RESUMEN
2',3'-Cyclic nucleotide 3'-phosphodiesterase (CNP) is one of the earliest myelin-related proteins to be specifically expressed in differentiating oligodendrocytes (ODCs) in the central nervous system (CNS) and is implicated in myelin biogenesis. CNP possesses an in vitro enzymatic activity, whose in vivo relevance remains to be defined, because substrates with 2',3,-cyclic termini have not yet been identified. To characterize CNP function better, we previously determined the structure of the CNP catalytic domain by NMR. Interestingly, the structure is remarkably similar to the plant cyclic nucleotide phosphodiesterase (CPDase) from A. thaliana and the bacterial 2'-5' RNA ligase from T. thermophilus, which are known to play roles in RNA metabolism. Here we show that CNP is an RNA-binding protein. Furthermore, by using precipitation analyses, we demonstrate that CNP associates with poly(A)(+) mRNAs in vivo and suppresses translation in vitro in a dose-dependent manner. With SELEX, we isolated RNA aptamers that can suppress the inhibitory effect of CNP on translation. We also demonstrate that CNP1 can bridge an association between tubulin and RNA. These results suggest that CNP1 may regulate expression of mRNAs in ODCs of the CNS.
Asunto(s)
Hidrolasas Diéster Fosfóricas/metabolismo , Biosíntesis de Proteínas , Inhibidores de la Síntesis de la Proteína/metabolismo , ARN Mensajero/metabolismo , Proteínas de Unión al ARN/metabolismo , 2',3'-Nucleótido Cíclico 3'-Fosfodiesterasa , Animales , Autorradiografía , Western Blotting , Células COS , Chlorocebus aethiops , Electroforesis en Gel de Poliacrilamida , Ratones , Oligodendroglía/metabolismo , Hidrolasas Diéster Fosfóricas/clasificación , Hidrolasas Diéster Fosfóricas/genética , Inhibidores de la Síntesis de la Proteína/clasificación , ARN Mensajero/genética , Proteínas de Unión al ARN/clasificación , Conejos , Ratas , Técnica SELEX de Producción de Aptámeros , Tubulina (Proteína)/metabolismoRESUMEN
The oligodendrocyte-myelin glycoprotein is a ligand of the neuronal Nogo receptor and a potent inhibitor of neurite outgrowth, but its physiological function remains to be elucidated. The oligodendrocyte-myelin glycoprotein is anchored solely in the outer leaflet of the plasma membrane via its glycosylphosphatidylinositol anchor, and through its leucine-rich repeat domain, it likely interacts with other proteins. In the present study, we compare its buoyancy and detergent solubility characteristics with those of other myelin proteins. Based on its detergent solubility profile and membrane fractionation using established ultracentrifugation procedures, we conclude that the oligodendrocyte-myelin glycoprotein is a lipid raft component that is closely associated with the axolemma. Moreover, it associates with caveolin-1 and caveolin-1-enriched membranes. We postulate that, by virtue of its concentration in lipid rafts and perhaps through interactions with caveolin-1, the oligodendrocyte-myelin glycoprotein may influence signaling pathways.
Asunto(s)
Encéfalo/metabolismo , Caveolina 1/metabolismo , Microdominios de Membrana/metabolismo , Vaina de Mielina/metabolismo , Glicoproteína Asociada a Mielina/metabolismo , Animales , Axones/química , Axones/metabolismo , Detergentes/química , Proteínas Ligadas a GPI , Microdominios de Membrana/química , Ratones , Proteínas de la Mielina , Vaina de Mielina/química , Glicoproteína Asociada a Mielina/química , Glicoproteína Mielina-Oligodendrócito , Ratas , Transducción de Señal/fisiología , SolubilidadRESUMEN
A protein fraction purified from bovine brain myelin, previously called arretin because of its ability to inhibit neurite outgrowth, has been identified as consisting predominantly of oligodendrocyte-myelin glycoprotein (OMgp). We show that it is a potent inhibitor of neurite outgrowth from rat cerebellar granule and hippocampal cells; from dorsal root ganglion explants in which growth cone collapse was observed; from rat retinal ganglion neurons; and from NG108 and PC12 cells. OMgp purified by a different procedure from both mouse and human myelin behaves identically in all bioassays tested.
