title: Diverse subcellular localizations of the insect CMP-sialic acid synthetases.
abstract: The occurrence and biological importance of sialic acid (Sia) and its metabolic enzymes in insects have been studied using Drosophila melanogaster. The most prominent feature of D. melanogaster CMP-Sia synthetase (DmCSS) is its Golgi-localization, contrasted with nuclear localization of vertebrate CSSs. However, it remains unclear if the Golgi-localization is common to other insect CSSs and why it happens. To answer these questions, Aedes aegypti (mosquito) CSS (AaCSS) and Tribolium castaneum (beetle) CSS (TcCSS) were cloned and characterized for their activity and subcellular localization. Our new findings show: (1) AaCSS and TcCSS share a common overall structure with DmCSS in terms of evolutionarily conserved motifs and the absence of the C-terminal domain typical to vertebrate CSSs; (2) when expressed in mammalian and insect cells, AaCSS and TcCSS showed in vivo and in vitro CSS activities, similar to DmCSS. In contrast, when expressed in bacteria, they lacked CSS activity because the N-terminal hydrophobic region appeared to induce protein aggregation; (3) when expressed in Drosophila S2 cells, AaCSS and TcCSS were predominantly localized in the ER, but not in the Golgi. Surprisingly, DmCSS was mainly secreted into the culture medium, although partially detected in Golgi. Consistent with these results, the N-terminal hydrophobic regions of AaCSS and TcCSS functioned as a signal peptide to render them soluble in the ER, while the N-terminus of DmCSS functioned as a membrane-spanning region of type II transmembrane proteins whose cytosolic KLK sequence functioned as an ER export signal. Accordingly, the differential subcellular localization of insect CSSs are distinctively more diverse than previously recognized.
authors.affiliation: Bioscience and Biotechnology Center, and Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya, Japan.|Program for Leading Graduate Schools, Integrative Graduate Education and Research Program in Green Natural Sciences, Nagoya University, Nagoya, Japan.
authors.forename: Wu
authors.initials: W
authors.lastname: Di
authors.affiliation: Bioscience and Biotechnology Center, and Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya, Japan.
authors.forename: Akiko
authors.initials: A
authors.lastname: Fujita
authors.affiliation: Bioscience and Biotechnology Center, and Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya, Japan.
authors.forename: Kayo
authors.initials: K
authors.lastname: Hamaguchi
authors.affiliation: University of Lille, CNRS, UMR 8576-UGS-Unité de Glycobiologie Structurale et Fonctionnelle, Lille, France.
authors.forename: Philippe
authors.initials: P
authors.lastname: Delannoy
authors.affiliation: Bioscience and Biotechnology Center, and Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya, Japan.|Program for Leading Graduate Schools, Integrative Graduate Education and Research Program in Green Natural Sciences, Nagoya University, Nagoya, Japan.
authors.forename: Chihiro
authors.initials: C
authors.lastname: Sato
authors.affiliation: Bioscience and Biotechnology Center, and Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya, Japan.|Program for Leading Graduate Schools, Integrative Graduate Education and Research Program in Green Natural Sciences, Nagoya University, Nagoya, Japan.
authors.forename: Ken
authors.initials: K
authors.lastname: Kitajima
mesh_terms: D000330:Aedes / Q000201:enzymology; D020816:Amino Acid Motifs / Q000235:genetics; D000818:Animals; D004331:Drosophila melanogaster / Q000201:enzymology / Q000235:genetics; D006056:Golgi Apparatus / Q000201:enzymology / Q000235:genetics; D009154:Mutation; D019158:N-Acetylneuraminic Acid / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D000216:N-Acylneuraminate Cytidylyltransferase / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D011487:Protein Conformation; D014231:Tribolium / Q000201:enzymology
chemical_list: D000216:N-Acylneuraminate Cytidylyltransferase; D019158:N-Acetylneuraminic Acid
keywords: beetle; CMP-sialic acid synthetase; Drosophila; ER export signal; mosquito