Research deals with the changes of
membrane glycosylation in disease and on their functional
significance. In particular:
1) the molecular mechanisms at the
basis of the altered expression of some glycosyltransferases in
cancer and its impact on cancer cell phenotype are studied. The
experimental model for these studies is comprised of colon and
bladder cancer cell lines genetically modified by
glycosyltransferase genes.
2) The study of glycosylation
changes in apoptosis has revealed that apoptotic cells acquire
selectively vitronectin from the extracellular microenvironment,
contributing to alter the glycophenotype.
3) The age-related changes of the
mechanisms of glycosylation of plasma membrane glycoproteins are
studied.
Glycosylation is one of the most important and frequent
post-translational modifications of proteins. The sugar structures
present on the membrane glycoproteins are the product of the
coordinated action of dozens of different glycosyltransferases. The
process of glycosylation is often altered in pathological
conditions, including cancer, and leads to the expression of
abnormal sugar structures on the membranes and on the secretions of
the tumor cells. These tumor-associated structures are important
for at least two reasons: first, because they can be used as
markers of neoplastic progression (the best known example is
provided by the antigen CA19.9, also known as sialyl Lewis a);
secondly their expression can change the phenotype of tumor cells.
Regarding the glycosylation in cancer, our research develops in the
following directions:
1) Study of the molecular mechanisms that lead to the biosynthesis
of tumor-associated sugar structures;
2) Study of the effects of expression of tumor-associated sugar
antigens on the phenotype of the tumor cells;
1) 2,6-sialyltransferase (ST6GAL1) is the enzyme that mounts the
sialic acid with 2,6-bond on the N-linked chains of glycoproteins.
This enzyme is often overexpressed in cancer cells because its
transcription is under the control of the RAS signaling pathway. In
previous years we have described the transcriptional mechanisms
leading to overexpression of this enzyme and of the sugar chains it
synthesized in colonic and liver tumors. The Sda carbohydrate
antigen and its cognate N-b1,4 acetylgalactosaminyltrasferase
(B4GALNT2) are expressed in the normal mucosa of the colon but are
greatly reduced in colon cancer, while the antigen sialyl Lewis x
(a ligand for the cell adhesion molecules of the selectin family
which plays a fundamental role in the process of metastasis) is
more espressed in tumor tissues. We have shown that this
modification, associated with malignancy, depends on the expression
of fucosyltrasferase 6 and the down-regulation of B4GALNT2.
2 The effects of the expression of tumor-associated sugar antigens
on the phenotype of the tumor cells can be studied by forcing the
expression of specific glycosyltransferases in cell lines using
gene transfer techniques. This can be done by transfection of the
gene of interest inserted into an expression vector plasmid or,
more efficiently, by retroviral transduction. Using one or the
other procedure we have previously obtained colon cancer cell lines
overexpressing ST6GAL1 or B4GALNT2 and lines of bladder cancer
overexpressing the sialyltransferases ST3GAL1 or ST6GALNAC1. We
reported that overexpression of ST6GAL1 led to a greater adhesion
of cells to extracellular substrates, such as fibronectin and
collagen IV and alters the morphology of the cell. In bladder
cancer, the phenotype was altered in cultured cells expressing
ST3GAL1 or ST6GALNAC1, while in samples of bladder cancer the
expression of the antigens synthesized by the two enzymes was
accompanied by the expression of specific clinical parameters and
response to therapy.
Regarding the alterations of glycosylation in apoptosis, we
observed that the apoptotic cells, but also those in primary
necrosis, became reactive with the lectin from Sambucus nigra that
binds to 2,6-sialylated structures. We have established that this
was not due to an alteration of the mechanisms of cellular
glycosylation associated with apoptosis, but rather to the
selective acquisition of vitronectin (a protein-rich of
2,6-sialylated structures) from the extracellular microenvironment.
The biological significance of this modification is under
investigation.
In aging, the glycosylation machinery of different organs and
tissues, particularly the liver and lymphocytes, undergoes several
modifications that lead to variations in the glycophenotype of
plasma proteins. The study that we have undertaken is aimed to
study the expression levels of plasma glycosyltransferases in
subjects of different ages and correlate it with inflammatory and
functional parameters and with the glycosylation of plasma
proteins.
