Trynity controls epidermal barrier function and respiratory tube maturation in Drosophila by modulating apical extracellular matrix nano-patterning

Abstract

The outer surface of insects is covered by the cuticle, which is derived from the apical extracellular matrix (aECM). The aECM is secreted by epidermal cells during embryogenesis. The aECM exhibits large variations in structure, function, and constituent molecules, reflecting the enormous diversity in insect appearances. To investigate the molecular principles of aECM organization and function, here we studied the role of a conserved aECM protein, the ZP domain protein Trynity, in Drosophila melanogaster. We first identified trynity as an essential gene for epidermal barrier function. trynity mutation caused disintegration of the outermost envelope layer of the cuticle, resulting in small-molecule leakage and in growth and molting defects. In addition, the tracheal tubules of trynity mutants showed defects in pore-like structures of the cuticle, and the mutant tracheal cells failed to absorb luminal proteins and liquid. Our findings indicated that trynity plays essential roles in organizing nano-level structures in the envelope layer of the cuticle that both restrict molecular trafficking through the epidermis and promote the massive absorption pulse in the trachea.

Nanopore formation in the cuticle of an insect olfactory sensillum

“…How the Gore-tex protein, which is localized to endosomal compartments, affects the extracellular events of cuticle envelope assembly is still unclear. One possibility is that Goretex regulates the trafficking of specific effector molecules for envelope assembly such as Trynity [31], as previously suggested for the function of Osi21/die4 in regulating endocytosed 20…”

“…Embryo cuticle preparation and live imaging of hatching behavior For cuticle preparations, larvae were placed on a glass slide with Hoyer's medium and lactic acid 5 (Wako) 1:1 and cover glass, and incubated at 60-65 °C overnight before observation with differential interference microscopy. For live imaging of hatching behavior, dechorionated stage-17 embryos with visible Malpighian tubules and slightly pigmented mouth cuticle placed on a plate with heptane glue (heptane with sticky tape), covered with water and imaged with a Digital Microscope VHX-6000 (Keyence) as described [31]. 10 LFP recording from the maxillary palp As the method of odor response assay LFP recording, not single sensilla recordings was chosen to avoid breaking cuticle.…”

Fluorescence-based 3D targeting of FIB-SEM acquisition of small volumes in large samples

“…An ultrastructural characterization that is necessary to understand the physiology and development of tracheal terminal cells can only be obtained using EM. Previous studies were limited to 2D TEM at embryonic development, or the earliest larval stages (Itakura et al, 2018; Öztürk-Çolak et al, 2016b; Jones et al, 2014; Nikolova and Metzstein, 2015) and the ultrastructure of the tracheal ECM has thus never been observed at the latest larval stage, even though this is when the terminal cells undertake most of their growth (JayaNandanan et al, 2014). Due to the size of the animal and the small number of tracheal terminal cells, a 3D EM analysis of these cells at later stages requires a precise targeting strategy (Fig.…”

Glycosylhydrolase genes control respiratory tubes sizes and airway stability

“…At the end of embryogenesis, tracheal cells need to internalize contents of the chitin-cable, in a process called airway clearance, which subsequently includes initial gas-filling of the branch system 26 , 27 . In parallel, tubes establish a chitin-cuticle at their apical surface, which forms a waterproof barrier and thus enables gas-transport 28 – 30 . This late embryonic tracheal cuticle is of crucial importance since any inhibition of tracheal oxygenation in first-instar larvae prevents further body growth and molting into the next larval stages 31 , 32 .…”

“…branch system 26,27 . In parallel, tubes establish a chitin-cuticle at their apical surface, which forms a waterproof barrier and thus enables gas-transport [28][29][30] . This late embryonic tracheal cuticle is of crucial importance since any inhibition of tracheal oxygenation in first-instar larvae prevents further body growth and molting into the next larval stages 31,32 .…”