Simple and complex carbohydrates (glycans) have long been known to play major metabolic, structural and physical roles in biological systems. Targeted microbial binding to host glycans has also been studied for decades. But such biological roles can only explain some of the remarkable complexity and organismal diversity of glycans in nature. Reviewing the subject about two decades ago, one could find very few clear-cut instances of glycan-recognition-specific biological roles of glycans that were of intrinsic value to the organism expressing them. In striking contrast there is now a profusion of examples, such that this updated review cannot be comprehensive. Instead, a historical overview is presented, broad principles outlined and a few examples cited, representing diverse types of roles, mediated by various glycan classes, in different evolutionary lineages. What remains unchanged is the fact that while all theories regarding biological roles of glycans are supported by compelling evidence, exceptions to each can be found. In retrospect, this is not surprising. Complex and diverse glycans appear to be ubiquitous to all cells in nature, and essential to all life forms. Thus, >3 billion years of evolution consistently generated organisms that use these molecules for many key biological roles, even while sometimes coopting them for minor functions. In this respect, glycans are no different from other major macromolecular building blocks of life (nucleic acids, proteins and lipids), simply more rapidly evolving and complex. It is time for the diverse functional roles of glycans to be fully incorporated into the mainstream of biological sciences.

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Biological Roles of Glycans

Protein glycosylation is a ubiquitous post-translational modification found in all domains of life Despite their significant complexity in animal systems, glycan structures have crucial biological and physiological roles, from contributions in protein folding and quality control to involvement in a large number of biological recognition events As a result, they impart an additional level of 'information content' to underlying polypeptide structures Improvements in analytical methodologies for dissecting glycan structural diversity, along with recent developments in biochemical and genetic approaches for studying glycan biosynthesis and catabolism, have provided a greater understanding of the biological contributions of these complex structures in vertebrates

Vertebrate protein glycosylation: diversity, synthesis and function

Chemical Diversity in the Sialic Acids and Related α-Keto Acids: An Evolutionary Perspective

Recognition molecules of the immunoglobulin superfamily have important roles in neuronal interactions during ontogeny, including migration, survival, axon guidance and synaptic targeting. Their downstream signal transduction events specify whether a cell changes its place of residence or projects axons and dendrites to targets in the brain, allowing the construction of a dynamic neural network. A wealth of recent discoveries shows that cell adhesion molecules interact with attractant and repellent guidance receptors to control growth cone and cell motility in a coordinate fashion. We focus on the best-studied subclasses, the neural cell adhesion molecule NCAM and the L1 family of adhesion molecules, which share important structural and functional features. We have chosen these paradigmatic molecules and their interactions with other recognition molecules as instructive for elucidating the mechanisms by which other recognition molecules may guide cell interactions during development or modify their function as a result of injury, learning and memory.

Neural recognition molecules of the immunoglobulin superfamily: signaling transducers of axon guidance and neuronal migration

Neoplastic transformation results in a wide variety of cellular alterations that impact the growth, survival, and general behavior of affected tissue. Although genetic alterations underpin the development of neoplastic disease, epigenetic changes can exert an equally significant effect on neoplastic transformation. Among neoplasia-associated epigenetic alterations, changes in cellular glycosylation have recently received attention as a key component of neoplastic progression. Alterations in glycosylation appear to not only directly impact cell growth and survival but also facilitate tumor-induced immunomodulation and eventual metastasis. Many of these changes may support neoplastic progression, and unique alterations in tumor-associated glycosylation may also serve as a distinct feature of cancer cells and therefore provide novel diagnostic and even therapeutic targets.

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Protein Glycosylation in Cancer

The structures of cellular oligosaccharides are determined by a series of processing reactions catalyzed by Golgi glycosidases and glycosyltransferases. While there are subtle cell type differences in Golgi enzyme subcompartmentation, in general, glycosylation enzymes are localized within the Golgi cisternae in the same sequence in which they act to modify oligosaccharide substrates. The possibility that this enzyme subcompartmentation may control the types of oligosaccharides expressed by a cell has led to an interest in the signals and mechanisms directing enzyme localization in the Golgi cisternae. All glycosidases and glycosyltransferases characterized thus far have very little sequence homology that might suggest a common Golgi retention signal, but they do share a similar domain structure. They are all type II transmembrane proteins consisting of an amino terminal cytoplasmic tail, a signal anchor transmembrane domain, a stem region, and a large luminal catalytic domain. Their lack of sequence homology suggests that these proteins' Golgi retention signals are not linear amino acid sequences, but most likely involve general characteristics or conformations of larger protein domains. The peptide sequences required for Golgi retention of the N-acetylglucosaminyltransferase I (GlcNAcTI), beta 1,4-galactosyltransferase (GalT) and alpha 2,6-sialytransferase (ST) have been extensively studied. To do this, researchers created mutant and chimeric proteins, expressed these in tissue culture cells, and localized these proteins using immunofluorescence microscopy or immunoelectron microscopy. The cell surface expression of deletion mutants suggested that the deleted sequences were necessary for Golgi retention. Then, if these sequences were fused to a non-Golgi reporter protein and this chimeric or hybrid protein was retained in the Golgi, then these sequences were also sufficient for Golgi retention. Due to differences in reporter proteins used to construct these chimeric proteins, different cell types used for protein expression, different levels of protein expression, and different methods of cell surface protein detection, these experiments have led to somewhat confusing results. However, in general, it appears that the GalT relies primarily on its transmembrane domain for Golgi retention, while the GlcNAcTI and ST have requirements for their transmembrane regions, sequences flanking these regions, and luminal stem sequences. Based on these results, two potential Golgi retention mechanisms have been proposed and are now being tested. The observation that glycosyltransferase transmembrane domains are frequently sufficient for Golgi retention has led to the first of these models, the bilayer thickness model. This model proposes that the shorter transmembrane domains of Golgi proteins prevent them from entering cholesterol-rich transport vesicles destined for the plasma membrane, and that this leads to Golgi retention. The second of these models is supported by the role of multiple protein domains in the Golgi retention of some proteins. This model, the oligomerization/ kin recognition model of Golgi retention, proposes that the formation of insoluble protein homo-oligomers or very large hetero-oligomers prevents protein movement into transport vesicles destined for later compartments. Initial work suggests that the bilayer thickness mechanism may play a role in the retention of some Golgi retained proteins; however, it is not the sole retention mechanism. Other evidence suggests that an oligomerization/kin recognition mechanism may be more common, but definitive proof for its general use in Golgi protein retention is lacking. More research is required to further elucidate the sequences and particularly the mechanisms of Golgi retention. (ABSTRACT TRUNCATED)

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Golgi localization of glycosyltransferases: more questions than answers

Sialylated N-glycans play essential roles in the immune system, pathogen recognition and cancer. This review approaches the sialylation of N-glycans from three perspectives. The first section focuses on the sialyltransferases that add sialic acid to N-glycans. Included in the discussion is a description of these enzymes' glycan acceptors, conserved domain organization and sequences, molecular structure and catalytic mechanism. In addition, we discuss the protein interactions underlying the polysialylation of a select group of adhesion and signaling molecules. In the second section, the biosynthesis of sialic acid, CMP-sialic acid and sialylated N-glycans is discussed, with a special emphasis on the compartmentalization of these processes in the mammalian cell. The sequences and mechanisms maintaining the sialyltransferases and other glycosylation enzymes in the Golgi are also reviewed. In the final section, we have chosen to discuss processes in which sialylated glycans, both N- and O-linked, play a role. The first part of this section focuses on sialic acid-binding proteins including viral hemagglutinins, Siglecs and selectins. In the second half of this section, we comment on the role of sialylated N-glycans in cancer, including the roles of β1-integrin and Fas receptor N-glycan sialylation in cancer cell survival and drug resistance, and the role of these sialylated proteins and polysialic acid in cancer metastasis.

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Sialylation of N-glycans: mechanism, cellular compartmentalization and function.

As an anti-adhesive, a reservoir for key biological molecules, and a modulator of signaling, polysialic acid (polySia) is critical for nervous system development and maintenance, promotes cancer metastasis, tissue regeneration and repair, and is implicated in psychiatric diseases. In this review, we focus on the biosynthesis and functions of mammalian polySia, and the use of polySia in therapeutic applications. PolySia modifies a small subset of mammalian glycoproteins, with the neural cell adhesion molecule, NCAM, serving as its major carrier. Studies show that mammalian polysialyltransferases employ a unique recognition mechanism to limit the addition of polySia to a select group of proteins. PolySia has long been considered an anti-adhesive molecule, and its impact on cell adhesion and signaling attributed directly to this property. However, recent studies have shown that polySia specifically binds neurotrophins, growth factors, and neurotransmitters and that this binding depends on chain length. This work highlights the importance of considering polySia quality and quantity, and not simply its presence or absence, as its various roles are explored. The capsular polySia of neuroinvasive bacteria allows these organisms to evade the host immune response. While this "stealth" characteristic has made meningitis vaccine development difficult, it has also made polySia a worthy replacement for polyetheylene glycol in the generation of therapeutic proteins with low immunogenicity and improved circulating half-lives. Bacterial polysialyltransferases are more promiscuous than the protein-specific mammalian enzymes, and new studies suggest that these enzymes have tremendous therapeutic potential, especially for strategies aimed at neural regeneration and tissue repair.

Karen J. Colley

Papers

Golgi localization of glycosyltransferases: more questions than answers

Sialylation of N-glycans: mechanism, cellular compartmentalization and function.

Polysialic acid: biosynthesis, novel functions and applications.

In Vivo Autopolysialylation and Localization of the Polysialyltransferases PST and STX

Hyposialylation of integrins stimulates the activity of myeloid fibronectin receptors.