Insertion of Inhbb into the Inhba locus rescues the Inhba-null phenotype and reveals new activin functions

Abstract

The activins (dimers of βA or βB subunits, encoded by the genes Inhba and Inhbb, respectively) are TGF-β superfamily members that have roles in reproduction and development1,2,3. Whereas mice homozygous for the Inhba-null allele demonstrate disruption of whisker, palate and tooth development, leading to neonatal lethality4,5, homozygous Inhbb-null mice are viable, fertile and have eye defects6,7. To determine if these phenotypes were due to spatiotemporal expression differences of the ligands or disruption of specific ligand-receptor interactions, we replaced the region of Inhba encoding the mature protein with Inhbb, creating the allele Inhbatm2Zuk (hereafter designated InhbaBK). Although the craniofacial phenotypes of the Inhba-null mutation were rescued by the InhbaBK allele, somatic, testicular, genital and hair growth were grossly affected and influenced by the dosage and bioactivity of the allele. Thus, functional compensation within the TGF-β superfamily can occur if the replacement gene is expressed appropriately. The novel phenotypes in these mice further illustrate the usefulness of insertion strategies for defining protein function.

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Figure 1: Generation of InhbaBK mice.
Figure 2: In situ hybridization of mouse ovary and embryo sections with βA and βB probes.
Figure 3: RNase protection assay and western-blot analysis.
Figure 4: Phenotypic analysis of wild-type and mutant mice.
Figure 5: Growth characteristics, survival and serum analysis of wild-type and mutant mice.

References

  1. 1

    Vale, W., Bilezikjian, L.M. & Rivier, C. Reproductive and other roles of inhibins and activins. in The Physiology of Reproduction (eds Knobil, E., Neill, J.D., Greenswald, G.S., Markert, C.L. & Pfaff, D.W.) 1861–1878 (Raven, New York, 1994).

  2. 2

    Lau, A.L., Shou, W., Guo, Q. & Matzuk, M.M. Transgenic approaches to study the functions of the transforming growth factor-b superfamily members. in Inhibin, Activin and Follistatin: Regulatory Functions in System and Cell Biology (eds Aono, T., Sugino, H. & Vale, W.) 220–243 (Springer, New York, 1997).

  3. 3

    Mather, J.P., Moore, A. & Li, R.H. Activins, inhibins, and follistatins: further thoughts on a growing family of regulators. Proc. Soc. Exp. Biol. Med. 215, 209–222 (1997).

  4. 4

    Matzuk, M.M. et al. Functional analysis of activins during mammalian development. Nature 374, 354–356 (1995).

  5. 5

    Ferguson, C.A. et al. Activin is an essential early mesenchymal signal in tooth development that is required for patterning of the murine dentition. Genes Dev. 12, 2636–2649 (1998).

  6. 6

    Vassalli, A., Matzuk, M.M., Gardner, H.A., Lee, K.F. & Jaenisch, R. Activin/inhibin bB subunit gene disruption leads to defects in eyelid development and female reproduction. Genes Dev. 8, 414–427 (1994).

  7. 7

    Schrewe, H., Gendron-Maguire, M., Harbison, M.L. & Gridley, T. Mice homozygous for a null mutation of activin bB are viable and fertile. Mech. Dev. 47, 43–51 (1994).

  8. 8

    Feng, Z.M., Madigan, M.B. & Chen, C.L. Expression of type II activin receptor genes in the male and female reproductive tissues of the rat. Endocrinology 132, 2593–2600 (1993).

  9. 9

    de Winter, J.P. et al. Activin receptor mRNA expression in rat testicular cell types. Mol. Cell. Endocrinol. 83, R1–8 (1992).

  10. 10

    Boitani, C., Stefanini, M., Fragale, A. & Morena, A.R. Activin stimulates Sertoli cell proliferation in a defined period of rat testis development. Endocrinology 136, 5438–5444 (1995).

  11. 11

    Kluin, P.M., Kramer, M.F. & de Rooij, D.G. Proliferation of spermatogonia and Sertoli cells in maturing mice. Anat. Embryol. 169, 73–78 (1984).

  12. 12

    Matzuk, M.M., Kumar, T.R. & Bradley, A. Different phenotypes for mice deficient in either activins or activin receptor type II. Nature 374, 356–360 (1995).

  13. 13

    Feijen, A., Goumans, M.J. & van den Eijnden-van Raaij, A.J. Expression of activin subunits, activin receptors and follistatin in postimplantation mouse embryos suggests specific developmental functions for different activins. Development 120, 3621–3637 (1994).

  14. 14

    Antonipillai, I., Wahe, M., Yamamoto, J. & Horton, R. Activin and inhibin have opposite effects on steroid 5 a-reductase activity in genital skin fibroblasts. Mol. Cell. Endocrinol. 107, 99–104 (1995).

  15. 15

    Mathews, L.S. & Vale, W.W. Expression cloning of an activin receptor, a predicted transmembrane serine kinase. Cell 65, 973–982 (1991).

  16. 16

    Dyson, S. & Gurdon, J.B. The interpretation of position in a morphogen gradient as revealed by occupancy of activin receptors. Cell 93, 557–568 (1998).

  17. 17

    Bunn, H.F. Induction of fetal hemoglobin in sickle cell disease. Blood 93, 1787–1789 (1999).

  18. 18

    Deconinck, N. et al. Expression of truncated utrophin leads to major functional improvements in dystrophin-deficient muscles of mice. Nature Med. 3, 1216–1221 (1997).

  19. 19

    Rafael, J.A., Tinsley, J.M., Potter, A.C., Deconinck, A.E. & Davies, K.E. Skeletal muscle-specific expression of a utrophin transgene rescues utrophin-dystrophin deficient mice. Nature Genet. 19, 79–82 (1998).

  20. 20

    Matzuk, M.M., Finegold, M.J., Su, J.-G.J., Hsueh, A.J.W. & Bradley, A. a-Inhibin is a tumour-suppressor gene with gonadal specificity in mice. Nature 360, 313–319 (1992).

  21. 21

    Ramirez-Solis, R. et al. Genomic DNA microextraction: a method to screen numerous samples. Anal. Biochem. 201, 331–335 (1992).

  22. 22

    Bradley, A. Teratocarcinomas and Embryonic Stem Cells: A Practical Approach 113–151 (IRL, Oxford, 1987).

  23. 23

    Elvin, J.A., Yan, C., Wang, P., Nishimori, K. & Matzuk, M.M. Molecular characterization of the follicle defects in the growth differentiation factor 9-deficient ovary. Mol. Endocrinol. 13, 1018–1034 (1999).

  24. 24

    Matzuk, M.M. et al. Transgenic models to study the roles of inhibins and activins in reproduction, oncogenesis, and development. Recent Prog. Horm. Res. 51, 123–154 (1996).

  25. 25

    Trudeau, V.L., Matzuk, M.M., Hache, R.J. & Renaud, L.P. Overexpression of activin-b A subunit mRNA is associated with decreased activin type II receptor mRNA levels in the testes of a-inhibin deficient mice. Biochem. Biophys. Res. Commun. 203, 105–112 (1994).

  26. 26

    Guo, Q. et al. Overexpression of mouse follistatin causes reproductive defects in transgenic mice. Mol. Endocrinol. 12, 96–106 (1998).

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Acknowledgements

We thank T.R. Kumar for measuring FSH levels; A. Lau and D. DeKretser for discussions; Q. Guo, P. Wang and Y. Wang for technical assistance; and W. Vale and J. Vaughan for providing antibodies to activin βA and βB proteins. This work was supported in part by NIH grants HD32067 (to M.M.M.), HD01156, HD27823 (to C.W.B.) and HD35708 (to T.K.W.), and the Robert Wood Johnson Foundation.

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Correspondence to Martin M. Matzuk.

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Brown, C., Houston-Hawkins, D., Woodruff, T. et al. Insertion of Inhbb into the Inhba locus rescues the Inhba-null phenotype and reveals new activin functions. Nat Genet 25, 453–457 (2000). https://doi.org/10.1038/78161

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