Fig. 1. Grh is expressed in the developing trachea and is required to prevent excessive tube extension. (A) Lateral view of the wild-type embryonic tracheal system at stage 16, visualised by antibodies against the 2A12 (lumenal) antigen. The major tracheal branches, dorsal trunk (DT), dorsal branch (DB), lateral trunk (LT), transverse connective (TC) and ganglionic branches (GB) are indicated. The rectangle outlines the part of the trachea that is shown in the subsequent panels, except for in F-I. (B-G) Tracheal lumens of wild-type (B,D,F) and grhs2140/Df(2R)Pcl7B mutant (C,E,G) embryos at stage 16. At the beginning of stage 16, grh mutants (C) have more elongated DTs than wild-type embryos (B). The DT growth continues as stage 16 proceeds, and at the end of this stage (D,E), the excessive tubular extension is evident in additional branches, including the TC (arrowheads in E), LT (not shown) and GB (F,G; ventral lateral view). (H-I) Dorsal view of wild-type (H) and grhs2140/Df(2R)Pcl7B mutant (I) embryos stained for 2A12 and DSRF, showing three pairs of unicellular terminal branches (arrowheads) emanating from the dorsal branches. Terminal branching and DSRF expression is normal in grh mutant embryos, but at the end of embryogenesis, the terminal branches become convoluted and often make loops in grh mutants (I, asterisk). (J,K) Wild type (J) and grh mutants (K) labelled with antisera against the nuclear protein Trh, which is expressed in all tracheal cells. The number of DT and DB cells is not greater in grh mutants than wild type. (L,M) Wild-type and (O,P) grhB37/grhB37 mutant embryos carrying the cytoplasmic trh-lacZ marker were labelled for Grh (red; L,O) and b-Gal (green; M,P). Grh is expressed in all tracheal cells in wild-type embryos (L; stage 14), and is absent in grh mutants (O; stage 14). (N,Q) Wild-type (N) and grhB37 embryos (Q) carrying the Grh activity reporter, GBE-lacZ, were double labelled with mAb2A12 (green) and anti-b-Gal (red). GBE-lacZ is expressed in the wild-type trachea (N; red nuclear staining), but is absent in grh mutants. Scale bar in A: 25 mm; B-K, 10 mm; L-Q, 20 mm. In this and all subsequent figures anterior is left and dorsal up.