Global overexpression of Hoxa2 depletes rhombomere 1 of granule cell precursors (A) Flatmounts of brains up to embryonic day (E) 6 were prepared by opening the neural tube along the dorsal midline: cb – cerebellum, mb – midbrain, hb – hindbrain, rl – rhombic lip, ml – ventral midline. (B) The expression of Pax6 (blue) at E6 in one half of the anterior hindbrain viewed as a flatmount (rostral top, mediolateral axis runs left to right). Granule cell precursors born at the rhombic lip express Pax6 as they migrate over dorsal rhombomere 1 (r1) to condense as the external granule cell layer (EGL [egl]). Pax6 is also expressed in a ventral wedge of r1 precursors (arrow) abutting the anterior r2 boundary and ventral midline. The relationship of this Pax6 positive precursor pool (blue) to the anterior boundary of Hoxa2 (red) is shown inset. (C) Overexpression of Hoxa2 (red) leads to a loss of Pax6 in the EGL and prominent ventral wedge (arrow). This is replaced by a weaker, broadened, ventral Pax6 domain similar to that in the hindbrain. (D) At E9, in transverse section through a control embryo, the neuregulin receptor erbB4 labels the EGL. Red label dorsally reflects high levels of expression of the control virus, RCANBP(B)Hoxa2 (red), within the ventricular zone of the cerebellum (cb). (E) Injection of the active RCASBP(B)Hoxa2 virus leads to a reduction in erbB4. Patches of expression (arrows) correspond to residual EGL overlying a locally thickened cerebellum. (F) Cresyl violet stains in control and (G) RCASBP(B)Hoxa2-infected cerebella reveal an absence of cell bodies within a superficial, subpial layer (arrows), normally occupied by the EGL, following Hoxa2 overexpression. Scale bar (D,E) = 500 μm.