Cell type differences in activity of the Streptomyces bacteriophage phiC31 integrase.
Genomic integration by the Streptomyces bacteriophage C31 integrase is a promising tool for non-viral gene therapy of various genetic disorders. We investigated the C31 integrase recombination activity in T cell derived cell lines, primary T lymphocytes and CD34(+) haematopoietic stem cells in comparison to mesenchymal stem cells and cell lines derived from lung-, liver- and cervix-tissue. In T cell lines, enhanced long-term expression above control was observed only with high amounts of integrase mRNA. Transfections of C31 integrase plasmids were not capable of mediating enhanced long-term transgene expression in T cell lines. In contrast, moderate to high efficiency could be detected in human mesenchymal stem cells, human lung, liver and cervix carcinoma cell lines. Up to 100-fold higher levels of recombination product was found in C31 integrase transfected A549 lung than Jurkat T cells. When the C31 integrase activity was normalized to the intracellular integrase mRNA levels, a 16-fold difference was found. As one possible inhibitor of the C31 integrase, we found 3- to 5-fold higher DAXX levels in Jurkat than in A549 cells, which could in addition to other yet unknown factors explain the observed discrepancy of C31 integrase activity.
Maucksch
Christof
C
Department of Pediatrics, Ludwig-Maximilians-University, 80337 Munich, Germany.
Aneja
Manish Kumar
MK
Hennen
Elisabeth
E
Bohla
Alexander
A
Hoffmann
Florian
F
Elfinger
Markus
M
Rosenecker
Joseph
J
Rudolph
Carsten
C
D048868:Adaptor Proteins, Signal Transducing / Q000032:analysis; D001435:Bacteriophages / Q000201:enzymology*; D002460:Cell Line; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D056970:Co-Repressor Proteins; D015870:Gene Expression; D017930:Genes, Reporter; D006801:Humans; D019426:Integrases / Q000378:metabolism*; D019169:Jurkat Cells; D018832:Molecular Chaperones; D009687:Nuclear Proteins / Q000032:analysis; D010957:Plasmids / Q000235:genetics; D016133:Polymerase Chain Reaction; D011995:Recombination, Genetic; D013302:Streptomyces / Q000821:virology; D013601:T-Lymphocytes / Q000378:metabolism; D014162:Transfection
D048868:Adaptor Proteins, Signal Transducing; D056970:Co-Repressor Proteins; C106985:DAXX protein, human; D018832:Molecular Chaperones; D009687:Nuclear Proteins; D019426:Integrases