Despite numerous studies on arylsulfatase A, the structure of its glycans is not well understood. It has been shown that the concentration of arylsulfatase A increases in the body fluids of patients with some forms of cancer, and the carbohydrate component of arylsulfatase A synthesized in tumor tissues and transformed cells undergoes increased sialylation, phosphorylation and sulfation. To understand the significance of any changes in the glycosylation of arylsulfatase A in cancer, it is important to know the structure of its carbohydrate component in normal tissue. In the present study we have analyzed carbohydrate moieties of human placental arylsylfatase A using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) followed by Western blotting on Immobilon P and on-blot deglycosylation using PNGase F for glycan release. Profiles of N-glycans were obtained by matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS). Oligosaccharides were sequenced using specific exoglycosidases, and digestion products were analyzed by MALDI MS and the computer matching of the resulting masses with those derived from a sequence database. Fifty picomoles (6 microg) of arylsulfatase A applied to the gel were sufficient to characterize its oligosaccharide content. The results indicated that human placental arylsulfatase A possesses only high-mannose-type oligosaccharides, of which almost half are core fucosylated. In addition, there was a minor species of high-mannose-type glycan bearing six mannose residues with a core fucose. This structure was not expected since high-mannose-type oligosaccharides basically have not been recognized as a substrate for the alpha1,6-fucosyltransferase.
The total protein glycosylation profile and specific activity of lysosomal enzymes were investigated in rat submandibular glands isolated from very young (1-month), young (1.5-months) and adult rats (3-months) rats. The specific activity of lysosomal hydrolases (i.e. acid phosphatase, arylsulfatases A and B, beta-N-acetyl-D-glucosaminidase, beta-galactosidase and beta-glucuronidase) decreased in parallel to increasing age of the animals. Furthermore, the thermal stability of acid phosphatase and beta-N-acetyl-D-glucosaminidase was influenced by the age of rats. Age-related changes in protein profile regarding the intensity of particular bands as well as the appearance of certain proteins limited to special age groups were also demonstrated as revealed by Coomassie and lectin staining. Moreover, the marked age-related increase in structures Man (alpha1-2, alpha1-3, alpha1-6) Man, Fuc (alpha1-6) GlcNAc as well as Gal (beta1-3) GlcNAc was observed, whereas staining with terminal NeuAc and GlcNAc showed an inverse correlation. The reaction with (beta1-6) branched N-glycans and Gal (beta1-3) Gal structures was limited to 1-month-old rats. No significant changes in a specific reaction with NeuAc (alpha2-3) Gal were observed. We speculate that the observed differences with respect to protein and glycosylation profiles between 1-month-old rats and older ones could be caused by a modification of the diet composition as well as by the functional and morphological maturation of the rat submandibular gland.
Cadherins are a large family of Ca2+dependent adhesion proteins. They are transmembrane or closely related to membrane glycoproteins localized in specialized adhesive junction. The expression of various cadherins may be concomitant with cancer progression steps and the term 'cadherin switch' has been created due to the observation of down-regulation of E-cadherin (suppressor of metastatic potential) and up-regulation of N-cadherin (promoter of metastatic potential) expression during tumour progression. These changes are thought to be closely related to epithelial-to-mesenchymal transition of cells of many different types of cancer including skin cancers, and accompany the increase of their motility and invasion abilities resulting in the metastasis formation. The cadherin polypeptide is a potential substrate for post-translational modification, for example, N-glycosylation, and its important role in the regulation of cadherin function has been described. The changed glycosylation of cadherins has been described in various skin cancers including melanoma and was consistent with cadherins' role in epithelial-to-mesenchymal transition. The detailed analysis of cadherin expression and cadherin-related glycosylation changes taking place during malignant transformation could be a key for better understanding of the nature of this process and may open new opportunities for the creation of more effective anticancer therapeutics and diagnostic tools.
Ectosomes are small heterogeneous membrane vesicles generated by budding from the plasma membrane in a variety of cell types and, more frequently, in tumor cells. They are shed into the extracellular space and are proposed as a novel form of intracellular communication in which information is transmitted from the originating cell to recipient cells without direct cell-to-cell contact. This review focuses on a single population of extracellular vesicles—ectosomes. We summarize recent studies of tumor-derived ectosomes which examine their biogenesis and protein cargo, and their influence on different aspects of cancer progression. We discuss possible clinical implications involving ectosomes as potential biomarkers, diagnostic tools and treatment targets in oncology. The unique composition of the molecules (cargo) that ectosomes carry, and their functional role, depends largely on the state of their originating cell. Through horizontal transfer of a variety of biologically active molecules (including proteins, lipids and nucleic acids) between donor and recipient cells, tumor-derived ectosomes may play functional roles in oncogenic transformation, tumor progression, invasion, metastasis, angiogenesis promotion, escape from immune surveillance, and drug resistance, thereby facilitating disease progression. The presence of tumor-derived ectosomes in body fluids such as the blood and urine of cancer patients makes them potentially useful prognostic and predictive biomarkers. Tumor-derived ectosomes also offer possible targets for multiple therapeutic strategies.
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