Cell type differences in activity of the Streptomyces bacteriophage phiC31 integrase.
Cell type differences in activity of the Streptomyces bacteriophage phiC31 integrase.
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细胞类型在链霉菌噬菌体PHIC31积分酶活性的活性差异。
DOI：
10.1093/nar/gkn532
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发表时间：
2008-10
影响因子：
14.9
通讯作者：
Rudolph C
中科院分区：
文献类型：
--
作者：
Maucksch C;Aneja MK;Hennen E;Bohla A;Hoffmann F;Elfinger M;Rosenecker J;Rudolph C
链霉菌噬菌体C31整合酶的基因组整合是一种有前途的非病毒基因治疗各种遗传性疾病的工具。我们研究了与间充质干细胞和来自肺、肝和子宫颈组织的细胞系相比，CD31整合酶在T细胞衍生的细胞系、原代T淋巴细胞和CD34+造血干细胞中的重组活性。在T细胞系中，仅在高整合酶mRNA量的情况下观察到高于对照的增强的长期表达。转染的C31整合酶质粒不能够介导增强的长期转基因表达的T细胞系。相比之下，在人间充质干细胞、人肺、肝和宫颈癌细胞系中可以检测到中到高效率。高达100倍的重组产物的水平被发现比Jurkat T细胞在A549肺中的CD31整合酶转染。当将DAXX C31整合酶活性归一化为细胞内整合酶mRNA水平时，发现了16倍的差异。作为DAXX C31整合酶的一种可能的抑制剂，我们发现Jurkat中的DAXX水平比A549细胞高3至5倍，这可以解释除了其他未知因素之外观察到的DAXX C31整合酶活性的差异。
Genomic integration by the Streptomyces bacteriophage ϕC31 integrase is a promising tool for non-viral gene therapy of various genetic disorders. We investigated the ϕC31 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 ϕC31 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 ϕC31 integrase transfected A549 lung than Jurkat T cells. When the ϕC31 integrase activity was normalized to the intracellular integrase mRNA levels, a 16-fold difference was found. As one possible inhibitor of the ϕC31 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 ϕC31 integrase activity.
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