murine DAF function and of murine DAF structure.