Sialic acids are widely expressed as terminal carbohydrates on glycoconjugates of eukaryotic cells. They are involved in a variety of cellular functions, such as cell adhesion or signal recognition. The key enzyme of sialic acid biosynthesis is the bifunctional UDP-N-acetylglucosamine-2-epimerase/N-acetylmannosamine kinase (GNE), which catalyzes the first two steps of sialic acid biosynthesis in the cytosol. Previously, we have shown that inactivation of the GNE by gene targeting causes early embryonic lethality in mice, whereas heterozygous GNE-deficient mice are vital. In this study we compared the amount of membrane-bound sialic acids of wildtype mice with those of heterozygous GNE-deficient mice. For that we quantified membrane-bound sialic acid concentration in various organs of wildtype- and heterozygous GNE-deficient mice. We found an organ-specific reduction of membrane-bound sialic acids in heterozygous GNE-deficient mice. The overall reduction was 25%. Additionally, we analyzed transferrin and polysialylated neural cell adhesion molecule (NCAM) by one- or two-dimensional gel electrophoresis. Transferrin-expression was unchanged in heterozygous GNE-deficient mice; however the isoelectric point of transferrin was shifted towards basic pH, indicating a reduced sialylation. Furthermore, the expression of polysialic acids on NCAM was reduced in GNE-deficient mice.

Sialic acid (Sia) is expressed as terminal sugar in many glycoconjugates and plays an important role during development and regeneration. Addition of homopolymers of Sia (polysialic acid; polySia/PSA) is a unique and highly regulated posttranslational modification of the neural cell adhesion molecule (NCAM). The presence of polySia affects NCAM-dependent cell adhesion and plays an important role during brain development, neural regeneration, and plastic processes including learning and memory. PolySia-NCAM is expressed on several neuroendocrine tumors of high malignancy and correlates with poor prognosis. Two closely related enzymes, the polysialyltransferases ST8SiaII and ST8SiaIV, catalyze the biosynthesis of polySia. This review summarizes recent knowledge on Sia biosynthesis and the correlation between Sia biosynthesis and polysialylation of NCAM and report on approaches to modify the degree of polySia on NCAM in vitro and in vivo. First, we describe the inhibition of polysialylation of NCAM in ST8SiaII-expressing cells using synthetic Sia precursors. Second, we demonstrate that the key enzyme of the Sia biosynthesis (UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase) regulates and limits the synthesis of polySia by controlling the cellular Sia concentration. Keywords: mannosamines, polysialyltransferase, sialic acid. In bacteria, polySia is often used as capsular polysaccharide, including neuroinvasive Escherichia coli K1 or Neisseria meningitidis Gp B (Jennings 1988). However, there are only three molecules, which are known to express polySia in mammals. These are two transmembrane glycoproteins, the neural cell adhesion molecule (NCAM) (Thiery et al. 1977;Finne et al. 1983;Ronn et al. 2000) and the voltagesensitive sodium channel alpha-subunit (Zuber et al. 1992) and a secreted glycoprotein in mammalian milk (CD36) (Yabe et al. 2003). However, the major carrier of polySia in mammals is NCAM (see Fig. 1). This was demonstrated by the fact that NCAM-deficient mice are polySia-negative. PolySia is synthesized in the Golgi apparatus by the activity of two closely related enzymes, the polysialyltransferases ST8SiaIV (Eckhardt et al. 1995;Nakayama et al. 1995) and ST8SiaII (Scheidegger et al. 1995;Yoshida et al. 1995). PolySia expression is high during embryogenesis, peaks in the perinatal phase and decreases rapidly in the adult, were § This work is dedicated to our friend Steven E. Pfeiffer.Address correspondence and reprint requests to Rüdiger Horstkorte, Institut für Physiologische Chemie, Medizinische Fakultät, Martin-Luther-Universität Halle-Wittenberg, Hollystrasse 1, 06114 Halle, Saale, Germany. E-mail: ruediger.horstkorte@medizin.uni-halle.de 1 These authors contributed equally to this review and are listed in alphabetical order.Abbreviations used: ES, embryonic stem; GNE, UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase; ManNAc, N-acetyl Dmannosamine; NCAM, neural cell adhesion molecule; polySia/PSA, polysialic acid; Sia, Sialic acid; VPA, valproic acid.