AS3_9 locus and its enhancer activity in the frontonasal region.

<p>(A) Location of human AS3_9 and surrounding genes. (B) The phastCons conservation track of AS3_9 in mouse (UCSC Genome Browser; mm10); the blue, yellow, and green bars denote the AmnSINE1, X6b_DNA, and MER117 regions, respectively. The construct used to produce AS3_9-<i>lacZ</i> mice is illustrated below. (C) Whole-mount (left), frontal view of the head (top right), and ventral view of the upper jaw (bottom right) showing <i>lacZ</i> expression in AS3_9-<i>lacZ</i> E13.5 embryos. (D) Coronal section of an AS3_9-<i>lacZ</i> E14.5 embryo. (E) ISH for <i>wnt5a</i> in E14.5 wild type. Arrowheads indicate the palatal shelves (C–E). MN, medial nasal process; LN, lateral nasal process; Max, maxillary process; Man, mandibular process. Scale bar (C–E): 0.5 mm.</p

<i>Wnt5a</i> expression in E14.5 coronal sections from the anterior and intermediate regions of the oral cavity in one wild-type and three AS3_9-ko embryos (#1–3).

<p>(A) Anterior region of a wild-type embryo. (B-D) Anterior region of three AS3_9-ko embryos. (E) Intermediate region of a wild-type embryo. (F-H) Intermediate region of three AS3_9-ko embryos. Arrowheads denote palatal shelves. Scale bar: 0.5 mm.</p

Functional dissection of AS3_9 enhancer.

<p>Top left: Conservation track from the UCSC Genome Browser (mm10) and TE regions in mouse. Shown are nine constructs (a–j) and their enhancer activity in the maxillary and mandibular processes, nasal processes, and palatal shelves (right). Lateral view (a'–j') and ventral view (a''–j'') of the upper jaw of E13.5 transgenic embryos harboring each deletion construct (a–i) and the construct with a mutated Msx1-binding site (j). <i>LacZ</i> expression was clearly observed in the maxillary and mandibular processes (yellow arrowheads), nasal processes (green arrowheads), and palatal shelves (red arrowheads). Scale bar: 1 mm.</p

<i>Wnt5a</i> expression in E14.5 coronal sections from the anterior and intermediate regions of the oral cavity in one wild-type and three AS3_9-ko embryos (#1–3).

<p>(A) Anterior region of a wild-type embryo. (B-D) Anterior region of three AS3_9-ko embryos. (E) Intermediate region of a wild-type embryo. (F-H) Intermediate region of three AS3_9-ko embryos. Arrowheads denote palatal shelves. Scale bar: 0.5 mm.</p

Functional dissection of AS3_9 enhancer.

<p>Top left: Conservation track from the UCSC Genome Browser (mm10) and TE regions in mouse. Shown are nine constructs (a–j) and their enhancer activity in the maxillary and mandibular processes, nasal processes, and palatal shelves (right). Lateral view (a'–j') and ventral view (a''–j'') of the upper jaw of E13.5 transgenic embryos harboring each deletion construct (a–i) and the construct with a mutated Msx1-binding site (j). <i>LacZ</i> expression was clearly observed in the maxillary and mandibular processes (yellow arrowheads), nasal processes (green arrowheads), and palatal shelves (red arrowheads). Scale bar: 1 mm.</p