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First published online 26 March 2008
doi: 10.1242/dev.019331

Development 135, 1597-1604 (2008)
Published by The Company of Biologists 2008
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Different autonomous myogenic cell populations revealed by ablation of Myf5-expressing cells during mouse embryogenesis

Nicole Gensch1,*, Thilo Borchardt1,*, Andre Schneider1, Dieter Riethmacher2 and Thomas Braun1,{dagger}

1 Max-Planck-Institute for Heart and Lung Research, Parkstr. 1, 61231 Bad Nauheim, Germany.
2 Human Genetics Division, University of Southampton, Southampton SO16 6YD, UK.


Figure 1
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  		Fig. 1. Contribution of Myf5-derived cells to different cell types of the paraxial mesoderm. (A-K) Expression of lacZ in E9.5 (A,B,E), E10.5 (C,D,F) and E15.5 (G-K) Rosa26lacZ embryos crossed either with myogenin-Cre (A,B,E) or with Myf5-Cre mice (C,D,F-K). lacZ staining (blue) reveals Myf5-derived cells in the sclerotome at E10.5 (F), in the rib cartilage (G,I), vertebrae (H) and skeletal muscles of the body wall (G) and of a limb (J). Arrows in G indicate the limited presence of Myf5-derived cells in the diaphragm of this embryo. Very few Myf5-derived cells are present in the neural tube and in the dorsal root ganglia (arrows in K). Arrowheads in K indicate lacZ-positive chondrocytes in a vertebra. Scale bars: 100 µm.

 

Figure 2
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  		Fig. 2. Efficient DTA-mediated ablation of Myf5-derived cells at E15.5. Myf5-Cre mice were crossed either to Z/AP reporter mice (A,C,E) or to Z/AP//DTA mice (B,D,F). AP staining (red) reveals Myf5-derived cells in different muscles of Myf5-Cre//Z/AP mice at E15.5 (A,C,E). Simultaneous activation of DTA by Myf5-Cre leads to a complete ablation of Myf5-derived cells in Myf5-Cre//DTA//Z/AP mice (B,D,F). (A,B) tongue muscles. (C-F) Limb and body wall muscles. Scale bars: 200 µm.

 

Figure 3
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  		Fig. 3. Loss of MyHC-expressing skeletal muscle cells in myogenin-Cre//DTA mice at E18.5. (A-L) Immunofluorescence staining with a MyHC antibody (MF20) of sections from Myf5-Cre//DTA (A,D,G,J), myogenin-Cre//DTA (B,E,H,K) and wild-type mice (C,F,I,L) at E14.5 (A-C) and E18.5 (D-L). Note the absence of MyHC-positive muscle cells in myogenin-Cre//DTA but not in Myf5-Cre//DTA mice, which show a virtually normal formation of skeletal muscles at E18.5. Sections from wild-type mice are shown for comparison. Scale bars: 200 µm.

 

Figure 4
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  		Fig. 4. Reduced expression of MRFs in Myf5-Cre//DTA mice at E10.5. Semi-quantitative RT-PCR analysis of Myf5, Myod1, myogenin, Myf6, Pax3 and Pax7 mRNAs from E10.5 and E14.5 wild-type and Myf5-Cre/DTA mice. Acidic ribosomal protein (ARS) served as a loading control.

 

Figure 5
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  		Fig. 5. Loss of Myf5-expressing cells in somites of Myf5-Cre//DTA mice at E10.5. Immunofluorescence staining of somites of Myf5-Cre//DTA (A,D,G), myogenin-Cre//DTA (B,E,H) and wild-type embryos (C,F,I) at E10.5 using a combination of Myf5 and Myod1, Myf5/Pax7 and Myf5/myogenin antibodies. Somites from the cranial part of embryos are shown. Myf5-expressing cells are lacking in Myf5-Cre//DTA but not in myogenin-Cre//DTA and wild-type embryos, and there is a massive reduction of Myod1- and myogenin-expressing cells in Myf5-Cre//DTA embryos. Broken lines indicate the outline of the embryos. Scale bars: 100 µm.

 

Figure 6
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  		Fig. 6. Compensatory increase of Myod1 expression in Myf5-Cre//DTA embryos. (A-F) In situ hybridization of Myf5-Cre//DTA (A-C) and wild-type (D-F) embryos at E10.5 (A,D), E11.5 (B,E) and E12.5 (C,F) with a probes against Myod1. No expression of Myod1 was observed in Myf5-Cre//DTA embryos at E10.5. At E11.5 an abnormal expression of Myod1 was present in the hypaxial part of the dermomyotome of Myf5-Cre//DTA embryos compared with the more widespread expression in wild-type embryos. At E12.5, no major differences in the expression of Myod1 were found between wild-type and mutant embryos. Scale bars: 150 µm.

 

Figure 7
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  		Fig. 7. Defects of the axial skeleton in Myf5-Cre//DTA mice at E18.5. (A-F) Macroscopic views of newborn myogenin-Cre//DTA (A,D), Myf5-Cre//DTA (B,E) and WT (C,F) before (A-C) and after Alzian Blue/Alizarin Red staining (D-F). Enlarged views of the deformed skeleton of Myf5-Cre//DTA mice (G-I). Note the abnormal posture of myogenin-Cre//DTA mice, owing to the absence of skeletal muscles. No primary malformations of the skeleton of myogenin-Cre//DTA mice are apparent.

 

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