Fig. 4. DAAM is required to organize apical actin into parallel running bundles in tracheal cells. In a wild-type (wt) tracheal tube, actin is organized into parallel running bundles that are perpendicular to the tube axis (A). The number and phasing of these actin bundles correspond to the taenidial fold pattern displayed on the tracheal cuticle (B). Actin bundles formed in the tracheal cells are located at the level of the adherens junctions (C,D). (C) A confocal projection of a wild-type tracheal tube where actin is visualized in red, while the adherens junctional marker, DE-cadherin is shown in green. (D) An optical xz section along the white line in C, apical is at the top. (E-G) The DAAM protein is largely colocalized with actin in the embryonic tracheal cells. Confocal sections have been collected from a one-segment wide region of the dorsal trunk of a stage 16 embryo. The cytoplasm of tracheal cells is labeled with GFP in green, actin is shown in red, DAAM is in blue. Arrow in F indicates the fusion cells located at the segmental boundary. DAAM is not expressed in these cells. (H-J) 3D projections of the same confocal sections shown in E-G. Sections were rotated within the XZ plane by 90°. There is strong colocalization of actin and DAAM at apical membranes of the tracheal tubes (J). In DAAM^Ex68 mutant tracheal tubes, not only is the cuticle pattern impaired (K), but actin organization is also severely altered in both the larval (L) and embryonic (M) tracheal cells. Actin cables formed are thinner and shorter than their wild-type counterparts (compare A with L, and E with M), and fail to organize into regularly aligned bundles. The formation of the apical actin bundles in the embryonic tracheal cells (marked by actin::GFP) is first detected at approximately 13 hours AEL (N). Scale bars: 20 µm in A for A,B; 50 µm in C for C,D; 50 µm in L for K,L; 10 µm in E-G for E-J; 10 µm in M for M.