Recent evidence for a similar lysosomal protein-sorting machinery in Drosophila Schneider S2 cells has been found by identifying a homolog of the mammalian mannose 6-phosphate receptor. Our findings support this hypothesis by demonstrating that many enzymes with hydrolytic activities which are known to concentrate in lysosomes contain oligo-mannosidic N-glycans. Another interesting observation was the occurrence of at least 10–25% of proteins without a protein signature for the attachment of an N-glycan structure. These observations suggest that mannose-containing O-glycosylation may be abundantly present in insect species. To our knowledge, the presence of mannose containing O-glycans in insects has only been described in D. melanogaster for the dystroglycan protein. Moreover, the O-mannosyltransferases that are responsible for the Oglycosylation were identified as POMT1 and POMT2. Recessive mutation in a pomt gene results in poorly viable flies with defects in muscle development, illustrating the influence of an aberration in O-mannosylation on normal development. Using the BLAST search algorithm, we were able to detect predicted protein sequences that are very homologous to POMT1 and POMT2, respectively, for T. castaneum, B. mori, A. mellifera as well as A. pisum. The construction of a phylogenetic tree for these predicted POMT proteins revealed that at least two distinct O-mannosyltransferases resembling POMT1 and POMT2 are conserved among the five insect species. Many proteins in the different glycoprotein sets have a known cytosolic localization such as actin, GSK2118436 Raf inhibitor tubulin or glycerol-3-phosphate dehydrogenase. Since POMTs are located in the lumen of the Golgi apparatus. However, several reports have demonstrated the existence of a cellular system involving retrograde transport of proteins from the ER to the cytosol. A dynamic and abundant O-glycosylation of serine and threonine was demonstrated for many cytoplasmic/nuclear proteins. For example, in Drosophila, post-translational O-GlcNAc modification was shown to be of importance for the regulation of Polycomb gene expression, while in vertebrates tubulin was even shown to contain sialyloligosaccharides. In addition, other types of cytoplasmic glycosylation may be present. Although at present the expression of a mannosyl transferase in the cytoplasm has never been shown, the addition of mannose residues or mannose containing oligosaccharides to the peptide backbone of cytoplasmic/nuclear proteins may occur in insects. Apart from its use as a tool for affinity chromatography, the snowdrop lectin was reported to exert strong insecticidal activity against different insect orders. Previously, midgut proteins such as ferritin, a-amylase or aminopeptidase were found to be targeted by mannose-binding plant lectins in several economically important pest insects. Indeed, these three midgut proteins were also found among the GNA binding glycoproteins in several insect species.