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Endogenous microRNA regulation suppresses transgene expression in hematopoietic lineages and enables stable gene transfer

Abstract

MicroRNAs (miRNAs) are small noncoding RNAs that regulate gene expression by repressing translation of target cellular transcripts. Increasing evidence indicates that miRNAs have distinct expression profiles and play crucial roles in numerous cellular processes, although the extent of miRNA regulation is not well known. By challenging mice with lentiviral vectors encoding target sequences of endogenous miRNAs, we show the efficiency of miRNAs in sharply segregating gene expression among different tissues. Transgene expression from vectors incorporating target sequences for mir-142-3p was effectively suppressed in intravascular and extravascular hematopoietic lineages, whereas expression was maintained in nonhematopoietic cells. This expression profile, which could not be attained until now, enabled stable gene transfer in immunocompetent mice, thus overcoming a major hurdle to successful gene therapy. Our results provide novel in situ evidence of miRNA regulation and demonstrate a new paradigm in vector design with applications for genetic engineering and therapeutic gene transfer.

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Figure 1: Endogenous miRNAs effectively de-target gene expression from human hematopoietic cells.
Figure 2: mir-142-3p mediates target mRNA degradation in hematopoietic cells in a robust and efficient manner.
Figure 3: In vivo expression pattern of miRNA-regulated lentiviral vectors.
Figure 4: mir-142-3p effectively segregates gene expression between hematopoietic and nonhematopoietic lineages throughout the organs of transgenic mice.
Figure 5: miRNA-regulated lentiviral vector enables stable gene transfer in immunocompetent mice.

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Notes

  1. NOTE: corrected online 28 April 2006 (details online). In the version of this article initially published online, an incorrect version of Figure 5 was used. The error has been corrected for all versions of the article.

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Acknowledgements

We thank E. Allievi and I. Benzoni (San Raffaele Conditional Mutagenesis Core Facility) for help in generating transgenic mice; B. Cullen for providing the miR-30aT, E. Hauben for human monocytes, F. Sanvito for histology, A. Cantore for technical assistance, and M. De Palma, A. Lombardo and D. Lillicrap for discussions. This work was supported by grants from Telethon (TIGET grant), the European Union (Projects LSHB-CT-2004-005276, RIGHT and LSHB-CT-2004-005242, CONSERT) and the Italian Ministry of Scientific Research (to L.N.). B.D.B. is the recipient of a Natural Science and Engineering Research Council of Canada fellowship.

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Correspondence to Luigi Naldini.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Fig. 1

Analysis of transgene expression in the peripheral blood of F1 TgN.PGK.GFP.142-3pT mice. (PDF 138 kb)

Supplementary Fig. 2

mir-142-3p sharply segregates gene expression between hematopoietic and nonhematopoietic lineages in the spleen of transgenic mice. (PDF 194 kb)

Supplementary Methods (PDF 52 kb)

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Brown, B., Venneri, M., Zingale, A. et al. Endogenous microRNA regulation suppresses transgene expression in hematopoietic lineages and enables stable gene transfer. Nat Med 12, 585–591 (2006). https://doi.org/10.1038/nm1398

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