Alterations of O-glycan biosynthesis in human colon cancer tissues

Abstract

Human colon cancer is associated with antigenic and structural changes in mucin-type carbohydrate chains (O-glycans). To elucidate the control of the biosynthesis of these O-glycans is colon cancer, we have studied glycosyltransferase and sulphotransferase activities involved in the assembly of elongated O-glycan structures. We analysed homogenates prepared from cancer tissue, adjacent normal and distal normal tissue from 20 patients. Several transferase activities showed pronounced changes in cancer tissue. The changes correlate with previous findings of a loss of O-glycans in cancer mucins, but did not always correlate with levels of Tn, sialyl-Tn, T and Lex antigens in homogenates or with the differentiation status and Duke's stages of the cancer tissue or the patient's blood type, sex and age. UDP-GlcNAc: Gal NAc-R beta 3-N-acetylglucosaminyltransferase (where GlcNAc is N-acetyl-D-glucosamine and GalNAc is N-acetyl-D-galactosamine) synthesizing O-glycan core 3, GlcNAc beta 1-3GalNAc-, CMP-sialic acid: GalNAc-peptide alpha 6-sialyltransferase synthesizing the sialyl-Tn antigen and sulphotransferase activities towards O-glycan core 1, Gal beta 1-3GalNAc-, were found to be decreased in cancer. UDP-GlcNAc: Gal beta 1-3GalNAc beta 6-N-acetylglucosaminyltransferase was also decreased in cancer concomitant with a loss of the ability to synthesize the I antigen and core 4, GlcNAc beta 1-6(GlcNAc beta 1-3) GalNAc-, CMP-sialic acid: Gal beta 1-3GalNAc-R alpha 3-sialyltransferase and GDP-fucose: Gal beta-R alpha 2-fucosyltransferase, synthesizing the blood group H determinant, were found to be 4- and 3- to 8-fold increased, respectively, in cancer compared to normal tissue. The data suggest that the biosynthesis of antigens and mucin-bound O-glycan structures in colon cancer is subject to complex control mechanisms.

Control of O-Glycan Branch Formation

“…The entire coding sequence of the C2/4GnT gene was represented in both clones and sequenced in full using automated sequencing (ABI377, Perkin-Elmer). Confirmatory sequencing was performed on a cDNA zen et al 4 The sequence of the 5Ј end of C2/4GnT mRNA including the translational start site and 5Ј-UTR was obtained by 5Ј rapid amplification of cDNA ends (35 cycles at 94°C for 20 s, 52°C for 15 s, and 72°C for 2 min) using total cDNA from the human COLO 205 cancer cell line with the antisense primer TSHC 48 (5Ј-GTGG-GAACTGTATGAACTTCC-3Ј) (see Fig. 2).…”

“…The membrane was probed overnight at 42°C as described previously (28) and washed twice for 30 min each at 42°C with 2 ϫ SSC, 0.1% SDS and twice for 30 min each at 52°C with 0.1 ϫ SSC, 0.1% SDS. Human multiple tissue Northern blots, MTN I and MTN II (CLONTECH), were 4 The website at the Swiss Institute for Experimental Cancer Research is http://www.isrec.isb-sib.ch/software/TMPRED_form.html). probed as described above and washed twice for 10 min each at room temperature with 2 ϫ SSC, 0.1% SDS; twice for 10 min each at 55°C with 1 ϫ SSC, 0.1% SDS; and once for 10 min with 0.1 ϫ SSC, 0.1% SDS at 55°C.…”

“…In addition, activity was detected with GlcNAc␤1-3Gal␤1-R for I antigen biosynthesis. This ␤6GlcNAc-transferase activity has been designated the M-form for the mucin-secreting tissue form of C2GnT in contrast to the recently cloned leukocyte form or L-form, which is restricted to the formation of core 2 by accepting Gal␤1-3GalNAc␣-R substrates exclusively (4,20,21). The C2/4GnT enzyme reported here shows broader acceptor substrate specificity in accepting core 1 as well as core 3 but does not resemble the mucin-secreting tissue form because it lacks IGnT activity.…”

“…Synthesis of the different O-glycan structures is cell-and tissuespecific. Different core structures are produced upon differentiation and malignant transformation (3)(4)(5)(6)(7). For example, increased formation of GlcNAc␤1-6GalNAc branching in O-glycans has been demonstrated during T-cell activation, during the development of leukemia, and for immunodeficiencies like Wiskott-Aldrich syndrome and AIDS (3,8,9).…”

A multipotential β-1,6-N-acetylglucosaminyl-transferase is encoded by bovine herpesvirus type 4

“…Changes in C2GnT expression have also been observed during oncogenesis. An increase in expression of core 2 branched oligosaccharides by cancer cells has been reported in leukemia, colonic carcinoma, and cells transfected with T24Hras (17,(19)(20)(21)(22).…”