Ethanol consumption decreases the synthesis of the mannose 6-phosphate/insulin-like growth factor II receptor but does not decrease its messenger RNA.
The mannose 6-phosphate/insulin-like growth factor II receptor (M6P/IGF-IIR) is a protein that facilitates the transport of acid hydrolases into the lysosome. We have shown that chronic ethanol consumption lowers the M6P/IGF-IIR content in rat hepatocytes. Here, we determined the steady-state level of mRNA encoding M6P/IGF-IIR, as well as the rate of receptor synthesis, to ascertain whether the ethanol-elicited reduction in receptor protein content is related to changes in either or both of these parameters. Rats were pair-fed the normal carbohydrate (NC) or low carbohydrate high-fat (LC) liquid diets containing either ethanol or isocaloric maltose-dextrin for 7-8 weeks. RNA was isolated from hepatocytes and from whole livers of these animals and subjected to reverse transcription-polymerase chain reaction (RT-PCR) to determine the mRNA levels encoding M6P/IGF-IIR. Hepatocytes isolated from these animals were also radiolabeled with Pro-mix L-[35S] in vitro cell labeling mix to measure incorporation into total cellular protein and the immunoprecipitated M6P/IGF-IIR protein. The steady-state levels of M6P/IGF-IIR mRNA in both hepatocytes and whole livers from ethanol-fed rats were the same as those from their respective controls regardless of whether they were fed the NC or the LC diets. Hepatocytes from ethanol-fed rats showed a 36% lower rate of total protein synthesis and an even greater reduction (70%) in receptor synthesis. When the relative rate of receptor synthesis was calculated, hepatocytes from ethanol-fed rats had a 53% lower relative rate of receptor synthesis compared with controls. Autoradiographic analysis of the immunoprecipitated receptor protein from ethanol-fed rats also indicated a 79% decline in the total M6P/IGF-IIR protein synthetic rate compared with pair-fed controls. We conclude that the ethanol-elicited reduction of M6P/IGF-IIR content was, in part, related to a concomitant reduction of receptor protein synthesis but not to a decline in its mRNA level. Thus, the ethanol-elicited decline in receptor protein synthesis may be due to defective M6P/IGF-IIR mRNA translation.
Haorah
James
J
Liver Study Unit, Research Service (151), The Veterans Affairs (VA) Medical Center, 4101 Woolworth Avenue, Omaha, NE 68105, USA. jhaorah@unmc.edu
MacDonald
Richard G
RG
Stoner
Julie A
JA
Donohue
Terrence M
TM
D000428:Alcohol Drinking; D000704:Analysis of Variance; D000818:Animals; D000431:Ethanol / Q000494:pharmacology*; D008297:Male; D008360:Mannosephosphates / Q000378:metabolism*; D011233:Precipitin Tests; D012333:RNA, Messenger / Q000187:drug effects / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D017527:Receptor, IGF Type 2 / Q000235:genetics / Q000378:metabolism*
D008360:Mannosephosphates; D012333:RNA, Messenger; D017527:Receptor, IGF Type 2; C027693:mannose-6-phosphate; D000431:Ethanol