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The SP-C promoter facilitates alveolar type II epithelial cell-specific plasmid nuclear import and gene expression

Gene Therapy volume 17, pages 541–549 (2010)Cite this article

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Abstract

Although nonviral gene therapy has great potential for use in the lung, the relative lack of cell-specific targeting has limited its applications. We have developed a new approach for cell-specific targeting based on selective nuclear import of plasmids in nondividing cells. Using a microinjection and in situ hybridization approach, we tested several potential DNA sequences for the ability to mediate plasmid nuclear import in alveolar type II epithelial (ATII) cells. Of these, only a sequence within the human surfactant protein C (SP-C) promoter was able to mediate nuclear localization of plasmid DNA specifically in ATII cells but not in other cell types. We have mapped the minimal import sequence to the proximal 318 nucleotides of the promoter, and demonstrate that binding sites for nuclear factor I, thyroid transcriptional factor 1, and GATA-binding protein 6 and the proteins themselves are required for import activity. Using intratracheal delivery of DNA followed by electroporation, we demonstrate that the SP-C promoter sequence will enhance gene expression specifically in ATII cells in mouse lung. This represents a new activity for the SP-C promoter, and thus ATII cell-specific nuclear import of DNA may prove to be a safe and effective method for targeted and enhanced gene expression in ATII cells.

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References

  1. Dean DA . Import of plasmid DNA into the nucleus is sequence specific. Exp Cell Res 1997; 230: 293–302.

    Article  CAS  PubMed Central  Google Scholar 

  2. Langle-Rouault F, Patzel V, Benavente A, Taillez M, Silvestre N, Bompard A et al. Up to 100-fold increase of apparent gene expression in the presence of Epstein-Barr virus oriP sequences and EBNA1: implications of the nuclear import of plasmids. J Virol 1998; 72: 6181–6185.

    CAS  PubMed  PubMed Central  Google Scholar 

  3. Dean DA, Dean BS, Muller S, Smith LC . Sequence requirements for plasmid nuclear entry. Exp Cell Res 1999; 253: 713–722.

    Article  CAS  PubMed Central  Google Scholar 

  4. Vacik J, Dean BS, Zimmer WE, Dean DA . Cell-specific nuclear import of plasmid DNA. Gene Ther 1999; 6: 1006–1014.

    Article  CAS  PubMed Central  Google Scholar 

  5. Mesika A, Grigoreva I, Zohar M, Reich Z . A regulated, NFkappaB-assisted import of plasmid DNA into mammalian cell nuclei. Mol Ther 2001; 3: 653–657.

    Article  CAS  PubMed Central  Google Scholar 

  6. Goncalves C, Ardourel MY, Decoville M, Breuzard G, Midoux P, Hartmann B et al. An optimized extended DNA kappa B site that enhances plasmid DNA nuclear import and gene expression. J Gene Med 2009; 11: 401–411.

    Article  CAS  Google Scholar 

  7. Wilson GL, Dean BS, Wang G, Dean DA . Nuclear import of plasmid DNA in digitonin-permeabilized cells requires both cytoplasmic factors and specific DNA sequences. J Biol Chem 1999; 274: 22025–22032.

    Article  CAS  PubMed Central  Google Scholar 

  8. Colin M, Moritz S, Fontanges P, Kornprobst M, Delouis C, Keller M et al. The nuclear pore complex is involved in nuclear transfer of plasmid DNA condensed with an oligolysine-RGD peptide containing nuclear localisation properties. Gene Ther 2001; 8: 1643–1653.

    Article  CAS  Google Scholar 

  9. Miller AM, Dean DA . Tissue-specific and transcription factor-mediated nuclear entry of DNA. Adv Drug Deliv Rev 2009; 61: 603–613.

    Article  CAS  Google Scholar 

  10. Carson JA, Fillmore RA, Schwartz RJ, Zimmer WE . The smooth muscle gamma-actin gene promoter is a molecular target for the mouse bagpipe homologue, mNkx3-1, and serum response factor. J Biol Chem 2000; 275: 39061–39072.

    Article  CAS  PubMed Central  Google Scholar 

  11. Browning CL, Culberson DE, Aragon IV, Fillmore RA, Croissant JD, Schwartz RJ et al. The developmentally regulated expression of serum response factor plays a key role in the control of smooth muscle-specific genes. Dev Biol 1998; 194: 18–37.

    Article  CAS  Google Scholar 

  12. Miller AM, Dean DA . Cell-specific nuclear import of plasmid DNA in smooth muscle requires tissue-specific transcription factors and DNA sequences. Gene Ther 2008; 15: 1107–1115.

    Article  CAS  PubMed Central  Google Scholar 

  13. Wikenheiser KA, Vorbroker DK, Rice WR, Clark JC, Bachurski CJ, Oie HK et al. Production of immortalized distal respiratory epithelial cell lines from surfactant protein C/simian virus 40 large tumor antigen transgenic mice. Proc Natl Acad Sci USA 1993; 90: 11029–11033.

    Article  CAS  Google Scholar 

  14. Glasser SW, Korfhagen TR, Wert SE, Bruno MD, McWilliams KM, Vorbroker DK et al. Genetic element from human surfactant protein SP-C gene confers bronchiolar-alveolar cell specificity in transgenic mice. Am J Physiol 1991; 261: L349–L356.

    CAS  PubMed  Google Scholar 

  15. Kelly SE, Bachurski CJ, Burhans MS, Glasser SW . Transcription of the lung-specific surfactant protein C gene is mediated by thyroid transcription factor 1. J Biol Chem 1996; 271: 6881–6888.

    Article  CAS  Google Scholar 

  16. Glasser SW, Burhans MS, Eszterhas SK, Bruno MD, Korfhagen TR . Human SP-C gene sequences that confer lung epithelium-specific expression in transgenic mice. Am J Physiol Lung Cell Mol Physiol 2000; 278: L933–L945.

    Article  CAS  Google Scholar 

  17. Bachurski CJ, Kelly SE, Glasser SW, Currier TA . Nuclear factor I family members regulate the transcription of surfactant protein-C. J Biol Chem 1997; 272: 32759–32766.

    Article  CAS  Google Scholar 

  18. Bachurski CJ, Yang GH, Currier TA, Gronostajski RM, Hong D . Nuclear factor I/thyroid transcription factor 1 interactions modulate surfactant protein C transcription. Mol Cell Biol 2003; 23: 9014–9024.

    Article  CAS  PubMed Central  Google Scholar 

  19. Korfhagen TR, Glasser SW, Wert SE, Bruno MD, Daugherty CC, McNeish JD et al. Cis-acting sequences from a human surfactant protein gene confer pulmonary-specific gene expression in transgenic mice. Proc Natl Acad Sci USA 1990; 87: 6122–6126.

    Article  CAS  PubMed Central  Google Scholar 

  20. Liu C, Glasser SW, Wan H, Whitsett JA . GATA-6 and thyroid transcription factor-1 directly interact and regulate surfactant protein-C gene expression. J Biol Chem 2002; 277: 4519–4525.

    Article  CAS  PubMed Central  Google Scholar 

  21. Dean DA, Machado-Aranda D, Blair-Parks K, Yeldandi AV, Young JL . Electroporation as a method for high-level non-viral gene transfer to the lung. Gene Ther 2003; 10: 1608–1615.

    Article  CAS  PubMed Central  Google Scholar 

  22. Young JL, Benoit JN, Dean DA . Effect of a DNA nuclear targeting sequence on gene transfer and expression of plasmids in the intact vasculature. Gene Ther 2003; 10: 1465–1470.

    Article  CAS  PubMed Central  Google Scholar 

  23. Young JL, Zimmer WE, Dean DA . Smooth muscle-specific gene delivery in the vasculature based on restriction of DNA nuclear import. Exp Biol Med 2008; 233: 840–848.

    Article  CAS  Google Scholar 

  24. Miller AM, Munkonge FM, Alton EW, Dean DA . Identification of Protein Cofactors Necessary for Sequence-specific Plasmid DNA Nuclear Import. Mol Ther 2009; 17: 1897–1903; .

    Article  CAS  PubMed Central  Google Scholar 

  25. Bruno MD, Bohinski RJ, Huelsman KM, Whitsett JA, Korfhagen TR . Lung cell-specific expression of the murine surfactant protein A (SP-A) gene is mediated by interactions between the SP-A promoter and thyroid transcription factor-1. J Biol Chem 1995; 270: 6531–6536.

    Article  CAS  Google Scholar 

  26. Bohinski RJ, Di Lauro R, Whitsett JA . The lung-specific surfactant protein B gene promoter is a target for thyroid transcription factor 1 and hepatocyte nuclear factor 3, indicating common factors for organ-specific gene expression along the foregut axis. Mol Cell Biol 1994; 14: 5671–5681.

    Article  CAS  PubMed Central  Google Scholar 

  27. Christophe-Hobertus C, Duquesne V, Pichon B, Roger PP, Christophe D . Critical residues of the homeodomain involved in contacting DNA bases also specify the nuclear accumulation of thyroid transcription factor-1. Eur J Biochem 1999; 265: 491–497.

    Article  CAS  Google Scholar 

  28. Park KS, Whitsett JA, Di Palma T, Hong JH, Yaffe MB, Zannini M . TAZ interacts with TTF-1 and regulates expression of surfactant protein-C. J Biol Chem 2004; 279: 17384–17390.

    Article  CAS  Google Scholar 

  29. Gill DR, Smyth SE, Goddard CA, Pringle IA, Higgins CF, Colledge WH et al. Increased persistence of lung gene expression using plasmids containing the ubiquitin C or elongation factor 1alpha promoter. Gene Ther 2001; 8: 1539–1546.

    Article  CAS  Google Scholar 

  30. Yew NS, Przybylska M, Ziegler RJ, Liu D, Cheng SH . High and sustained transgene expression in vivo from plasmid vectors containing a hybrid ubiquitin promoter. Mol Ther 2001; 4: 75–82.

    Article  CAS  Google Scholar 

  31. Gossen M, Bujard H . Tight control of gene expression in mammalian cells by tetracycline-responsive promoters. Proc Natl Acad Sci USA 1992; 89: 5547–5551.

    Article  CAS  Google Scholar 

  32. Brunner S, Sauer T, Carotta S, Cotten M, Saltik M, Wagner E . Cell cycle dependence of gene transfer by lipoplex, polyplex and recombinant adenovirus. Gene Ther 2000; 7: 401–407.

    Article  CAS  Google Scholar 

  33. Escriou V, Carriere M, Bussone F, Wils P, Scherman D . Critical assessment of the nuclear import of plasmid during cationic lipid-mediated gene transfer. J Gene Med 2001; 3: 179–187.

    Article  CAS  Google Scholar 

  34. Berglund L, Bjorling E, Oksvold P, Fagerberg L, Asplund A, Szigyarto CA et al. A genecentric human protein Atlas for expression profiles based on antibodies. Mol Cell Proteomics 2008; 7: 2019–2027.

    Article  CAS  Google Scholar 

  35. Park KS, Korfhagen TR, Bruno MD, Kitzmiller JA, Wan H, Wert SE et al. SPDEF regulates goblet cell hyperplasia in the airway epithelium. J Clin Invest 2007; 117: 978–988.

    Article  CAS  PubMed Central  Google Scholar 

  36. Gasiorowski JZ, Dean DA . Postmitotic nuclear retention of episomal plasmids is altered by DNA labeling and detection methods. Mol Ther 2005; 12: 460–467.

    Article  CAS  PubMed Central  Google Scholar 

  37. Kouretas D, Karinch AM, Rishi A, Melchers K, Floros J . Conservation analysis of rat and human SP-A gene identifies 5′ flanking sequences of rat SP-A that bind rat lung nuclear proteins. Exp Lung Res 1993; 19: 485–503.

    Article  CAS  Google Scholar 

  38. Bohinski RJ, Huffman JA, Whitsett JA, Lattier DL . Cis-active elements controlling lung cell-specific expression of human pulmonary surfactant protein B gene. J Biol Chem 1993; 268: 11160–11166.

    CAS  PubMed  Google Scholar 

  39. Rust K, Bingle L, Mariencheck W, Persson A, Crouch EC . Characterization of the human surfactant protein D promoter: transcriptional regulation of SP-D gene expression by glucocorticoids. Am J Respir Cell Mol Biol 1996; 14: 121–130.

    Article  CAS  Google Scholar 

  40. Oshima RG, Baribault H, Caulin C . Oncogenic regulation and function of keratins 8 and 18. Cancer Metastasis Rev 1996; 15: 445–471.

    Article  CAS  PubMed Central  Google Scholar 

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Acknowledgements

We thank J Gasiorowski for technical advice and members of the Dean lab for stimulating discussions. We also thank L Davies (Oxford) for help with enhanced green fluorescent protein (EGFP) visualization in cryosections. This study was supported in part by National Institutes of Health grants EB00903 and HL81148 and a predoctoral training fellowship from the Midwest Affiliate of the American Heart Association (JVD).

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Authors and Affiliations

  1. Division of Pulmonary and Critical Care Medicine, Feinberg School of Medicine, Northwestern University, Chicago, IL, USA

    J V DeGiulio & D A Dean

  2. Division of Neonatology, School of Medicine and Dentistry, University of Rochester, Rochester, NY, USA

    C D Kaufman & D A Dean

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  1. J V DeGiulio
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  3. D A Dean
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Correspondence to D A Dean.

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DeGiulio, J., Kaufman, C. & Dean, D. The SP-C promoter facilitates alveolar type II epithelial cell-specific plasmid nuclear import and gene expression. Gene Ther 17, 541–549 (2010). https://doi.org/10.1038/gt.2009.166

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