Figure 2.

BmprIa^flox/- hindlimbs are bidorsal at and around the time of the motor axon dorsoventral projection choice. The dorsoventral character of limb mesenchyme was defined using molecular markers. Brn4-cre^Tg/-, BmprIa^flox/- (BmprIa^flox/-) mutant hindlimbs are completely dorsalized. (a-d) Upper panels: control hindlimbs. At E11.0 axons have reached the base of the hindlimb (a,b). Plzf is a general limb marker (b), and Lmx1b is a dorsal limb marker (a,b). EphA4 marks proximal dorsal mesenchyme (c) and Σephrin-A marks ventral mesenchyme (d) at E11.5. Note coincidence of proximal expression borders. Lower panels: BmprIa^flox/- mutant hindlimbs. Marker expression demonstrates they are completely dorsalized (Lmx1b+ Σephrin-A-) (a,b,d) and that they are proximally dorsal (EphA4+) (c). (e) Summary of molecular dorsoventral code marker expression in control (upper) or bidorsal BmprIa^flox/- mutant (lower) hindlimb. RNA in situ hybridization (Plzf), immunostaining (Lmx1b, EphA4) and AP fusion protein staining (Σephrin-A) were performed on adjacent transverse cryosections and images were digitally superimposed. n = 12 (a,b), 8 (c), 5 (d) embryos. Top, dorsal; left, medial. Dotted lines: limb outline. NF, neurofilament. Asterisks indicate the position of the plexus.