Selective requirement of a deacetylase domain for Emp24independent luminal secretion in the Drosophila trachea
(English)Manuscript (preprint) (Other academic)
Vermiform and Serpentine are secreted putative chitin deacetylases (ChLDs). They are deposited into the tracheal lumen to terminate tube elongation during morphogenesis. Deletion analysis of a Serp-GFP reporter had revealed that the deacetylase domain is essential for its luminal localization. We transferred the deacetylase domain from Serp to Gasp, another tracheal luminal protein, which requires the Emp24 adaptor for ER exit. The GaspDeac-GFP chimera was normally secreted in emp24 mutants indicating that the deacetylase domain contains potential ER-exit signals. To explore this possibility we identified and characterized conserved sequence motifs in Serp deacetylase domain. We generated amino acid substitution mutants altering the three putative Nglycosylation sites, the predicted enzymatic activity cluster and three phylogenetically conserved motifs. We tested the cellular localization of the constructs in S2 cultured cells and the trachea of transgenic Drosophila embryos. Residue substitutions in the putative catalytic site neither interfered with Serp secretion nor with its ability to rescue the tracheal tube elongation defects of serp mutants. Mutations of the N-glycosylation sites gradually reduced the luminal deposition of Serp-GFP constructs suggesting that increased glycosylation enhances apical Serp secretion. By contrast, substitutions in each of the three uncharacterized amino acid stretches completely blocked the ER-exit of Serp-GFP constructs. The mutated proteins were N-glycosylated suggesting that the motifs may be involved in a subsequent protein-folding step or facilitate ER exit through interactions with unidentified specific adaptors.
Research subject Developmental Biology
IdentifiersURN: urn:nbn:se:su:diva-32570OAI: oai:DiVA.org:su-32570DiVA: diva2:280975