Hydroxylation of CMP-NeuAc controls the expression of N-glycolylneuraminic acid in GM3 ganglioside of the small intestine of inbred rats.
An enzymatic activity responsible for the hydroxylation of CMP-NeuAc into CMP-N-glycolylneuraminic acid (CMP-NeuGc) was found in the cytosolic fraction after cellular fractionation of the mucosa of rat small intestine. It was maximum in the presence of NADPH or NADH, but it was reduced by 50% by addition of 1 mM EDTA. The Km value for CMP-NeuAc was 0.6 microM. The CMP-NeuAc hydroxylase activity paralleled the expression of the GM3 (NeuGc) phenotype in the epithelium of the small intestine and was not measurable in the mutant rats BN and SHR that only expressed GM3 (NeuAc). Furthermore, the only form of CMP-sialic acid present in the intestinal mucosa of the mutants was CMP-NeuAc, whereas in the other strains CMP-NeuGc accounted for 70-85% of the native CMP-sialic acids. Wild-type and CMP-NeuAc hydroxylase-deficient inbred rats were mated. Individuals of F1 and backcross generations were typed for the phenotypes GM3(NeuGc)/GM3(NeuAc) and the activity of CMP-NeuAc hydroxylase in the small intestine. It was found that the expression of NeuGc in GM3 depends on a single autosomal dominant gene and correlates with the activity of CMP-NeuAc hydroxylase. Two tissues other than small intestine, kidney and spleen, which expressed GM3(NeuGc) in BN and SHR, also expressed the CMP-NeuAc hydroxylase activity, as in the other strains. It was concluded that the key enzyme responsible for the presence of NeuGc in GM3 is a CMP-NeuAc hydroxylase and that mutant rats carry a defect that is specific to intestine. The comparative analysis of the respective contribution of NeuGc and NeuAc to the glycoprotein sialic acids of the small intestine showed that CMP-NeuAc hydroxylase is also responsible for part of the NeuGc present in the glycoproteins. However, the occurrence of 20-30% of NeuGc in the intestinal glycoproteins of the CMP-NeuAc hydroxylase-deficient rats indicated that there is another enzyme providing intestinal glycoproteins with NeuGc and operating under a different genetic control.
Bouhours
J F
JF
Institut National de la Santé et de la Recherche Médicale, Unité 76, Paris, France.
Bouhours
D
D
D000818:Animals; D003569:Cytidine Monophosphate N-Acetylneuraminic Acid / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D005260:Female; D005679:G(M3) Ganglioside / Q000235:genetics*; D005732:Gangliosides; D015870:Gene Expression; D006900:Hydroxylation; D007413:Intestinal Mucosa / Q000378:metabolism; D007421:Intestine, Small / Q000254:growth & development / Q000378:metabolism*; D008297:Male; D006899:Mixed Function Oxygenases / Q000235:genetics* / Q000378:metabolism; D009438:Neuraminic Acids / Q000378:metabolism*; D051381:Rats; D011914:Rats, Inbred BN; D011917:Rats, Inbred Lew; D011918:Rats, Inbred SHR; D011919:Rats, Inbred Strains; D012794:Sialic Acids / Q000378:metabolism*; D014018:Tissue Distribution
D005679:G(M3) Ganglioside; D005732:Gangliosides; D009438:Neuraminic Acids; D012794:Sialic Acids; C032592:N-glycolylneuraminic acid; D003569:Cytidine Monophosphate N-Acetylneuraminic Acid; D006899:Mixed Function Oxygenases; C074383:CMPacetylneuraminate monooxygenase