Identification of MARPP (14-20), morphine- and cyclic AMP-regulated phosphoproteins of 14-20 kDa, as myelin basic proteins: evidence for their acute and chronic regulation by morphine in rat brain.
MARPP(14-20), morphine- and cyclic AMP-regulated phosphoproteins of 14, 17-18, and 20 kDa, were identified originally by one-dimensional electrophoresis as a group of proteins whose state of phosphorylation was decreased by acute morphine and increased by chronic morphine in the rat locus coeruleus. We now show that MARPP(14-20) represent myelin basic proteins based on biochemical and immunochemical criteria. First, MARPP(14-20) were found to have isoelectric points of about 11 based on their migration on non-equilibrium pH gradient electrophoresis. Second, MARPP(14-20) were greatly enriched in myelin fractions of brain, and were not detectable, or present at very low levels, in other subcellular fractions of brain. Third, analysis of phosphorylated MARPP(14-20) by one- and two-dimensional gel electrophoresis demonstrated precise comigration with immunolabeled myelin basic proteins. In contrast, MARPP(14-20) were distinguished from histones, another group of low molecular weight, highly basic phosphoproteins, in these subcellular fractionation and immunochemical studies. Finally, we confirmed using two-dimensional electrophoresis, that changes observed previously by one-dimensional electrophoresis in the phosphorylation of MARPP(14-20) in response to acute and chronic morphine, and to acute forskolin, occur in myelin basic proteins. It was also found that changes in the state of phosphorylation of myelin basic proteins in response to chronic morphine occur without a change in the total amount of the proteins as determined by immunoblot analysis. The results demonstrate that the phosphorylation of myelin basic proteins is regulated by morphine in the nervous system, and raise the possibility that regulation of these proteins contributes to mechanisms underlying some of the acute and chronic actions of opiates in brain.
Guitart
X
X
Laboratory of Molecular Psychiatry, Yale University School of Medicine, New Haven, CT 06508.
Nestler
E J
EJ
D000818:Animals; D001921:Brain / Q000187:drug effects / Q000378:metabolism*; D000242:Cyclic AMP / Q000494:pharmacology*; D008297:Male; D008970:Molecular Weight; D009020:Morphine / Q000494:pharmacology*; D004676:Myelin Basic Protein / Q000302:isolation & purification*; D010750:Phosphoproteins / Q000302:isolation & purification* / Q000502:physiology; D010766:Phosphorylation; D051381:Rats; D011919:Rats, Inbred Strains; D013347:Subcellular Fractions / Q000032:analysis
D004676:Myelin Basic Protein; D010750:Phosphoproteins; D009020:Morphine; D000242:Cyclic AMP