Myf-5 Revisited: Loss of Early Myotome Formation Does Not Lead to a Rib Phenotype in Homozygous Myf-5 Mutant Mice

Abstract

The Myf-5/MRF4 locus has been targeted several times. Most alleles published so far generated significant, though different, rib pattern formation deficits. The explanation for the rib phenotype concentrated on defects in myotome formation caused by a lack or a massive reduction of Myf-5 expression. The results of three different mutations introduced into the MRF4 gene, which is located approximately 8 kb upstream of Myf-5 on mouse chromosome 10 seemed to support this view (Olson et al. 1996xOlson, E.N., Arnold, H.H., Rigby, P.W., and Wold, B.J. Cell. 1996; 85: 1–4Abstract | Full Text | Full Text PDF | PubMed | Scopus (364)See all ReferencesOlson et al. 1996).The generation of the two new Myf-5 alleles reported here, which do not show any malformations of the ribs or other skeletal elements despite the absence of Myf-5 mRNA in somites and the complete loss of early myotome formation, exclude this explanation. We can rule out a direct involvement of the Myf-5 and/or MRF4 proteins in the generation of the short rib phenotype. Neither the Myf-5ΔloxP nor the “weakest” MRF4 mutations, which are true null-alleles of the Myf-5 and the MRF4 gene, respectively, nor Myf-5ΔloxP/Myf-6m1 mice and Myf-5ΔloxP/Myf-5m1 compound heterozygous mice show any rib truncations. These findings now strongly suggest that an alteration of the activity of a third or more unknown gene(s) is the real cause for the skeletal malformations seen in Myf-5m1, Myf-5lacZ, and the various MRF4 knockout mouse strains. This does not necessarily mean that no cross-talk between different somitic layers is necessary for rib formation. A number of separate experiments indicate that the dermomyotome but not necessarily the myotomal cell layer is involved in the morphogenesis of the distal parts of the ribs (6xHenderson, D.J., Conway, S.J., and Copp, A.J. Dev. Biol. 1999; 209: 143–158Crossref | PubMed | Scopus (56)See all References, 7xHuang, R., Zhi, Q., Schmidt, C., Wilting, J., Brand-Saberi, B., and Christ, B. Development. 2000; 127: 527–532PubMedSee all References). It is still feasible that inductive influences from the dermomyotome are necessary for early specification of lateral sclerotome. Alteration of a gene that is normally active in the dermomyotome by the Myf-5 mutation may therefore be causative for sclerotomal malformations.How does the phenotype of the knockout of the Myogenin gene that resides on a differerent chromosome but also results in a rib phenotype (Hasty et al. 1993xHasty, P., Bradley, A., Morris, J.H., Edmundson, D.G., Venuti, J., Olson, E.N., and Klein, W.H. Nature. 1993; 364: 501–506Crossref | PubMed | Scopus (588)See all ReferencesHasty et al. 1993) fit into this picture? (1) The myogenin phenotype is rather different from the Myf-5 phenotype. (2) It was never very likely to depend on the same mechanism, since the lack of Myogenin did not result in a lack of myotome formation and the development of primary muscle cells in Myogenin mutant embryos was not severely affected (Venuti et al. 1995xVenuti, J.M., Morris, J.H., Vivian, J.L., Olson, E.N., and Klein, W.H. J. Cell. Biol. 1995; 128: 563–576Crossref | PubMed | Scopus (162)See all ReferencesVenuti et al. 1995). It appears most likely that either the increase of the relative numbers of myoblasts during primary or secondary myogenesis affects the delicate balance between dermomyotome and sclerotome or at a later stage the interaction of the rib blastema with surrounding tissues. This may lead to a phenocopy of some aspects of the Myf-5 phenotype without affection of the gene that causes the phenotype in Myf-5 mutants.Sequence analysis of the greater Myf-5 locus is in progress. Expression analysis of potential open reading frames from this region during embryonic development in wild-type, Myf-5ΔloxP, and Myf-5m1 mice will allow us to determine whether these genes might be involved in formation and growth of the sclerotome, rib primordia, or chondrocytes. Final proof of the role of these genes will await genetic manipulation of their expression in transgenic mice.The Myf-5 example emphasizes the fact that our current ability to build valid models explaining the phenotype of targeted mutations is strongly biased by our knowledge of neighboring genes. Although a number of cases have been described where genes of the same multigene group in a cluster were affected by a targeted mutation, no examples have been described where an unrelated gene was influenced in cis by gene targeting experiments.Additional data related to this article can be found online (http://www.cell.com/cgi/content/full/102/1/17/DC1).Acknowledgments: work in the authors' laboratory was supported by the DFG, FCI, and the Volkswagen Foundation.*To whom correspondence should be addressed (e-mail: thomas.braun@medizin.uni-halle.de).†Both authors contributed equally to the work.‡Present address: Mice&More, Martinistr. 52, 20251 Hamburg, Germany.