Long-term ethanol exposure impairs glycosylation of both N- and O-glycosylated proteins in rat liver☆

Abstract 

Carbohydrate residues of glycoproteins play important roles in their functions. We have previously shown that long-term ethanol treatment in rats alters the normal glycosylation pattern of plasma transferrin and apolipoprotein (apo) E. Glycosylation of proteins is a posttranslational process that is regulated by both glycosyltransferases and glycosidases, the resident enzymes of hepatic subcellular organelles. In this investigation using rat transferrin and apo E as model N- and O-glycosylated proteins, respectively, we have explored the effects of long-term ethanol treatment on the (1) incorporation of various labeled sugar precursors into these specific glycoproteins, (2) activities of mannosyltransferase, galactosyltransferase, and sialyltransferases, and (3) hepatic synthetic rate of N-acetyl glucosamine (GlcNAc) α2,6-sialyltransferase (2,6-ST). The relative ratio of labeled sugar to leucine incorporation (glycosylation index) showed a 43% (P < .01) decrease for relative mannosylation of transferrin molecule at both the microsomal and Golgi level in the ethanol group (AN) versus the control group (CN). For apo E, relative mannosylation was reduced by 48.9% (P < .01) and 46.9% (P < .01), respectively, at the microsomal and Golgi level in the AN versus CN. More importantly, relative sialation of transferrin was reduced by 86% (P < .001) in AN as compared with CN. Relative sialation of apo E was reduced by 35% (P < .01) in AN as compared with CN. A comparison of the effects of long-term ethanol treatment on key glycosylating enzymes in the liver showed that the activities of mannosyltransferase and galactosyltransferase were decreased by 24.4% (P < .01) and 21.1% (P < .01), respectively, whereas the activity of 2,6-ST was decreased markedly by 52.9% (P < .001) AN versus CN. This inhibitory effect of ethanol was specific for 2,6-ST, since the activities of GlcNAc α-2,3-sialyltransferase and [4,5-³H]-N-acetylgalactosamine (GalNAc) α-2,3-ST were decreased only by 31.6% (P < .01) and 4% (NS), respectively, by the same long-term ethanol treatment. It was further shown that this inhibition of 2,6-ST activity was due to a concomitant 48% (P < .001) specific inhibition of its synthetic rate caused by long-term ethanol treatment. Thus, our results have clearly established that long-term ethanol treatment leads to a marked specific inhibition of the sialation step of both N- and O-glycosylated proteins by inhibiting the hepatic synthetic rate of 2,6-ST.

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