Involvement of mind the gap in the organization of the tracheal apical extracellular matrix in Drosophila and Nilaparvata lugens

Abstract

The tracheal apical extracellular matrix (aECM) is vital for expansion of the tracheal lumen and supports the normal structure of the lumen to guarantee air entry and circulation in insects. Although it has been found that some cuticular proteins are involved in the organization of the aECM, unidentified factors still exist. Here, we found that mind the gap (Mtg), a predicted chitin-binding protein, is required for the normal formation of the apical chitin matrix of airway tubes in the model holometabolous insect Drosophila melanogaster. Similar to chitin, the Mtg protein was linearly arranged in the tracheal dorsal trunk of the tracheae in Drosophila. Decreased mtg expression in the tracheae seriously affected the viability of larvae and caused tracheal chitin spiral defects in some larvae. Analysis of mtg mutant showed that mtg was required for normal development of tracheae in embryos. Irregular taenidial folds of some mtg mutant embryos were found on either lateral view of tracheal dorsal trunk or internal view of transmission electron microscopy analysis. These abnormal tracheae were not fully filled with gas and accompanied by a reduction in tracheal width, which are characteristic phenotypes of tracheal aECM defects. Furthermore, in the hemimetabolous brown planthopper (BPH) Nilaparvata lugens, downregulation of NlCPAP1-N (a homolog of mtg) also led to the formation of abnormal tracheal chitin spirals and death. These results suggest that mtg and its homolog are involved in the proper organization of the tracheal aECMs in flies and BPH, and that this function may be conserved in insects.

Genome‐wide in vitro and in vivo RNAi screens reveal Fer3 to be an important regulator of kkv transcription in Drosophila

“…Mutant clones were generated using the CRISPR/Cas9 system and Vasa‐Cas9 alleles. Fer3 mutant generation was performed in accordance with a previously described method (Yue et al ., 2019). The genomic locations of the sites targeted by the guide RNAs are shown in Figure S1, and the primers used in this procedure are listed in the Supplementary Materials and Methods.…”

“…The isolation of total RNA from each sample and quantitative reverse transcription (qRT)‐PCR procedures were performed as previously described (Yue et al ., 2019). Fly Act42A (FlyBase ID: FBgn0000043) was used as an internal control (Ponton et al ., 2011).…”