Forebrain patterning defects in bel mutants. (A,B) In bel mutants (B), dlx2 expression is extremely reduced in anterior/dorsal diencephalon (arrows) and is slightly reduced in the telencephalon (brackets), compared with in wild type (A). Inset in A shows a similar loss of dlx2 expression in the anterior diencephalon after injection of lhx2 antisense MO into a wild-type embryo. (C,D) In bel mutants (D), nk2.1b expression is regionally disrupted in the diencephalon (arrows) and expanded at a small region at the optic recess (arrowhead), compared with in wild type (C). E, F) fgf8 expression is extremely reduced in the optic stalk region (arrowheads) in bel mutants (F), compared with wild type (E). Insets show anterior views of the head, with fgf8 expression absent at the midline (arrowheads) in bel mutants. (G, H) In bel mutants (H), pax2.1 expression is absent in the medial optic stalk (arrowheads) but remains laterally. (I,J) zic2.1 expression is similarly absent across the midline (arrowheads) in bel mutants (J). (K, L) The number of phospho-histone-labeled mitotic cells is reduced in the dorsal/anterior diencephalon of bel mutants (brackets). (M) Graph showing a significant reduction in the number of proliferating cells in the mutant diencephalon (asterisk, P<0.01). (N, O) Anterior views showing the loss of dlx2 expression in the anterior diencephalon of bel mutants (arrows, compare with lateral views in A,B). (P-U) Fluorescein dextran-labeled cells (red/brown) were transplanted between wild-type and bel mutant embryos. (P,R,T) Transplanted wild-type cells express dlx2 (blue) in a wild-type (P) and bel mutant (R,T) background (seven embryos). (Q) Transplanted bel mutant cells do not express dlx2 in the ventral midline in bel mutants. (S,U) In a wild-type background (19 embryos), bel mutant cell clones do not express dlx2, but populate the midline and are intermingled with wild-type cell clones. (T,U) Higher magnification view of transplanted cells in the diencephalon. (T) dlx2 transcripts (blue) and lineage tracer (red) are present in wild-type cells (purple arrowheads) in a bel mutant forebrain. (U) By contrast, alternating blue (dlx2-positive wild type) and red (lineage tracer containing dlx2-negative bel mutant) cells occupy the ventral midline in a wild-type background (red and blue arrowheads). (A-F,K,L) Lateral views of the forebrain, anterior left, eyes removed; (G-J) ventral views of the forebrain, anterior up; (N-U) anterior views of the forebrain, dorsal up; (T,U) Black dots mark the position of the optic recess. Scale bars: 15 μm.
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