Mucins in
ulcerative colitis and
colon cancer share common properties of reduced sulfation and increased oncofetal
carbohydrate antigen expression. It has previously been shown that there is no simple correlation between these changes and the activity of the relevant glycosyl-, sialyl-, and sulfo-
transferases. We examined
mucin sulfation and expression of oncofetal Thomsen-Friedenreich (
TF) antigen (galactosyl beta1-3N-acetylgalactosamine alpha-) in the goblet cell-differentiated human
colon cancer cell line LS174T following treatment with
bafilomycin A(1, )which raises intra-Golgi pH, or
monensin, which disrupts medial-trans Golgi transport. Cells were dual-labeled with
sodium [(35)S]-
sulfate and D-[6-(3)H(N)]-
glucosamine hydrochloride, or labeled with L-[U-(14)C]-
threonine alone.
Mucin was purified using
Sepharose CL-4B gel filtration.
Mucin sulfo-Lewis(a) and
TF antigen expression were assessed using the F2 anti-
sulfo-Lewis(a)
monoclonal antibody and
peanut agglutinin binding respectively. Bafilomycin (0.01 microM; 48 h) reduced total
mucin sulfation, expressed relative to incorporation of
glucosamine, to 0.50 +/- 0.04 d.p.m. [(35)S]-
sulfate per d.p.m. [(3)H]-
glucosamine compared to control, 0.84 +/- 0.05 (p < 0.001, n = 16). This was accompanied by 50.3 +/- 8.0% increased expression of
TF antigen (p < 0.01) and 50.1 +/- 5.5% decreased expression of
sulfo-Lewis(a) (p < 0.01). The reduced
sulfate:glucosamine ratio was largely due to increased incorporation of
glucosamine into newly synthesized
mucin rather than reduction in total
sulfate incorporation. In contrast,
monensin only reduced total
mucin glycosylation at concentrations > 0.1 microM and had no significant effect on
mucin sulfation or TF expression. Intra-Golgi alkalinization affects
mucin glycosylation, resulting in decreased
mucin sulfation and increased expression of
TF antigen, changes that mimic those seen in cancerous and premalignant human colonic epithelium.