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.

Anionic and zwitterionic moieties as widespread glycan modifications in non-vertebrates

“…The single Drosophila sialyltransferase has been functionally and biochemically characterised [50,51]. In other insect species, sialyltransferase genes have been identified and parts of the CMP-sialic acid biosynthesis pathway revealed [52,53]; however, only upon glycoengineering can it be safely concluded that Lepidoptera will express sialic acid on recombinant glycoproteins [54].…”

Diversity of sialic acids and sialoglycoproteins in gametes and at fertilization

“…Frontiers in Cell and Developmental Biology frontiersin.org deuterostomes, and Arthropods (Di et al, 2017;Petit et al, 2018). Interestingly, in the zebrafish Danio rerio, there are two CSS with different substrate specificity and spatial expression one localizing in the nucleus and showing higher specificity towards Neu5Ac and the other localizing to the cytosol and showing higher specificity towards Kdn (Schaper et al, 2012).…”

“…Subsequent use of Sias depends on their activation to the sugar-nucleotide cytidine monophosphate sialic acid (CMP-sialic acid or CMP-Sia) which is catalyzed by the cytidine monophosphate N -acetylneuraminic acid synthetase or CMP-sialic acid synthetase (CMAS or CSS) in the nucleus of most vertebrate cells ( Figure 2A ). In Metazoa, orthologous sequences of human CMAS gene were retrieved in the Cnidaria, deuterostomes, and Arthropods ( Di et al, 2017 ; Petit et al, 2018 ). Interestingly, in the zebrafish Danio rerio , there are two CSS with different substrate specificity and spatial expression one localizing in the nucleus and showing higher specificity towards Neu5Ac and the other localizing to the cytosol and showing higher specificity towards Kdn ( Schaper et al, 2012 ).…”

Exploration of the Sialic Acid World

“…Interestingly, also insects were shown to contain sialic acids and to express corresponding enzymes. [8][9][10][11] Sialic acids were found in some viruses, various bacteria, protozoa, and pathogenic fungi. They are absent in plants and in most animals in the evolutionary tree lower than the echinoderms.…”