Abstract：
We have adopted a real-time assay based on a dual-split reporter to assess cell-cell fusion mediated by the measles virus (MeV) membrane fusion machinery. This reporter system is comprised of two expression vectors, each encoding a segment of Renilla luciferase fused to a segment of GFP. To regain function, the two segments need to associate, which is dependent on cell-cell fusion between effector cells expressing the MeV fusion machinery and target cells expressing the corresponding MeV receptor. By measuring reconstituted luciferase activity, we can follow the kinetics of cell-cell fusion and quantify the extent of fusion. This assay lends itself to the study of the MeV fusion machinery comprised of the attachment and fusion glycoproteins, the matrix protein, and the MeV receptors. Moreover, entry inhibitors targeting attachment or fusion can be readily screened using this assay. Finally, this assay can be easily adopted to study the entry of other members of the Paramyxoviridae, as we have demonstrated for the henipaviruses.
摘要：
我们采用了基于双分裂报告子的实时测定法，以评估由麻疹病毒（MeV）膜融合机制介导的细胞-细胞融合。该报告系统由两个表达载体组成，每个编码与GFP片段融合的Renilla荧光素酶片段。要恢复功能，这两个部分需要关联，这依赖于表达MeV融合机制的效应细胞和表达相应MeV受体的靶细胞之间的细胞-细胞融合。通过测量重组的荧光素酶活性，我们可以跟踪细胞-细胞融合的动力学并量化融合的程度。该测定有助于研究由附着和融合糖蛋白组成的MeV融合机制，基质蛋白，和MeV受体。此外，使用该测定可以容易地筛选靶向附着或融合的进入抑制剂。最后,该测定法可以很容易地用于研究副粘病毒科其他成员的进入，正如我们已经证明的对流感病毒。
