关键词: 腺相关病毒, RAB蛋白, 囊泡运输, 呼吸道上皮细胞

Abstract: Objective To test whether type 2 adeno-associated virus(AAV2) is able to undergo vesicular sorting and traffic into the late endosome, and then evaluate the endosome-contained AAV2 possess different transduction activities, thus to imply the role of endosomal trafficking in the activation of AAV2 transduction in transformed human bronchial epithelial cells. Methods Using the RAB7 GFP fusion protein as the specific late endosome marker, its colocalization with AF568 fluorochrome labeled AAV2 was analyzed by confocal microscopy. The infected AAV2 was retrieved by gradient centrifugation and magnetic beads-based immunoisolation together with the endosome, and then subjected to the reinfection of IB3 cells. The efficiency of the AAV2 transgene expression was compared under the conditions without and with the modulation of RAB7 functions. Results It was found that a significant portion of the endocytosed AAV2 was able to traffic into the late endosome in IB3 cells. The AAV2 recovered from the late endosome exerted better transduction activities than the native viruses. Conclusion The results implied that the vesicular sorting of AAV2 into the late endosome might be an important step for the activation of the AAV mediated transgene expression.

Key words: adeno-associated virus, RAB proteins, vesicular trafficking, airway epithelial cells