Fig. 5. Six1 is not required for myotomal differentiation or myogenic precursor cell migration. Immunochemistry experiments performed on transverse sections of E9.5 (A-C) and E11 (D-M) embryos did not show any differences between Six1^–/–, Six1^+/– and wild-type littermates. Presented here are only the results obtained with Six1^–/– embryos (A-K), except for L and M which show the expression of Six4 and myogenin in Six1^+/– embryos. D, dorsal; V, ventral. (A-C) Absence of Six1 does not impair early somitic differentiation: at E9.5 immunostaining with Myf5 (A), myogenin (B) and Pax3 (C) antibodies shows that Myf5 and myogenin accumulate correctly in the myotome (double arrowheads in A and B) and that Pax3 is normal in the dermomyotome (arrowhead in C). (D-F) Six1 does not control the expression of Pax3 in the dermomyotome and does not impair Pax3-dependent migration of hypaxial progenitor. (D) At E11, Pax3 expression is detected in the dermomyotome (arrowhead) and in migrating myogenic cells delaminating from the lateral edge of the dermomyotome at the forelimb level. (F) ß-galactosidase immunostaining revealed Six1-expressing cells in the myotome, in the lateral part of the dermomyotome and in migrating cells. (E) Double staining for Pax3 and ß-gal demonstrate that most of the Pax3-expressing cells in the lateral part of the dermomyotome, as well as most of the migrating precursors, also co-express Six1 (arrowhead), whereas differentiated myotomal cells express only Six1 (double arrowhead) and median dermomyotomal cells express only Pax3 (arrow). (G,H) Absence of Six1 does not impair Pax3-dependent migration into the limbs. Immunostaining with Pax3 antibodies shows that Pax3 accumulates correctly in migrating myogenic cells of both ventral and dorsal regions of the forelimb bud (G) at E11. (H) Six1 does not control the expression of Myf5 in limb buds: immunostaining with Myf5 antibodies shows that Myf5 can also be detected in dorsal and ventral myogenic regions of the forelimb bud at E11. (I-M) Six1 is not required for the activation of Myf5, Six4 and myogenin expression in the myotome. (I) Myf5 expression (double arrowheads) is not altered in Six1^–/– myotome. (J,L) Six4 antibodies allow detection of Six4 accumulation in the dermomyotome (single arrowhead) and myotome (double arrowheads) in Six1^–/– (J) and Six1^+/– (L) embryos. Expression of Six4 in the myotome could compensate the absence of Six1 (K,M). Myogenin expression is detected with a specific antibody in both Six1^–/– (K) and Six1^+/– (M) embryos in the myotome (double arrowheads).