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The tail of the Xenopus tadpole will regenerate completely after transection. Much of the mass of the regenerate is composed of skeletal muscle, but there has been some uncertainty about the source of the new myofibres. Here, we show that the growing tail contains many muscle satellite cells. They are active in DNA replication, whereas the myonuclei are not. As in mammals, the satellite cells express pax7. We show that a domain-swapped construct, pax7EnR, can antagonize pax7 function. Transgenic tadpoles were prepared containing pax7EnR driven by a heat-inducible promoter. When induced, this reduces the proportion of satellite cells formed in the regenerate. A second amputation of the resulting tails yielded second regenerates containing notochord and spinal cord but little or no muscle. This shows that inhibition of pax7 action does not prevent differentiation of satellite cells to myofibres, but it does prevent their maintenance as a stem cell population.
Fig. 1. Expression pattern of pax7 in Xenopus early development. Whole-mount in situ hybridization was performed with the pax7 antisense RNA probe. (A-C) Dorsal view of stage 13 (A), stage 16 (B) and stage 19 (C) embryos, anterior towards left. The inset in B is an anterior transverse section of a stage 16 embryo, showing that pax7 is expressed in the sensorial layer of neural ectoderm and in the lateral plate mesoderm. The inset in the bottom left-hand corner in C shows the segmented pattern of pax7 expression in a stage 19 embryo. The inset in the top right-hand corner is a view of an anterior transverse section. (D-F) Lateral view of stage 25, 33 and 35 embryos, anterior towards left. The arrowheads in D,E indicate the expression domain of pax7 in the pronephros. Arrows in D,E show the chevron pattern of pax7 expression in somites. The inset in F is an anterior view of stage 35 embryo. The lines in F indicate the relative position of cross-section planes in H-J,L. (G) Sagittal section of stage 35 embryos. The inset indicates the transcripts of pax7 in spinal cord concentrated on the dorsal side. (H) Transverse section through the midbrain of a stage 35 embryo. (I) Pax7 expression in the pituitary anlage is marked by an asterisk. (J,L) Transverse section through the trunk of a stage 35 embryo. (K) Parasagittal section of stage 35 embryo showing pax7 transcripts in posterior pronephric anlage. (M) Horizontal section of stage 35 embryohead. (N) Parasagittal section of stage 35 embryo. The black arrows in H,M,N indicate the mesenchyme cells with pax7 expression anterior to the eye. The white arrow in N indicates that pax7 transcripts locate in the edges of myotomes. (O) Parasagittal view of the tail of stage 35 embryo. Abbreviations: e, eye; mb, midbrain; n, notochord; pa, pituitary anlage; pr, pronephros; sc, spinal cord; so, somite.
Fig. 2. Pax7 antibody detection in stage 46 tadpole. Immunostaining with anti-Pax7 monoclonal antibody was carried out on transverse sections of stage 46 tadpoles. (A-C) Detection of Pax7 with DAB staining (brown). The tissues were counterstained with 0.5% Methyl Green solution. (A) Midbrain; (B) hindbrain; (C) spinal cord. (D) Immunostaining of Pax7 (red) and DAPI (blue) on cross-section of tadpolehead. The arrow in D indicates expression of Pax7 in eyemuscle. (E) Co-immunostaining of Pax7 (red) and laminin (green) on cross-section of tadpoletailmuscle. Abbreviations: hb, hindbrain; mb, midbrain; pg, pituitary gland; sc, spinal cord. Scale bars: 20 μm.
