The polyadenylated RNA directing the synthesis of the rat myelin basic proteins is present in both free and membrane-bound forebrain polyribosomes.
Free and membrane-bound polyribosomes were isolated from the forebrain of actively myelinating 24-day-old rats. The poly(A)+ RNA (polyadenylated RNA) extracted from both fractions was translated in vitro in reticulocyte lysates [Hall & Lim (1981) Biochem. J. 196. 327-336] in the presence or absence of a heterologous microsomal membrane fraction from dog pancreas. The rat myelin basic proteins synthesized in vitro were isolated by CM-cellulose chromatography and by immunoprecipitation with purified anti-(myelin basic protein) antibody. The large (mol.wt. 18 500) and small (mol.wt. 16 000) myelin basic proteins were translational products of poly(A)+ RNA from both free and membrane-bound polyribosomes. The identity of the myelin basic proteins was verified by analysis of peptides generated by the cathepsin D digestion of the immunoprecipitated proteins synthesized in vitro, in comparison with authentic rat myelin basic proteins. Although several other translational products of membrane-bound polyribosomal poly(A)+ RNA were modified when microsomal membranes were present during translation, molecular weights of the myelin basic proteins themselves were unchanged. The myelin basic proteins synthesized in vitro also did not differ significantly in size from the authentic myelin basic proteins, indicating that these membrane proteins are unlikely to be synthesized as substantially larger precursor molecules. The presence of the specific mRNA species on both free and membrane-bound polyribosomes is compatible with the extrinsic location of the myelin basic proteins on the cytoplasmic surface of the myelin membrane.
D000818:Animals; D002402:Cathepsin D; D002403:Cathepsins; D002462:Cell Membrane / Q000378:metabolism; D011232:Chemical Precipitation; D002852:Chromatography, Ion Exchange; D004027:Diencephalon / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D066298:In Vitro Techniques; D008297:Male; D004676:Myelin Basic Protein / Q000096:biosynthesis* / Q000276:immunology / Q000302:isolation & purification; D011061:Poly A / Q000378:metabolism*; D011132:Polyribosomes / Q000378:metabolism*; D014176:Protein Biosynthesis; D012313:RNA / Q000378:metabolism*; D012333:RNA, Messenger; D051381:Rats; D011919:Rats, Inbred Strains; D013687:Telencephalon / Q000378:metabolism*
D004676:Myelin Basic Protein; D012333:RNA, Messenger; D011061:Poly A; D012313:RNA; D002403:Cathepsins; D002402:Cathepsin D