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Abstract

Glycosylation is key posttranslational modification for membrane-bound and secreted proteins that can influence both the secondary structure and the function of the protein backbone. In order to investigate the effect of altered cellular glycosylation potential, we have generated a number of clonal cell lines over-expressing the alpha2,3(N) sialyltransferase enzyme (ST3N). In general, there was a decrease in total sialyltransferase (ST) enzyme activity in the clones transfected with the ST3N cDNA, with this decrease being inversely proportional to the quantity of the mRNA coding for the enzyme. The ST3N enzyme was, however, functional and there was an increase in both MAA lectin staining and the expression of polysialic acid, which is attached to the NCAM protein backbone primarily via an alpha2,3 linkage. These results suggest that the overexpression of a sialyltransferase may upset the sialylation potential of the cell.

Functional Consequences of Differential O-glycosylation of MUC1, MUC4, and MUC16 (Downstream Effects on Signaling)

“…Furthermore, a significant percentage of cancers exhibit hypermethylation of the Cosmc gene, resulting in decreased expression and increased formation of Tn and STn epitopes [ 52 , 53 ]. Another potential factor is deregulation of enzymes that extend or terminate extension of O -glycans (e.g., sialyl transferases), which has also been observed in a variety of cancers [ 54 , 55 , 56 , 57 , 58 , 59 ]. This may explain observed decreases in expression of Core 3 and 4 structures in gastric and colorectal cancers [ 60 , 61 ].…”

The role of protein glycosylation in the control of cellular N‐sialyltransferase activity

“…One primary problem in the field of glycobiology is the lack of sensitive tools with which to investigate the effects of altered glycosylation on protein structure, function and processing. There are inherent problems with over‐expressing (or knocking out) glycosyltransferase enzymes in cell lines by transfection as this has been reported to upset the subcellular distribution or functional roles of other Golgi enzymes [19]. While the generation of proteins with mutated glycosylation sites (usually Asn residues) or treatment with agents such as tunicamycin both act to completely block the glycosylation process, they are not sufficiently sensitive to provide any information on the role of individual sugar residues.…”

“…One primary problem in the ¢eld of glycobiology is the lack of sensitive tools with which to investigate the e¡ects of altered glycosylation on protein structure, function and processing. There are inherent problems with over-expressing (or knocking out) glycosyltransferase enzymes in cell lines by transfection as this has been reported to upset the subcellular distribution or functional roles of other Golgi enzymes [19].…”

“…Determining the role of individual sugar residues of oligosaccharide chains in protein function and processing can be particularly difficult as there are relatively few sufficiently selective tools with which to modify the activities of individual glycosyltransferase or glycosidase enzymes. While the generation of stably transfected cell lines is one approach that can be taken [18], the over‐expression of certain glycosyltransferase enzymes may actually disrupt the cellular glycosylation machinery [19]. However, a number of mutant Chinese hamster ovary (CHO) cell lines have previously been generated, each of which exhibits a specific glycosylation defect [20].…”