Intracellular trafficking and fate of chimeric adenovirus 5/F35 in human B lymphocytes.

BACKGROUND

Investigation of the molecular processes that control the development and function of lymphocytes is essential for our understanding of humoral immunity, as well as lymphocyte-associated pathogenesis. Adenovirus-mediated gene transfer provides a powerful tool for investigating these processes. However, we observed variation in transgene expression among normal human peripheral blood B lymphocytes from different donors and at distinct stages of differentiation. It is recognized that efficient gene transfer is highly dependent on the intracellular route by which the viruses travel within the host cell. Thus, we aimed to examine this aspect in the present study.

METHODS

We analyzed the binding, uptake, intracellular trafficking and fate of CY3-labelled Ad5/F35 vectors in lymphoid cell lines and primary B cells. Furthermore, we decreased protein synthesis levels and rapid endocytosis in a plasma cell line exhibiting a high level of protein synthesis activity and activated transcription and endocytosis in primary B cells, which are less active than plasma cells.

RESULTS

Major differences in intracellular trafficking pattern between B cells and plasma cell line U266 were identified that explain the observed divergence in transgene expression efficiency. Importantly, modification of the transcriptional or translational activity of U266 cells reverted the Ad5/F35 endocytic trafficking to that seen in B cells, with a loss of transgene expression, whereas activation of B cells with phorbol 12-myristate 13-acetate had the opposite effects.

CONCLUSIONS

Taken together, these results suggest that Ad5/F35 is more efficiently transduced in cells with a strong transcriptional activity as a result of differences in intracellular trafficking. This finding extends our current knowledge of the mechanisms of adenovirus-mediated gene transfer.