[The tissue localization and secretion of mucin-D in Drosophila melanogaster]. / Tkanevaia lokalizatsiia i sekretsiia mutsina-D Drosophila melanogaster.

Glycoprotein with biochemical characteristics that allow us to classify it as a glycoprotein of mucin-type was isolated from cultured embryonic cells of Drosophila melanogaster. This is the first finding of mucin-type glycoprotein in insects. Using high-affinity monoclonal antibodies against a carbohydrate epitope, we demonstrated that the accumulation of this glycoprotein in the culture fluid of Drosophila cell line and cultured cells of other insects was inhibited by secretion inhibitors. 20-hydroxyecdysone, a hormone responsible for molting and metamorphosis in insects, inhibits the accumulation of this glycoprotein in the culture fluid, as well as in cells of Drosophila Dm cell line. In another cell line (Schneider-2), where there was practically no secretion of this glycoprotein, the hormone induced its increased accumulation in the cells. Mucin glycoproteins recognized by monoclonal antibodies have been found in embryos, imaginal disc, fat body, testicles, and ovaries, but not in salivary glands or muscles of Drosophila.

Insect mucin-type glycoprotein: immunodetection of the O-glycosylated epitope in Drosophila melanogaster cells and tissues.

A mucin-type glycoprotein (GP) from cultured embryonic cells of Drosophila melanogaster was isolated and used to raise monoclonal antibodies (MAbs). Epitope(s) recognized by MAbs were sensitive to the treatment by O-glycanase, which specifically cleaves off O-linked mucin-type Gal(beta 1,3)GalNAc disaccharide, representing the major part of the carbohydrate moiety of Drosophila GP. Using high-affinity MAbs against carbohydrate epitopes of the Drosophila mucin GP we demonstrated its accumulation in culture medium, as well as in cultured cells, which proved to be regulated by 20-hydroxyecdysone. Mucin GPs carrying Gal(beta 1,3)GalNAc disaccharide recognized by the MAbs were immunochemically localized in several Drosophila tissues of ectodermal, mesodermal and germ line origin, including epidermal and follicle cells capable of their secretion.

[The tissue localization of "chitinoprotein", detectable by using specific antibodies, in the development of Drosophila melanogaster]. / Tkanevaia lokalizatsiia "khitinoproteina", vyiavliaemaia s pomoshch'iu spetsificheskikh antitel, v razvitii Drosophila melanogaster.

The biosynthesis of cognate glycoproteins with chitinase-sensitive carbohydrate moiety ("chitinoproteins") was detected after incubation of cultured cells of different insect species with 3H-glucosamine (Kramerov et al., Insect Biochem. v. 20; 769-775, 1990). It was also demonstrated that production of the specific chitinoprotein takes place during the development of D. melanogaster as revealed by immunoblotting and autoradiographic analysis of crude tissue extracts. An investigation of the developmental pattern of tissue localization of Drosophila chitinoprotein was performed using antibodies raised in rabbit after immunization with a purified preparation of the chitinoprotein (ChiP) from Drosophila embryonic cultured cells. The paraffin-embedded thin (5 microns) sections of organisms fixed in Bouin fixative were stained immunohistochemically with primary antibodies and peroxidase-conjugated secondary antibodies followed by enhancement of the precipitated DAB product with osmium tetroxide. Preimmune serum and antiserum preadsorbed with the purified ChiP preparation were used as negative controls yielding no specific staining of tissue sections. Negative staining with specific anti-ChiP antibodies was demonstrated for salivary glands, gut, muscles, central and peripheral nerve system and some other tissues. A complex pattern of tissue-specific ChiP localization in a variety of tissues of ectodermal, mesodermal and germ line origin was revealed. The mesodermal derivatives--hemocytes and oenocytes capable of producing the components of cuticle as well as epidermal cells of larvae and imago clearly demonstrated staining of cytoplasmic vesicles, which in the latter case were exocytosed and included into the newly formed endocuticle. Another cell type known to produce cuticle--epithelial cells of imaginal discs (primordia of adult organs)--were also stained with antibodies. One can suppose that ChiP is involved in biogenesis of insect cuticle, probably, as a protein precursor of chitin formation. It was quite surprising to observe a rather strong staining of fat body cells and follicle cells of adult ovaries. The follicular epithelium secreted the stained granules into a growing oocyte that accumulated large amounts of this immunopositive material until transformation into a mature egg. In an early embryo ChiP is localized in blastodermal and pole cells, but not in yolk. This is, probably, the result of segregation of ChiP to the periphery of an egg during the final stage of its maturation and subsequent cellularization in the beginning of embryogenesis. Later ChiP can be found in ectodermal cells and hemocyte-like cells. It should be noted that not all amounts of ChiP detected in embryos are maternally inherited, for an active ChiP biosynthesis takes place in dissociated embryonic cells after incubation with labelled sugar precursor.(ABSTRACT TRUNCATED AT 400 WORDS)