RESUMEN
Notch signaling is a key regulator of angiogenesis, in which sprouting is regulated by an equilibrium between inhibitory Dll4-Notch signaling and promoting Jagged-Notch signaling. Whereas Fringe proteins modify Notch receptors and strengthen their activation by Dll4 ligands, other mechanisms balancing Jagged and Dll4 signaling are yet to be described. The intermediate filament protein vimentin, which has been previously shown to affect vascular integrity and regenerative signaling, is here shown to regulate ligand-specific Notch signaling. Vimentin interacts with Jagged, impedes basal recycling endocytosis of ligands, but is required for efficient receptor ligand transendocytosis and Notch activation upon receptor binding. Analyses of Notch signal activation by using chimeric ligands with swapped intracellular domains (ICDs), demonstrated that the Jagged ICD binds to vimentin and contributes to signaling strength. Vimentin also suppresses expression of Fringe proteins, whereas depletion of vimentin enhances Fringe levels to promote Dll4 signaling. In line with these data, the vasculature in vimentin knockout (VimKO) embryos and placental tissue is underdeveloped with reduced branching. Disrupted angiogenesis in aortic rings from VimKO mice and in endothelial 3D sprouting assays can be rescued by reactivating Notch signaling by recombinant Jagged ligands. Taken together, we reveal a function of vimentin and demonstrate that vimentin regulates Notch ligand signaling activities during angiogenesis.
Asunto(s)
Neovascularización Fisiológica , Receptores Notch/metabolismo , Vimentina/metabolismo , Animales , Aorta/metabolismo , Embrión de Pollo , Endocitosis , Células Endoteliales de la Vena Umbilical Humana , Humanos , Técnicas In Vitro , Proteína Jagged-1/genética , Proteína Jagged-1/metabolismo , Ligandos , Ratones , Ratones de la Cepa 129 , Ratones Noqueados , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Transducción de Señal , Activación Transcripcional , Vimentina/deficiencia , Vimentina/genéticaRESUMEN
The sphingolipids, sphingosine 1-phosphate (S1P) and sphingosylphosphorylcholine (SPC), can induce or inhibit cellular migration. The intermediate filament protein vimentin is an inducer of migration and a marker for epithelial-mesenchymal transition. Given that keratin intermediate filaments are regulated by SPC, with consequences for cell motility, we wanted to determine whether vimentin is also regulated by sphingolipid signalling and whether it is a determinant for sphingolipid-mediated functions. In cancer cells where S1P and SPC inhibited migration, we observed that S1P and SPC induced phosphorylation of vimentin on S71, leading to a corresponding reorganization of vimentin filaments. These effects were sphingolipid-signalling-dependent, because inhibition of either the S1P2 receptor (also known as S1PR2) or its downstream effector Rho-associated kinase (ROCK, for which there are two isoforms ROCK1 and ROCK2) nullified the sphingolipid-induced effects on vimentin organization and S71 phosphorylation. Furthermore, the anti-migratory effect of S1P and SPC could be prevented by expressing S71-phosphorylation-deficient vimentin. In addition, we demonstrated, by using wild-type and vimentin-knockout mouse embryonic fibroblasts, that the sphingolipid-mediated inhibition of migration is dependent on vimentin. These results imply that this newly discovered sphingolipid-vimentin signalling axis exerts brake-and-throttle functions in the regulation of cell migration.
Asunto(s)
Movimiento Celular/fisiología , Esfingolípidos/metabolismo , Vimentina/metabolismo , Animales , Línea Celular , Línea Celular Tumoral , Fibroblastos/metabolismo , Humanos , Lisofosfolípidos/metabolismo , Ratones , Fosforilación/fisiología , Fosforilcolina/análogos & derivados , Fosforilcolina/metabolismo , Receptores de Lisoesfingolípidos/metabolismo , Transducción de Señal/fisiología , Esfingosina/análogos & derivados , Esfingosina/metabolismo , Receptores de Esfingosina-1-Fosfato , Quinasas Asociadas a rho/metabolismoRESUMEN
Intermediate filament (IF) proteins comprise a large family with more than 70 members. Initially, IFs were assumed to provide only structural reinforcement for the cell. However, IFs are now known to be dynamic structures that are involved in a wide range of cellular processes during all stages of life, from development to ageing, and during homeostasis and stress. This Commentary discusses some lesser-known functional and regulatory aspects of IFs. We specifically address the emerging roles of nestin in myogenesis and cancer cell migration, and examine exciting evidence on the regulation of nestin and lamin A by the notch signalling pathway, which could have repercussions for our understanding of the roles of IF proteins in development and ageing. In addition, we discuss the modulation of the post-translational modifications of neuronally expressed IFs and their protein-protein interactions, as well as IF glycosylation, which not only has a role in stress and ageing, but might also regulate IFs during development. Although many of these recent findings are still preliminary, they nevertheless open new doors to explore the functionality of the IF family of proteins.
Asunto(s)
Envejecimiento/metabolismo , Filamentos Intermedios/metabolismo , Envejecimiento/genética , Animales , Humanos , Proteínas de Filamentos Intermediarios/genética , Proteínas de Filamentos Intermediarios/metabolismo , Filamentos Intermedios/genética , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Nestina , Procesamiento Proteico-Postraduccional/genética , Procesamiento Proteico-Postraduccional/fisiología , Transducción de Señal/genética , Transducción de Señal/fisiologíaRESUMEN
The AP-1 transcription factor c-Jun has been shown to be essential for stress-induced apoptosis in several models. However, the molecular mechanisms underlying the proapoptotic activity of c-Jun are poorly understood. We identify the apoptosis-antagonizing transcription factor (AATF) as a novel nucleolar stress sensor, which is required as a cofactor for c-Jun-mediated apoptosis. Overexpression or down-regulation of AATF expression levels led to a respective increase or decrease in the amount of activated and phosphorylated c-Jun with a proportional alteration in the induction levels of the proapoptotic c-Jun target genes FasL and TNF-α. Accordingly, AATF promoted commitment of ultraviolet (UV)-irradiated cells to c-Jun-dependent apoptosis. Whereas AATF overexpression potentiated UV-induced apoptosis in wild-type cells, c-Jun-deficient mouse embryonic fibroblasts were resistant to AATF-mediated apoptosis induction. Furthermore, AATF mutants defective in c-Jun binding were also defective in inducing AP-1 activity and c-Jun-mediated apoptosis. UV irradiation induced a translocation of AATF from the nucleolus to the nucleus, thereby enabling its physical association to c-Jun. Analysis of AATF deletion mutants revealed that the AATF domains required for compartmentalization, c-Jun binding, and enhancement of c-Jun transcriptional activity were all also required to induce c-Jun-dependent apoptosis. These results identify AATF as a nucleolar-confined c-Jun cofactor whose expression levels and spatial distribution determine the stress-induced activity of c-Jun and the levels of c-Jun-mediated apoptosis.