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Abstract

The MUC1 glycoprotein, epitectin, a component of the human bladder epithelium, was purified from human urine. Sedimentation equilibrium analysis and gel filtration using polysaccharide or protein standards revealed a polydisperse preparation with molecular weights ranging from about 0.9 to 1.3 x 10(6). This suggests that in the native state epitectin exists as aggregates of three or four monomer units of 350-400 kDa. Epitectin was found to have significant affinity to hexyl-, octyl- or phenyl agarose indicating that hydrophobic interactions and possibly carbohydrate-carbohydrate interactions may be responsible for the self-association. Chemical and enzymic deglycosylation of [125I]-labeled urine epitectin and metabolically labeled H.Ep.2 epitectin resulted in extremely polydisperse products. The buoyant densities of epitectin purified from urine and H.Ep.2 cells were found to be 1.39-1.40 g ml(-1), suggesting that the total carbohydrate content of these preparations is not significantly different. The O-linked saccharides of epitectin were fractionated by HPLC and analyzed by permethylation and FAB-MS. The neutral saccharides from both sources contain three common structures, namely Gal1 --> 3GalNAc, GlcNAc1 --> 6 (Gal1 --> 3) GalNAc and Gal1 --> 4GlcNAc --> 6 (Gal1 --> 3)GalNAc. The sialic acid of urine epitectin consisted entirely of N-acetylneuraminic acid. The two sources of epitectin, in vitro labeled on sialic acid, were found to have the same sialyl oligosaccharides but in different proportions. Metabolic labeling and N-glycanase susceptibility experiments firmly established the presence of N-linked saccharides in epitectin as minor components. The remarkable similarities in the total carbohydrate content, the carbohydrate composition and structures of saccharides between epitectin from urine, a non-malignant source, and H.Ep.2 cells is surprising in view of the prevailing view that MUC1 glycoproteins of cancer cells are underglycosylated compared to those produced by non-malignant cells.

Use of CA15‑3 for screening breast cancer: An antibody‑lectin sandwich assay for detecting glycosylation of CA15‑3 in sera

“…The O -glycan profiles of CA15-3 from breast milk, urine and breast carcinoma cell lines have been investigated ( 25 – 27 ). The glycan of CA15-3 in culture supernatants and cell lysates of breast carcinoma cell lines was found to have a truncated precursor structure ( 25 – 28 ). On the other hand, little attention has been paid to the N -glycan profiles of CA15-3 during carcinogenesis.…”

Glycosylation of uroplakins. Implications for bladder physiopathology

“…The expression of T and Tn glycotops on urothelium has been upregulated after stimulation of cell layers lining the urinary tract with epidermic growth factor (EGF) [35]. So far, certain mucins, such as MUC1 (epitectin) and proteoglycans, such as syndycan-1 have been isolated and characterized [36, 37]. As shown by Higuchi  et al [38] the rabbit mucins and glycoproteins of urothelium surface layers consisted of weakly sialylated and neutral oligosaccharides O-glycosidically linked to serine and threonine residues.…”

“…Expression of sialic acid on glycoproteins might indicate that the urothelium is not simply a inert barrier but can modulate the composition of soluble urinary proteins, which may play pathophysiological roles in the lower urinary tract [39]. Several urothelium-derived glycoproteins have been detected in urine of cows [1], rabbits [34, 36], pigs [40], rats [35] and people [37, 40, 41]. Using glycoproteomic technique Halim et al .…”

Recombinant MUC1 Probe Authentically Reflects Cell-specific O-Glycosylation Profiles of Endogenous Breast Cancer Mucin

“…Mono-and disialylated structures were also present but accounted for less than 25% of total glycans (15). Urinary MUC1, in contrast, exhibits an O-glycan profile with significantly shorter neutral and acidic glycans, which are based on core 1 as well as core 2 structures (16). According to three reports, tumor-associated glycoforms, which had been isolated from T47D (17) and BT 20 (18) breast carcinoma cell line supernatants or from cell lysates (19), were demonstrated to contain truncated precursor structures like core-GalNAc or the core 1 disaccharide Gal(␤1-3)GalNAc as well as its mono-and disialylated derivatives.…”