Effect of cycloheximide on palmitylation of PO protein of the peripheral nervous system myelin.
Incubation of rat sciatic nerve slices with Krebs-Ringer bicarbonate buffer containing [3H]palmitic acid resulted in the acylation of the PO glycoprotein and a 24 kDa protein of the peripheral nerve myelin. Radioactivity was removed from PO after treating PO with hydroxylamine (83%) and methanolic KOH (97%). These results provided evidence that the radioactivity incorporated into PO was not due to the metabolic conversion of [3H]palmitic acid into amino acids or sugars. PO was more heavily labelled in the homogenate than in the myelin membrane in 8-day-old rat nerve between 5 min and 2 h of incubation. These results suggested that PO may be primarily acylated in the cell body. Incubation of purified myelin with [1-14C]palmitoyl-CoA resulted in the non-enzymic acylation of PO. This provided evidence of the absence of fatty acyltransferase from the purified peripheral nerve myelin. Glycosylation of PO has been shown to occur in the Golgi complex, and monensin inhibited glycosylation of PO in the homogenate and myelin by 53 and 61% respectively. These results suggest that the processing of PO in the Golgi complex and the assembly of PO into myelin is impaired by monensin. However, fatty acylation of PO was unimpaired by monensin, suggesting that the addition of fatty acids may not occur in the Golgi complex. There was a progressive decrease in the acylation of PO between 5 min (28%) and 2 h (61%) in the presence of cycloheximide, as the pool of previously synthesized PO was gradually depleted. These results also provide evidence that palmitylation of PO is not coupled to protein synthesis, and acylation of this protein probably occurs in the early subcompartment of the Golgi complex, which appears to be insensitive to monensin.
Agrawal
H C
HC
Department of Pediatrics, Washington University School of Medicine, St. Louis, MO 63110.
Agrawal
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D
D000215:Acylation; D000818:Animals; D002474:Cell-Free System; D003513:Cycloheximide / Q000494:pharmacology*; D006031:Glycosylation; D006878:Hydroxides / Q000494:pharmacology; D019811:Hydroxylamine; D006898:Hydroxylamines / Q000494:pharmacology; D066298:In Vitro Techniques; D008562:Membrane Glycoproteins / Q000096:biosynthesis / Q000378:metabolism*; D008985:Monensin / Q000494:pharmacology; D018993:Myelin P0 Protein; D009185:Myelin Proteins / Q000096:biosynthesis / Q000378:metabolism*; D009186:Myelin Sheath / Q000187:drug effects*; D011188:Potassium / Q000494:pharmacology; D017680:Potassium Compounds; D051381:Rats; D011919:Rats, Inbred Strains; D012584:Sciatic Nerve / Q000187:drug effects*; D014415:Tunicamycin / Q000494:pharmacology
D006878:Hydroxides; D006898:Hydroxylamines; D008562:Membrane Glycoproteins; D018993:Myelin P0 Protein; D009185:Myelin Proteins; D017680:Potassium Compounds; D014415:Tunicamycin; D019811:Hydroxylamine; D008985:Monensin; D003513:Cycloheximide; D011188:Potassium; C029943:potassium hydroxide