Differential Effects of Unnatural Sialic Acids on the Polysialylation of the Neural Cell Adhesion Molecule and Neuronal Behavior

Abstract

In this study we have examined how unnatural sialic acids can alter polysialic acid expression and influence the adhesive properties of the neural cell adhesion molecule (NCAM). Unnatural sialic acids are generated by metabolic conversion of synthetic N-acyl mannosamines and are typically incorporated into cell-surface glycoconjugates. However, N-butanoylmannosamine and Npentanoylmannosamine are effective inhibitors of polysialic acid (PSA) synthesis in stably transfected HeLa cells expressing NCAM and the polysialyltransferase STX. These cells were used as substrates to examine the effect of inhibiting PSA synthesis on the development of neurons derived from the chick dorsal root ganglion. N-butanoylmannosamine blocked polysialylation of NCAM and significantly reduced neurite outgrowth comparable with enzymatic removal of PSA by endoneuraminidases. As a result, neurite outgrowth was similar to that observed for non-polysialylated NCAM. In contrast, previous studies have shown that N-propanoyl sialic acid (SiaProp), generated from N-propanoylmannosamine, is readily accepted by polysialyltransferases and permits the extension of poly(SiaProp) on NCAM. Despite being immunologically distinct, poly(SiaProp) can promote neurite outgrowth similarly to natural polysialic acid. Thus, subtle structural differences in PSA resulting from the incorporation of SiaProp residues do not alter the antiadhesive properties of polysialylated NCAM.

Tissue-based metabolic labeling of polysialic acids in living primary hippocampal neurons

“…We envisioned that the primary neurons would tolerate external perturbations better when they were interconnected to each other and supplemented by tissue scaffolds and nutrients. Our hypothesis was supported indirectly by a previous report, in which ManNProp, a potent inhibitor of a polysialyltransferase (ST8SiaII) (41,46), was metabolized and expressed on cell membranes in hippocampal slice culture and in vivo, although at an extremely low efficiency, probably by another polysialyltransferase (ST8SiaIV) (7,47). The use of tissue culture was also hinted as a means of increasing the cellular tolerance to unnatural sialic acids and their precursors, although no data were presented (43).…”

“…For example, the growth rate of NG108-15 neuroblastoma-glioma hybrid cells was slowed sharply by feeding of 1 mM N-glycolyl-D-mannosamine pentaacetate, and the cell viability decreased to 76% after 70 h of feeding (11). In addition, 0.1 mM peracetylated ManNAc (Ac 4 ManNAc) derivatives were reported to be toxic to terminally differentiated human embryonic carcinoma NT2 cells (41). Although ManNProp (5 mM) was not cytotoxic, it altered the fate of cerebellar neurons in the microexplant culture containing neurons and glial cells (42).…”

Characterization of the Metabolic Flux and Apoptotic Effects of O-Hydroxyl- and N-Acyl-modified N-Acetylmannosamine Analogs in Jurkat Cells

“…In practice, sialic acid engineering methods have already been demonstrated to regulate cellular responses ranging from adhesion to proliferation (7,8) and have shown promise for use in biomedical applications such as inhibition of viral binding (9), modulation of the immune system (10), and selective delivery of diagnostic (11) or therapeutic agents (2) to cancer cells. ManNAc analogs can also modify the expression of polysialic acid (12,13), a linear polysaccharide composed of entirely of ␣-2,8-linked sialic acid, which is implicated in the complex neural processes (14), synaptic plasticity (15,16), and tumor metastasis (17). In addition to research and medical applications, ManNAc analogs also hold potential for biotechnological applications, such as increasing the product quality of recombinant sialoglycoconjugates (18).…”

Selection of Poly-α 2,8-Sialic Acid Mimotopes from a Random Phage Peptide Library and Analysis of Their Bioactivity

“…This has been achieved by genetic engineering (7), EndoN enzymatic digestion (6,10,28), or by the use of a small molecule, N-butanoylmannosamine, which is able to reversibly inhibit PSA synthesis in vitro (29,30). However, the feasibility of enhancing PSA activity has not yet been demonstrated either in vitro or in vivo.…”