Differential expression of five sialyltransferase genes in human tissues.

The molecular and cell biology of glycosyltransferases

Primary structure of Gal beta 1,3(4)GlcNAc alpha 2,3-sialyltransferase determined by mass spectrometry sequence analysis and molecular cloning. Evidence for a protein motif in the sialyltransferase gene family.

Sialic acids on vertebrate cell surfaces mediate many biological roles. Altered expression of certain sialic acid types or their linkages can have prognostic significance in human cancer. A classic but unexplained example is enhanced alpha2-6-sialylation on N-glycans resulting from overexpression of the Golgi enzyme beta-galactoside:alpha2-6-sialyltransferase (ST6Gal-I). Previous data supporting a role for the resulting Sia alpha 2-3Gal beta 1-4GlcNAc (Sia6LacNAc) structure in tumor biology were based on in vitro studies in transfected carcinoma cells, in which increased Sia6LacNAc on beta1-integrins enhanced their binding to ligands, and stimulated cell motility. Here, we examine for the first time the in vivo role of the ST6Gal-I enzyme in the growth and differentiation of spontaneous mammary cancers in mice transgenic for a mouse mammary tumor virus promoter-driven polyomavirus middle T antigen, a tumor in which beta1-integrin function is important for tumorigenesis and in maintaining the proliferative state of tumor cells. Tumors induced in St6gal1-null animals were more differentiated compared with those in the wild-type background, both by histologic analysis and by protein expression profiles. Furthermore, we show the St6gal1-null tumors have selectively altered expression of genes associated with focal adhesion signaling and have decreased phosphorylation of focal adhesion kinase, a downstream target of beta1-integrins. This first in vivo evidence for a role of ST6Gal-I in tumor progression was confirmed using a novel approach, which conditionally restored St6gal1 in cell lines derived from the null tumors. These findings indicate a role for ST6Gal-I as a mediator of tumor progression, with its expression causing a less differentiated phenotype, via enhanced beta1-integrin function.

https://cancerres.aacrjournals.org/content/canres/68/2/388.full.pdf

alpha 2-6-Linked sialic acids on N-glycans modulate carcinoma differentiation in vivo.

Protein N-glycosylation is a metabolic process that has been highly conserved in evolution. In all eukaryotes, N-glycosylation is obligatory for viability. It functions by modifying appropriate asparagine residues of proteins with oligosaccharide structures, thus influencing their properties and bioactivities. N-glycoprotein biosynthesis involves a multitude of enzymes, glycosyltransferases, and glycosidases, encoded by distinct genes. The majority of these enzymes are transmembrane proteins that function in the endoplasmic reticulum and Golgi apparatus in an ordered and well-orchestrated manner. The complexity of N-glycosylation is augmented by the fact that different asparagine residues within the same polypeptide may be modified with different oligosaccharide structures, and various proteins are distinguished from one another by the characteristics of their carbohydrate moieties. Furthermore, biological consequences of derivatization of proteins with N-glycans range from subtle to significant. In the past, all these features of N-glycosylation have posed a formidable challenge to an elucidation of the physiological role for this modification. Recent advances in molecular genetics, combined with the availability of diverse in vivo experimental systems ranging from yeast to transgenic mice, have expedited the identification, isolation, and characterization of N-glycosylation genes. As a result, rather unexpected information regarding relationships between N-glycosylation and other cellular functions--including secretion, cytoskeletal organization, proliferation, and apoptosis--has emerged. Concurrently, increased understanding of molecular details of N-glycosylation has facilitated the alignment between N-glycosylation deficiencies and human diseases, and has highlighted the possibility of using N-glycan expression on cells as potential determinants of disease and its progression. Recent studies suggest correlations between N-glycosylation capacities of cells and drug sensitivities, as well as susceptibility to infection. Therefore, knowledge of the regulatory features of N-glycosylation may prove useful in the design of novel therapeutics. While facing the demanding task of defining properties, functions, and regulation of the numerous, as yet uncharacterized, N-glycosylation genes, glycobiologists of the 21st century offer exciting possibilities for new approaches to disease diagnosis, prevention, and cure.

Protein N-Glycosylation: Molecular Genetics and Functional Significance

The rat beta-galactoside alpha 2,6-sialytransferase gene is differentially utilized by liver and kidney in the generation of mRNAs that predict substantially divergent polypeptides. In order to determine the biosynthetic relationship between these sialyltransferase mRNA isoforms, genomic sequences were isolated and analysed. Five exons that span at least 40 kb of DNA carry the coding information for the liver beta-galactoside alpha 2,6-sialyltransferase protein. An additional exon contains only sequences for the 5'-untranslated leader of the liver mRNA. In contrast, the predominant kidney mRNAs from this gene share only three coding exons that specify the carboxyl terminal 42% of the liver sialyltransferase protein sequence. In addition, these kidney mRNAs contain information from two other exons that comprise the 5' divergent region of these transcripts. Primer extension and S1 nuclease protection analysis demonstrate that the hepatic and kidney specific mRNAs are transcriptionally initiated at different sites within the sialyltransferase gene. While the hepatic sialyltransferase mRNAs are transcribed from the first exon, the kidney transcripts are initiated from a site within the third intron. Genomic regions upstream of both transcriptional initiation sites can regulate expression of the bacterial chloramphenicol acetyltransferase gene in transiently transfected L cells. Together, the data implicate multiple promoters as a principle mechanism in the generation of kidney and liver gene product diversity in sialyltransferase expression.

Rebeccah F. Young

Papers

Rat β-galactoside α2,6-sialyltransferase genomic organization: alternate promoters direct the synthesis of liver and kidney transcripts