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TRIM5? variations influence transduction efficiency with lentiviral vectors in both human and rhesus CD34(+) cells in vitro and in vivo.

ABSTRACT: Human immunodeficiency virus type 1 (HIV-1) vectors can transduce human hematopoietic stem cells (HSC), but transduction efficiency varies among individuals. The innate immune factor tripartite motif-containing protein 5? (TRIM5?) plays an important role for restriction of retroviral infection. In this study, we examined whether TRIM5? could account for variations in transduction efficiency using both an established rhesus gene therapy model and human CD34(+) cell culture. Evaluation of TRIM5? genotypes (Mamu-1, -2, -3, -4, -5, and TrimCyp) in 16 rhesus macaques that were transplanted with transduced CD34(+) cells showed a significant correlation between TRIM5? Mamu-4 and high gene marking in both lymphocytes and granulocytes 6 months after transplantation. Since significant human TRIM5? coding polymorphisms were not known, we evaluated TRIM5? expression levels in human CD34(+) cells from 14 donors. Three days after HIV-1 vector transduction, measured transduction efficiency varied significantly among donors and was negatively correlated with TRIM5? expression levels. In summary, transduction efficiency in both rhesus and human CD34(+) cells was influenced by TRIM5? variations (genotypes and expression levels). Our findings are important for both understanding and mitigating the variability of transduction efficiency for rhesus and human CD34(+) cells.

SUBMITTER: Evans ME

PROVIDER: S-EPMC3916046 | biostudies-literature | 2014 Feb

REPOSITORIES: biostudies-literature

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TRIM5α variations influence transduction efficiency with lentiviral vectors in both human and rhesus CD34(+) cells in vitro and in vivo.

Evans Molly E ME Kumkhaek Chutima C Hsieh Matthew M MM Donahue Robert E RE Tisdale John F JF Uchida Naoya N

Molecular therapy : the journal of the American Society of Gene Therapy 20131023 2

Human immunodeficiency virus type 1 (HIV-1) vectors can transduce human hematopoietic stem cells (HSC), but transduction efficiency varies among individuals. The innate immune factor tripartite motif-containing protein 5α (TRIM5α) plays an important role for restriction of retroviral infection. In this study, we examined whether TRIM5α could account for variations in transduction efficiency using both an established rhesus gene therapy model and human CD34(+) cell culture. Evaluation of TRIM5α g ...[more]

PMID: 24153115

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Project description:RNA interference (RNAi) is widely used in gene knockdown analysis and as a tool to screen host genes involved in viral infection. Owing to the limitations of transducing cells with synthetic small interfering RNAs (siRNAs), lentiviral short hairpin RNA (shRNA) vectors are more widely used. However, we found that stable transduction with lentiviral shRNA vectors inhibited hepatitis C virus (HCV) propagation in human hepatoma cells. We found by microRNA (miRNA) microarray analysis that this inhibition was induced by the alteration of host miRNA expression. In addition to one miRNA (miR-196b-5p) previously reported to be involved in HCV infection, other miRNAs (miR-216a-5p, -216b-5p, 217, and -30b-5p) were found to influence HCV infection in this study. Further studies suggested that this effect was independent of the transcription of shRNAs. The lentiviral vector itself and the integration site of the lentiviral vector might determine the change in miRNA expression. Moreover, the upregulation of JUN contributed to the dysregulation of miR-216a-5p, -216b-5p, and -217 in stably transduced cells. Although the changes in miRNA expression were beneficial for inhibiting HCV infection in our study, this off-target effect should be considered when transduction with lentiviral vectors is performed for other purposes, especially in therapy.IMPORTANCE We found that stable transduction with lentiviral shRNA was able to nonspecifically inhibit HCV infection by the dysregulation of host miRNAs. Previous studies showed that the overexpression of shRNAs oversaturated the host miRNA pathways to inhibit HCV infection. In contrast, the miRNA machinery was not affected in our study. Knockout studies suggested that the nonspecific effect was independent of the transcription of shRNAs. The lentiviral vector itself and the integration sites in the host genome determined the changes in miRNAs. Stable transduction with lentiviral vectors was able to increase the expression of JUN, which in turn upregulated miR-216a-5p, miR-216b-5p, and miR-217. miR-216a-5p and miR-216b-5p might inhibit HCV by suppressing the host autophagic machinery. Our study suggested a novel nonspecific effect of lentiviral vectors, and this side effect should be considered when transduction with lentiviral vectors is performed for other purposes, especially in therapy.

Dataset Information

TRIM5? variations influence transduction efficiency with lentiviral vectors in both human and rhesus CD34(+) cells in vitro and in vivo.

Publications

TRIM5α variations influence transduction efficiency with lentiviral vectors in both human and rhesus CD34(+) cells in vitro and in vivo.

Similar Datasets

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