O- and N-glycosylation are the most common and complex of the post-translational modifications. Both are enzymatic processes and it was suggested that both could be regulated by cAMP cascade at the early stages. N-glycosylation starts with the formation of lipid-linked oligosaccharides and this process is catalysed by crucial glycosyltransferase - dolichol phosphate mannose synthase. The results of several studies strongly suggest that the cAMP acting through a cAMP-dependent protein kinase A-mediated protein phosphorylation/dephosphorylation cycle may modulate activation of this enzyme. It was shown that cAMP can also up regulate another enzyme involved in phosphodolichole synthesis - cis-prenyltransferase. The mechanism acting here is the alteration of the rate of its gene expression. cAMP cascade is also involved in regulation of O-glycosylation since phosphorylation of human glutamine:fructose-6-phosphate amidotransferase results in depletion of O-GlcNAc structure formation. These observation suggested an important role of GPCRs and their ligand in regulation of N- and O-glycan synthesis.
N-glycosylation of integrins plays an important role in cancer progression. Increased αvβ3 integrin expression during melanoma progression is well-documented but the role of its glycans in tumorigenesis is still poorly understood. In the present study we used the WM793 primary melanoma cell line and its highly metastatic variant, WM1205Lu, to examine αvβ3 glycosylation. Lectin precipitation, enzyme digestion and the use of swainsonine (SW) showed that αvβ3 integrin glycosylation differs significantly between primary and metastatic melanoma cells. High-mannose structures and complex glycans with bisecting N-acetylglucosamine (GlcNAc) were more abundant in both subunits of primary cells. We also observed a shift in the sialylation of αvβ3 integrin related to reduction of α2-6-linked sialic acid expression and an increase of α2-3 sialylation of both subunits in melanoma progression. Metastatic melanoma migration on vitronectin (VN) was reduced in the presence of antibody against αvβ3 and the lectins phytohemagglutinin-L (PHA-L), Sambucus nigra agglutinin (SNA) and Maackia amurensis (MAA) in woundhealing assays. Our results show that the acquisition of metastatic competence by melanoma cells is accompanied by alteration of αvβ3 integrin glycosylation and that both αvβ3 and β1-6-branched sialylated complex-type N-glycans promote metastatic melanoma migration on VN.
We have examined the diversity between primary uveal (92-1 and Mel202) and cutaneous (FM55P and IGR-39) melanoma cells in their interaction with vitronectin, and established the effect of integrins and β1,6-branched N-oligosaccharides on this process. The adhesion level of uveal melanoma cells to vitronectin was at least twice lower than that of cutaneous ones, but all cells tested repaired scratch wounds on vitronectin-coated surfaces with similar speed. Swainsonine treatment, by reducing the amount of β1,6-branches, significantly decreased cell attachment in all cases, but reduction of wound healing efficiency was compromised only in cutaneous melanoma cell. Functional blocking antibodies used in adhesion and migration assays revealed that integrin αvβ3 was strongly involved in adhesion and migration only in cutaneous melanoma cells, but its role here was less pronounced than that of integrin αvβ5. However, in uveal melanoma the specific anti-αvβ5 integrin antibody had no impact on migration speed. Therefore, the anti-α3β1 integrin antibody was used in order to explain the nature of uveal melanoma interaction with vitronectin, which caused a mild decrease in adhesion efficiency and reduced their motility. Expression of αvβ5 integrin differed between the cell lines, but there was no distinct pattern to distinguish uveal melanoma from cutaneous melanoma. In conclusion, αvβ5, but not αvβ3 integrin is heavily involved in uveal melanoma cell interaction with vitronectin. The role of β1,6-branched N-glycans in the adhesion, but not during migration, of all cells to vitronectin has been confirmed.
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.
Melanoma is the most aggressive of all skin cancers and is exceptionally resistant to therapies. During melanoma progression, cancer cells reprogram their proliferation and survival pathways and achieve resistance to treatment-induced apoptosis. Galectin-3 (gal-3) is a member of the lectin family and is involved in such biological processes as cell adhesion, growth and differentiation, the cell cycle, and apoptosis. Gal-3 also plays an important role in tumor development and metastasis. The relationship between gal-3 expression and these processes is specific to the tumor type and the stage of cancer progression. The biological functions of gal-3 depend on its localization in the cell. In the present study, human metastatic melanoma A-375 cells, characterized by weak endogenous expression of gal-3, were transfected with gal-3 cDNA and cisplatin-induced apoptosis was measured. Data from AnnexinV and mitochondrial membrane potential analysis revealed that gal-3 did not protect the A-375 melanoma cells against cisplatin. This result probably is associated with its nuclear localization in the cells.
The endothelium is made of a layer of cells located on the inner surface of blood and lymphatic vessels. It is considered as the integral organ of human body. It is formed of endothelial cells anchoring in extracellular matrix and at the same time tightly connected one with another through various types of cellular junctions. There is a number of adhesion molecules: proteins and glycans, taking part in this cell-to-cell and cell-to-ECM adhesion, and many of them are involved in variety of biomechanical and biochemical processes occurring in the endothelial cells. One of the most characteristic feature of the endothelial cells is the presence of the Weible-Palade bodies as well as numerous caveolae and cellular membrane-anchored enzymes regulating blood clotting. It has been shown that there is a specific type of endothelial cells called high endothelial cells, which are characteristic for the lymphatic vessels. This high endothelium facilitates physiological transmigration of leukocytes form vessels to surrounding lymphatic tissue. The endothelium is known to have the uppermost role in maintaining the constant flow of blood in vessels through its regulatory influence on blood clotting processes. The endothelial cells produces a wide variety of factors including: anticoagulants, vasodilators - causing relaxation of vessels, vasoconstrictors - leading to constriction of blood vessels, as well as pro- and antiangiogenic factors that regulates formation of new blood vessels. Therefore, it seems to be very important to fully analyze the physiology of endothelium. In vitro cultures of endotelial cells, despite their limitations, seems to be and reliable model s for biochemical and molecular analysis of this tissue under physiological and pathological conditions.
'%3e%3cpath fill='%23012169' d='M0 0v30h60V0z'/%3e%3cpath stroke='%23fff' stroke-width='6' d='m0 0 60 30m0-30L0 30'/%3e%3cpath stroke='%23C8102E' stroke-width='4' d='m0 0 60 30m0-30L0 30' clip-path='url(%23b)'/%3e%3cpath stroke='%23fff' stroke-width='10' d='M30 0v30M0 15h60'/%3e%3cpath stroke='%23C8102E' stroke-width='6' d='M30 0v30M0 15h60'/%3e%3c/g%3e%3c/svg%3e)
 scale(5.56106)'%3e%3ccircle r='1'/%3e%3cg id='d'%3e%3cg id='c'%3e%3cpath id='b' d='M.195-.98 0-1.39l-.195.408' transform='rotate(11.25)'/%3e%3cuse xlink:href='%23b' transform='rotate(22.5)'/%3e%3cuse xlink:href='%23b' transform='rotate(45)'/%3e%3c/g%3e%3cuse xlink:href='%23c' transform='rotate(67.5)'/%3e%3c/g%3e%3cuse xlink:href='%23d' transform='scale(-1 1)'/%3e%3c/g%3e%3cg transform='translate(0 58.787) scale(8.1434)'%3e%3ccircle r='1'/%3e%3cg id='g'%3e%3cg id='f'%3e%3cpath id='e' d='M.259.966 0 1.576l-.259-.61'/%3e%3cuse xlink:href='%23e' transform='rotate(180)'/%3e%3c/g%3e%3cuse xlink:href='%23f' transform='rotate(90)'/%3e%3c/g%3e%3cuse xlink:href='%23g' transform='rotate(30)'/%3e%3cuse xlink:href='%23g' transform='rotate(60)'/%3e%3c/g%3e%3c/g%3e%3c/svg%3e)