Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Production of a severe cystic fibrosis mutation in mice by gene targeting

Abstract

We have used gene targeting in embryonic stem cells to introduce an HPRT mini–gene into the coding sequence of the murine cystic fibrosis gene (cftr). This insertion introduces a termination codon in frame with the cftr coding sequence to terminate prematurely the CFTR protein within the first nucleotide binding domain. Animals homozygous for the cftr disruption fail to thrive and display a range of symptoms including meconium ileus, distal intestinal obstructions, gastrointestinal mucus accumulation and blockage of pancreatic ducts. The animals also show lacrimal gland pathology. Tracheal and caecal transepithelial current measurements demonstrate the lack of a cAMP activatable Clchannel. These animals will prove useful for the evaluation of new therapeutic drugs and gene therapy strategies.

This is a preview of subscription content, access via your institution

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

Prices may be subject to local taxes which are calculated during checkout

Similar content being viewed by others

References

  1. Hodson, M., Norman, A.P. & Batten, J.C. eds Cystic Fibrosis (Bailliere Tindall, London, 1983).

    Google Scholar 

  2. Anderson, M.P. et al. Nucleoside triphosphates are required to open the CFTR chloride channel. Cell 67, 775–784 (1991).

    Article  CAS  PubMed  Google Scholar 

  3. Tsui, L.-C. The spectrum of cystic fibrosis mutations. Trends Genet. 8, 392–398 (1992).

    Article  CAS  PubMed  Google Scholar 

  4. Riordan, J.R. et al. Identification of the cystic fibrosis gene: cloning and characterisation of complementary DNA. Science 245, 1066–1073 (1989).

    Article  CAS  PubMed  Google Scholar 

  5. Kerem, B. et al. Identification of the cystic fibrosis gene:genetic analysis. Science 245, 1073–1080 (1989).

    Article  CAS  PubMed  Google Scholar 

  6. Colledge, W.H., Ratcliff, R., Foster, D., Williamson, R. & Evans, M.J. : Cystic fibrosis mouse with intestinal obstruction. Lancet 340, 680 (1992).

    Article  CAS  PubMed  Google Scholar 

  7. Snouwaert, J.N. et al. An animal model for cystic fibrosis made by gene targeting. Science 257, 1083–1088 (1992).

    Article  CAS  PubMed  Google Scholar 

  8. Dorin, J.R. et al. Cystic fibrosis in the mouse by targeted insertional mutation. Nature 359, 211–215 (1992).

    Article  CAS  PubMed  Google Scholar 

  9. Clarke, L.L. et al. Defective epithelial chloride transport in a gene targeted mouse model of cystic fibrosis. Science 257, 1125–1128 (1992).

    Article  CAS  PubMed  Google Scholar 

  10. Hunton, D.B., Long, W.K. & Tsumagari, H.Y. Meconium ileus equivalent: an adult complication of fibrocystic disease. Gasteroenterology 50, 99–107 (1966).

    CAS  Google Scholar 

  11. Hadorn, B. et al. Quantitative assessment of exocrine pancreatic function in infants and children. J. Pediatr. 73, 39–48 (1968).

    Article  CAS  PubMed  Google Scholar 

  12. Smith, S.N., Alton, E.W.F. & Geddas. D.M. Ion transport characteristics of the murine trachea. Clin. Sci. 82, 667–672 (1992).

    Article  CAS  Google Scholar 

  13. Kristidis, P. et al. Genetic determination of the exocrine pancreas function in cystic fibrosis. Am. J. hum. Genet. 50, 1178–1184 (1992).

    CAS  PubMed  PubMed Central  Google Scholar 

  14. Donnison, A.B., Shwachman, H. & Gross, R.E. A review of 164 children with meconium ileus seen at the Children's Hospital Medical Centre, Boston. Pediatrics 37, 833–843 (1966).

    CAS  PubMed  Google Scholar 

  15. Moens, C.B., Auerbach, A.B., Conlon, R.A., Joyner, A.L. & Rossant, J. A targeted mutation reveals a role for N-myc in branching morphogenesis in the embryonic mouse lung. Genes Devel. 6, 691–704 (1992).

    Article  CAS  PubMed  Google Scholar 

  16. Willumsen, N.J. & Boucher, R.C. Activation of an apical C1 conductance by Ca2+ ionophores in CF airway epithelia. Am. J. Physiol., 256, 226–233 (1989).

    Article  Google Scholar 

  17. Wagner, J.A. et al. Activation of chloride channels in normal and cystic fibrosis airway epithelial cells by multifunctional calcium/calmodulin dependent protein kinase. Nature 349, 793–796 (1991).

    Article  CAS  PubMed  Google Scholar 

  18. Anderson, M.P. & Welch, M.J. Calcium and cAMP activate different chloride channels in the apical membrance of normal and cystic fibrosis epithelia. Proc. Natl. Acad. Sci. U.S.A. 88, 6003–6007 (1991).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Wagner, J.A., et al. Antisense oligodeoxynucleotides to the cystic fibrosis transmembrane conductance regulator inhibit cAMP activated but not calcium-activated chloride currents. Proc. natn. Acad. Sci. U.S.A. 89, 6785–6789 (1992).

    Article  CAS  Google Scholar 

  20. Chang, S.H. et al. Defective intracellular transport and processing of CFTR is the molecular basis of most cystic fibrosis. Cell 63, 827–834 (1990).

    Article  Google Scholar 

  21. Gregory, R.J. et al. Maturation and function of cystic fibrosis transmembrane conductance regulator variants bearing mutations in putative nucleotide binding domains 1 and 2. Molec. cell Biol. 11, 3886–3893 (1991).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Kartner, N., Augustinos, O., Jensen, T.J., Naismith, A.L. & Riordan, J.R. Mislocalization of ΔF508 CFTR in cystic fibrosis sweat gland. Nature Genet. 1, 321–327 (1992).

    Article  CAS  PubMed  Google Scholar 

  23. Henderson, R.M., Ashford, M.L.J., MacVinish, L.J. & Cuthbert, A.W. Chloride channels and anion fluxes in a human colonic epithelium (HCA–7). Br. J. Pharmacol. 106, 109–114 (1992).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Egan, M. et al. Defective regulation of outwardly rectifying Cl- channels by protein kinase A corrected by insertion of CFTR. Nature 358, 581–584. (1992).

    Article  CAS  PubMed  Google Scholar 

  25. Boucher, R.C., Stutts, M.J., Knowles, M.R., Cantley, L. & Gatzy, J.T. Na+ transport in cystic fibrosis respiratory epithelia. Abnormal basal route and response to adenylate cyclase activations. J. clin. Invest. 78, 1245–1252 (1986).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Cotton, C.U., Knowles, M.R., Gatzy, J.T. & Boucher, R.C. Abnormal apical cell membrance in cystic fibrosis respirtory epithelium. An in vitro electrophysiologic analysis. J. clin. Invest. 79, 80–85 (1987).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Knowles, M.R., Spock, A., Fisher, N., Gatszy, J.T. & Boucher, R.C. Abnormal ion permeation through cystic fibrosis respiratory epithelium. Science 221, 1067–1070 (1983).

    Article  CAS  PubMed  Google Scholar 

  28. Tata, F. et al. Cloning the mouse homolog of the human cytic fibrosis transmembrance conductance regulator gene. Genomics 10, 301–307 (1991).

    Article  CAS  PubMed  Google Scholar 

  29. Van der Lugt, L., Maandag, E., te Riele, H., Laird, P.W. & Berns, A. A pgkhprt fusion as a selectable marker for targeting of genes in mouse embryonic stem cells:disruption of the T-Cell receptor delta-chain-encoding gene. Gene 105, 263–267 (1991).

    Article  CAS  PubMed  Google Scholar 

  30. Adra, C.N., Boer, P.M. & McBurney, M.W. Cloning and expression of the mouse pgk-1 gene and the nucleotide sequence of its promoter. Gene 60, 65–74 (1987).

    Article  CAS  PubMed  Google Scholar 

  31. Kuehn, M.R., Bradley, A., Robertson, E.J. & Evans, M.J. A potential animal model for Lesch-Nyhan sydrome through introduction of HPRT mutations into mice. Nature 326, 295–298 (1987).

    Article  CAS  PubMed  Google Scholar 

  32. Evans, M.J. Potential for genetic manipulation of mammals. Molec. biol. Med. 6, 557–565 (1989).

    CAS  PubMed  Google Scholar 

  33. Ratcliff, R. et al. Disruption of the cystic fibrosis transmembrace conductance regulator gene in embryonic stem cells by gene targeting. Trans. Res. 1, 177–181 (1992).

    Article  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Rights and permissions

Reprints and permissions

About this article

Cite this article

Ratcliff, R., Evans, M., Cuthbert, A. et al. Production of a severe cystic fibrosis mutation in mice by gene targeting. Nat Genet 4, 35–41 (1993). https://doi.org/10.1038/ng0593-35

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/ng0593-35

This article is cited by

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing