The expression of polysialic acid (polySia) on the neuronal cell adhesion molecule (NCAM) is called NCAM-polysialylation, which is strongly related to the migration and invasion of tumor cells and aggressive clinical status. Thus, it is important to select a proper drug to block tumor cell migration during clinical treatment. In this study, we proposed that lactoferrin (LFcinB11) may be a better candidate for inhibiting NCAM polysialylation when compared with CMP and low-molecular-weight heparin (LMWH), which were determined based on our NMR studies. Furthermore, neutrophil extracellular traps (NETs) represent the most dramatic stage in the cell death process, and the release of NETs is related to the pathogenesis of autoimmune and inflammatory disorders, with proposed involvement in glomerulonephritis, chronic lung disease, sepsis, and vascular disorders. In this study, the molecular mechanisms involved in the inhibition of NET release using LFcinB11 as an inhibitor were also determined. Based on these results, LFcinB11 is proposed as being a bifunctional inhibitor for inhibiting both NCAM polysialylation and the release of NETs.

The polysialyltransferases ST8SIA2 and ST8SIA4 and their product, polysialic acid (polySia), are known to be related to cancers and mental disorders. ST8SIA2 and ST8SIA4 have conserved amino acid (AA) sequence motifs essential for the synthesis of the polySia structures on the neural cell adhesion molecule. To search for a new motif in the polysialyltransferases, we adopted the in silico Individual Meta Random Forest program that can predict disease-related AA substitutions. The Individual Meta Random Forest program predicted a new eight-amino-acids sequence motif consisting of highly pathogenic AA residues, thus designated as the pathogenic (P) motif. A series of alanine point mutation experiments in the pathogenic motif (P motif) showed that most P motif mutants lost the polysialylation activity without changing the proper enzyme expression levels or localization in the Golgi. In addition, we evaluated the enzyme stability of the P motif mutants using newly established calculations of mutation energy, demonstrating that the subtle change of the conformational energy regulates the activity. In the AlphaFold2 model, we found that the P motif was a buried β-strand underneath the known surface motifs unique to ST8SIA2 and ST8SIA4. Taken together, the P motif is a novel buried β-strand that regulates the full activity of polysialyltransferases from the inside of the molecule.

The overexpression of polysialic acid (polySia) on neural cell adhesion molecules (NCAM) promotes hypersialylation, and thus benefits cancer cell migration and invasion. It has been proposed that the binding between the polysialyltransferase domain (PSTD) and CMP-Sia needs to be inhibited in order to block the effects of hypersialylation. In this study, CMP was confirmed to be a competitive inhibitor of polysialyltransferases (polySTs) in the presence of CMP-Sia and triSia (oligosialic acid trimer) based on the interactional features between molecules. The further NMR analysis suggested that polysialylation could be partially inhibited when CMP-Sia and polySia co-exist in solution. In addition, an unexpecting finding is that CMP-Sia plays a role in reducing the gathering extent of polySia chains on the PSTD, and may benefit for the inhibition of polysialylation. The findings in this study may provide new insight into the optimal design of the drug and inhibitor for cancer treatment.

The recent chemical identification of polysialylated glycoproteins in the jelly coat and on the cell surface of the sea urchin egg raises important questions about their biosynthesis and possible function. Using CMP-[14 C]-Neu5Ac as substrate and cell free preparations from eggs and embryos of the sea urchin Lytechinus pictus, we have identified a membrane associated CMP-Neu5Ac:poly-α2,8 sialosyl sialyltransferase (polyST) that transferred Neu5Ac from CMP-Neu5Ac to an endogenous acceptor membrane protein of approximately 38kDa. An average of five to six [14 C]-Neu5Ac residues were transferred to the glycan moiety of this protein. The membrane-associated polyST also catalyzed the polysialylation of several exogenous mammalian ganglioside acceptors, including GD3 . Given that no structurally similar naturally occurring polysialylated gangliosides have been described, nor were observed in the present study, we conclude that a single polyST activity catalyzes sialylation of the endogenous acceptor protein and the gangliosides. Using an excess of GD3 as an exogenous acceptor, it was established that the expression of the polyST in L. pictus embryos increased rapidly at the mesenchyme blastula stage and reached a maximum at the gastrula stage. The finding that this polyST in the sea urchin embryo is developmentally regulated raises the possibility that it may play a role in the changing cell and tissue interactions that occur during gastrulation and the early stages of spicule formation.

Alzheimer\'s disease (AD) is the most common form of dementia and is characterized by a substantial reduction of neuroplasticity. Our previous work demonstrated that neurons involved in memory function may lose plasticity because of decreased protein levels of polysialylated neural cell adhesion molecule (PSA-NCAM) in the entorhinal cortex (EC) of the human AD brain, but the cause of this decrease is unclear.
To investigate genes involved in PSA-NCAM regulation which may underlie its decrease in the AD EC.
We subjected neurologically normal and AD human EC sections to multiplexed fluorescent in situ hybridization and immunohistochemistry to investigate genes involved in PSA-NCAM regulation. Gene expression changes were sought to be validated in both human tissue and a mouse model of AD.
In the AD EC, a cell population expressing a high level of CALB2 mRNA and a cell population expressing a high level of PST mRNA were both decreased. CALB2 mRNA and protein were not decreased globally, indicating that the decrease in CALB2 was specific to a sub-population of cells. A significant decrease in PST mRNA expression was observed with single-plex in situ hybridization in middle temporal gyrus tissue microarray cores from AD patients, which negatively correlated with tau pathology, hinting at global loss in PST expression across the AD brain. No significant differences in PSA-NCAM or PST protein expression were observed in the MAPT P301S mouse brain at 9 months of age.
We conclude that PSA-NCAM dysregulation may cause subsequent loss of structural plasticity in AD, and this may result from a loss of PST mRNA expression. Due PSTs involvement in structural plasticity, intervention for AD may be possible by targeting this disrupted plasticity pathway.

Schizophrenia is a serious psychiatric disorder that affects the social life of patients. Psychiatric disorders are caused by a complex combination of genetic (G) and environmental (E) factors. Polysialylation represents a unique posttranslational modification of a protein, and such changes in neural cell adhesion molecules (NCAMs) have been reported in postmortem brains from patients with psychiatric disorders. To understand the G × E effect on polysialylated NCAM expression, in this study, we performed precise measurements of polySia and NCAM using a disrupted-in-schizophrenia 1 (DISC1)-mutant mouse (G), a mouse model of schizophrenia, under acute stress conditions (E). This is the first study to reveal a lower number and smaller length of polySia in the suprachiasmatic nucleus of DISC1 mutants relative to those in wild-type (WT) mice. In addition, an analysis of polySia and NCAM responses to acute stress in five brain regions (olfactory bulb, prefrontal cortex, suprachiasmatic nucleus, amygdala, and hippocampus) revealed that the pattern of changes in these responses in WT mice and DISC1 mutants differed by region. These differences could indicate the vulnerability of DISC1 mutants to stress.

Polysialic acid (polySia, PSA) is a unique constituent of the glycocalyx on the surface of bacterial and vertebrate cells. In vertebrates, its biosynthesis is highly regulated, not only in quantity and quality, but also in time and location, which allows polySia to be involved in various important biological phenomena. Therefore, impairments in the expression and structure of polySia sometimes relate to diseases, such as schizophrenia, bipolar disorder, and cancer. Some bacteria express polySia as a tool for protecting themselves from the host immune system during invasion. PolySia is proven to be a biosafe material; polySia, as well as polySia-recognizing molecules, are key therapeutic agents. This review first comprehensive outlines the occurrence, features, biosynthesis, and functions of polySia and subsequently focuses on the related diseases.

Polysialyltransferases (polySTs) are glycosyltransferases that synthesize polymers of sialic acid found in vertebrates and some bacterial pathogens. Bacterial polySTs have utility in the modification of therapeutic proteins to improve serum half-life, and the potential for tissue engineering. PolySTs are membrane-associated proteins and as recombinant proteins suffer from inherently low solubility, low expression levels and poor thermal stability. To improve their physicochemical and biochemical properties, we applied a directed evolution approach using a FACS-based ultrahigh-throughput assay as a simple, robust and reliable screening method. We were able to enrich a large mutant library and, in combination with plate-based high-throughput secondary screening, we discovered mutants with increased enzymatic activity and improved stability compared to the wildtype enzyme. This work presents a powerful strategy for the screening of directed evolution libraries of bacterial polySTs to identify better catalysts for in vitro polysialylation of therapeutics.

Sialic acid (Sia) is involved in many biological activities and commonly occurs as a monosialyl residue at the nonreducing terminal end of glycoconjugates. The loss of activity of UDP-GlcNAc2-epimerase/ManNAc kinase, which is a key enzyme in Sia biosynthesis, is lethal to the embryo, which clearly indicates the importance of Sia in embryogenesis. Occasionally, oligo/polymeric Sia structures such as disialic acid (diSia), oligosialic acid (oligoSia), and polysialic acid (polySia) occur in glycoconjugates. In particular, polySia, a well-known epitope that commonly occurs in neuroinvasive bacteria and vertebrate brains, is one of the most well-known and biologically/neurologically important glycotopes in vertebrates. The biological effects of polySia, especially on neural cell-adhesion molecules, have been well studied, and in-depth knowledge regarding polySia has been accumulated. In addition, the importance of diSia and oligoSia epitopes has been reported. In this chapter, the recent advances in the study of diSia, oligoSia, and polySia residues in glycoproteins in neurology, and their history, definition, occurrence, analytical methods, biosynthesis, and biological functions evaluated by phenotypes of gene-targeted mice, biochemical features, and related diseases are described.

Mental disorders, such as schizophrenia, bipolar disorder, and autism spectrum disorder, are challenging to manage, worldwide. Understanding the molecular mechanisms underlying these disorders is essential and required. Studies investigating such molecular mechanisms are well performed and important findings are accumulating apace. Based on the fact that these disorders are due in part to the accumulation of genetic and environmental risk factors, consideration of multi-molecular and/or multi-system dependent phenomena might be important. Acidic glycans are an attractive family of molecules for understanding these disorders, because impairment of the fine-tuned glycan system affects a large number of molecules that are deeply involved in normal brain function. One of the candidates of this important family of glycan epitopes in the brain is polysialic acid (PSA/polySia). PSA is a well-known molecule because of its role as an oncodevelopmental antigen and is also widely used as a marker of adult neurogenesis. Recently, several reports have suggested that PSA and PSA-related genes are associated with multiple mental disorders. The relationships among PSA, PSA-related genes, and mental disorders are reviewed here.