O-Glycosylation modifies and regulates a variety of intracellular proteins. Plakoglobin, which functions in both cell-cell
adhesion and signal transduction, is modified by O-glycosylation; however, the significance is unknown. To investigate the functional consequence of plakoglobin O-glycosylation, we cloned and overexpressed in keratinocytes murine O-GlcNAc transferase (mOGT). Over expression of mOGT in murine keratinocytes resulted in (i) glycosylation of plakoglobin and
(ii) increased levels of plakoglobin due to post-translational stabilization of plakoglobin. Additionally, overexpression
of mOGT in keratinocytes correlated with increased staining for cell-cell adhesion proteins and greater cell-cell adhesion.
These observations suggest that O-glycosylation functions to regulate the post-translational stability of plakoglobin and keratinocyte cell-cell adhesion.
"However, this has never been demonstrated. In addition to FN, other glycoproteins associated with cell adhesion, such as cytokeratins (125, 126), plakoglobin (127, 128), E-cadherin (129), and beta-catenin (63, 130) are known to express their peculiar O-glycophenotypes. Although the EMT process is deeply associated with glycosylation changes, there is little information regarding the impact of the atypical O-linked glycans adopted by these glycoproteins in cancer cells undergoing EMT. "
[Show abstract][Hide abstract] ABSTRACT: Glycosylation changes are a feature of disease states. One clear example is cancer cells, which commonly express glycans at atypical levels or with different structural attributes than those found in normal cells. Epithelial-mesenchymal transition (EMT) was initially recognized as an important step for morphogenesis during embryonic development, and is now shown to be one of the key steps promoting tumor metastasis. Cancer cells undergoing EMT are characterized by significant changes in glycosylation of the extracellular matrix (ECM) components and cell-surface glycoconjugates. Current scientific methodology enables all hallmarks of EMT to be monitored in vitro and this experimental model has been extensively used in oncology research during the last 10 years. Several studies have shown that cell-surface carbohydrates attached to proteins through the amino acids, serine, or threonine (O-glycans), are involved in tumor progression and metastasis, however, the impact of O-glycans on EMT is poorly understood. Recent studies have demonstrated that transforming growth factor-beta (TGF-β), a known EMT inducer, has the ability to promote the up-regulation of a site-specific O-glycosylation in the IIICS domain of human oncofetal fibronectin, a major ECM component expressed by cancer cells and embryonic tissues. Armed with the knowledge that cell-surface glycoconjugates play a major role in the maintenance of cell homeostasis and that EMT is closely associated with glycosylation changes, we may benefit from understanding how unusual glycans can govern the molecular pathways associated with cancer progression. This review initially focuses on some well-known changes found in O-glycans expressed by cancer cells, and then discusses how these alterations may modulate the EMT process.
Frontiers in Oncology 03/2014; 4(33):59. DOI:10.3389/fonc.2014.00059
"O-glycosylation of classical cadherins is not well studied, but Zhu and col- leagues  studied E-cadherin trafficking to and from the membrane under stress conditions. In contrast to the observations of Hu et al  , in which enhanced O-glycosylation of plakoglobin stabilized adherens junctions and desmosomes, Zhu and colleagues  found that ER stress induced O-glycosylation of newly synthesized E-cadherin as well as of β-catenin resulted in failure of this newly synthesized E-cadherin to be transported to the cell surface with subsequent decreased intercellular adhesion. These observations suggest that the effects of O-glycosylation are likely to be variable and context dependent. "
[Show abstract][Hide abstract] ABSTRACT: Post-translational protein modification, including phosphorylation, is generally quick and reversible, facilitating rapid biologic adjustments to altered cellular physiologic demands. In addition to protein phosphorylation, other post-translational modifications have been identified. Intracellular protein O-glycosylation, the addition of the simple sugar O-linked N-acetylglucosamine (O-GlcNAc) to serine/threonine residues, is a relatively recently identified post-translational modification that has added to the complexity by which protein function is regulated. Two intracellular enzymes, O-GlcNAc transferase and O-GlcNAcase, catalyze the addition and removal, respectively, of O-GlcNAc to serine and threonine side-chain hydroxyl groups. Numerous proteins, including enzymes, transcription factors, receptors and structural proteins have been shown to be modified by intracellular O-glycosylation. In this review, the mechanism and relevance of O-GlcNAc protein modification are discussed in the context of cell adhesion and several representative diseases.
"In particular, the possible competition between O-GlcNAcylation and phosphorylation (OGT/kinases targeting the same serines/threonines) has become a topic of vigorous research, due to the possibility of O-GlcNAc regulating a number of phosphorylation-dependent signal transduction pathways. Many studies have used knockout/knockdown/overexpression approaches to modulate levels of the OGA/OGT proteins, thus testing the effects of hyper/hypo-O-GlcNAcylation on particular cellular processes (e.g., Slawson et al., 2005; Hu et al., 2006; Yang et al., 2008). However, an alternative approach has been to use small-molecule inhibitors on live cells to inhibit OGA, inducing hyper-O-GlcNAcylation (potent inhibitors of OGT are not yet available). "
[Show abstract][Hide abstract] ABSTRACT: Streptozotocin is a natural product that selectively kills insulin-secreting beta cells, and is widely used to generate mouse models of diabetes or treat pancreatic tumors. Several studies suggest that streptozotocin toxicity stems from its N-nitrosourea moiety releasing nitric oxide and possessing DNA alkylating activity. However, it has also been proposed that streptozotocin induces apoptosis by inhibiting O-GlcNAcase, an enzyme that, together with O-GlcNAc transferase, is important for dynamic intracellular protein O-glycosylation. We have used galacto-streptozotocin to chemically dissect the link between O-GlcNAcase inhibition and apoptosis. Using X-ray crystallography, enzymology, and cell biological studies on an insulinoma cell line, we show that, whereas streptozotocin competitively inhibits O-GlcNAcase and induces apoptosis, its galacto-configured derivative no longer inhibits O-GlcNAcase, yet still induces apoptosis. This supports a general chemical poison mode of action for streptozotocin, suggesting the need for using more specific inhibitors to study protein O-GlcNAcylation.
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