Figure 1. Dynamics of taenidial fold and actin ring formation.

(A–F) Dorsal Trunk detail of wild-type embryos stained with fluostain to label the chitin structures. Maximum projections of confocal Z sections showing the dynamics of intraluminal chitin filament and taenidial folds during late stages of embryonic development. Chitin structures are schematically represented under each image. Chitin filament: at late stage 16, intraluminal chitin filament is thick and dense (A); as the embryo develops, it becomes less and less dense (B, C) until it turns into a thin chitin fibre that runs in zigzags along the tube diameter (D); in the last steps of embryogenesis, the intraluminal chitin filament is completely cleared from the lumen (E, F). Taenidial folds: at late stage 16, taenidial folds are newly formed and thin (A); as the taenidial folds become thicker, it is apparent that the taenidial folds at fusion points are not formed yet (B); later, the taenidial folds at fusion points are also formed which generates a continuous taenidial structure along the tube (C); in the final steps, as the intraluminal chitin filament is cleared from the lumen, the taenidial folds reach their the most mature form (D–F). Scale bars = 10 μm. (G–J) wild-type embryonic (G–H) and 3^rd instar larval (I–J) trachea stained with fluostain (red) and phalloidin (green) showing taenidial folds and F-actin bundles together (G, H, I, J) or separately (G’, G’’, H’, H’’ and I’, I’’). F-actin organisation in structures perpendicular to the main tube axis occurs at stage 16 prior to taenidial fold appearance (G). When taenidia become apparent (H), they are positioned over the actin bundles and this co-localisation continues throughout larval stages (I, J). Scale bars = 10 μm. (K) TEM detail of wild-type DT taenidia, the open triangle points to actin filament cross-sections (diameter around 7 nm). Scale bar 250 nm. (L) Detail of DT showing apical cell borders (labelled by anti-DE-Cad in green) and how taenidia (labelled by fluostain in red) span continuously beyond cell-cell borders from one cell to the other.

DOI: http://dx.doi.org/10.7554/eLife.09373.003

Figure 1—figure supplement 1. Time course of actin ring and taenidial fold formation.

(A–F) Projections of confocal sections of wild-type DTs from early stage 16 to stage 17. F-actin is detected by phalloidin (red) and Chitin by fluostain (blue). The F-actin (red) and chitin (blue) structures are schematically represented under each image. First F-actin bundles are formed at the fusion points during the fusion process of the DT (A’, arrows). These bundles are formed as a result of highly orchestrated cell shape changes during fusion events. At mid-stage 16, F-actin rings become visible along the DT while the taenidial folds are not fully formed yet and hence not labelled with fluostain (B). Then, very thin taenidial folds become visible (C). At first, they are not formed at fusion points (D). Later, taenidial folds at fusion points are also formed, thus generating a continuous taenidial structure along the tube (E). Finally, the taenidial folds and F-actin bundles reach their most mature form as the trachea start to fill with air (F). Both stainings are shown in the merge images (A-F). The fluostain (A’-F’) and phalloidin (A’’-F’’) stainings are shown separately.

DOI: http://dx.doi.org/10.7554/eLife.09373.004