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Spontaneous transgenesis of human B lymphocytes

Gene Therapy volume 11, pages 42–51 (2004)Cite this article

Abstract

DNA can cross the cell membrane by natural means, but the functional relevance of this phenomenon has not been fully elucidated. Here, we analyzed spontaneous transgenesis of human B cells using plasmid DNA coding for a functional immunoglobulin (Ig) heavy chain gene under the control of a B-cell-specific promoter. Using polymerase chain reaction (PCR), reverse transcriptase-PCR, and flow cytometry in combination, spontaneous transgenesis was documented in Burkitt's lymphoma cell lines, Epstein–Barr virus-transformed cell lines, and peripheral blood B lymphocytes of the mature naïve phenotype (IgM+/IgD+/CD27−). By immunoelectron microscopy, the internalized DNA was seen in the lysosomes/late endosomes and in the cytosol proximal to the nucleus. Importantly, spontaneously transgenic B cells processed and presented to major histocompatibility complex (MHC)-restricted T lymphocytes a peptide expressed in the transgenic product. This is the first demonstration that primary B lymphocytes possess a program for the spontaneous internalization of DNA, which in turn imparts the cell with new immunological functions. As spontaneous transgenesis is obtained using a nonviral vector, does not require prior cell activation, and is not associated with chromosomal integration, the findings reported here open new possibilities for genetic manipulations of mature naïve B lymphocytes for therapy and vaccination.

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Acknowledgements

We are thankful to Drs MG Farquhar, W Loomis, and J Urbain for critical reading of the manuscript and their helpful suggestions. We also thank G Almanza and I LacKamp (Cosmo Bioscience) for their help in the performance of PCR work and A Necchi (University of Pavia) for assistance with the electron microscopy. This work was supported in part by a grant from the School of Medicine of the University of California, San Diego, and NIH Grants RO1CA77427 and R21AI49771 (to MZ). GF was a recipient of a short-term mobility grant from Consiglio Nazionale delle Ricerche and from the Compagnia di SanPaolo (Italy).

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Author notes

  1. G Filaci

    Present address: Division of Internal Medicine and Clinical Immunology, Department of InternalMedicine and Center of Excellence for Biomedical Research, University of Genova, Italy

  2. H-D Chang

    Present address: Deutsches Rheumaforschungszentrum, 10115, Berlin, Germany

  3. G Filaci and M Gerloni: The first two authors contributed equally to this work

Authors and Affiliations

  1. The Department of Medicine and Cancer Center, University of California, San Diego, La Jolla, CA, USA

    G Filaci, M Gerloni, M Rizzi, P Castiglioni, H-D Chang, MC Wheeler & M Zanetti

  2. Cosmo Bioscience, La Jolla, CA, USA

    M Gerloni

  3. The Department of Surgical and Morphological Sciences, Division of Human Pathology, University of Genova, Italy

    R Fiocca

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Filaci, G., Gerloni, M., Rizzi, M. et al. Spontaneous transgenesis of human B lymphocytes. Gene Ther 11, 42–51 (2004). https://doi.org/10.1038/sj.gt.3302132

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