RESUMO
Viruses have evolved strategies to ensure efficient translation using host cell ribosomes and translation factors. In addition to cleaving translation initiation factors required for host cell translation, poliovirus (PV) uses an internal ribosome entry site (IRES). Recent studies suggest that viruses exploit specific ribosomal proteins to enhance translation of their viral proteins. The ribosomal protein receptor for activated C kinase 1 (RACK1), a protein of the 40S ribosomal subunit, was previously shown to mediate translation from the 5' cricket paralysis virus and hepatitis C virus IRESs. Here we found that translation of a PV dual-luciferase reporter shows a moderate dependence on RACK1. However, in the context of a viral infection we observed significantly reduced poliovirus plaque size and titers and delayed host cell translational shut-off. Our findings further illustrate the involvement of the cellular translational machinery during PV infection and how viruses usurp the function of specific ribosomal proteins.
Assuntos
Hepacivirus/genética , Hepatite C/metabolismo , Sítios Internos de Entrada Ribossomal , Poliomielite/metabolismo , Poliovirus/genética , Receptores de Quinase C Ativada/metabolismo , Hepacivirus/metabolismo , Hepatite C/genética , Hepatite C/virologia , Interações Hospedeiro-Patógeno , Humanos , Poliomielite/genética , Poliomielite/virologia , Poliovirus/metabolismo , Biossíntese de Proteínas , Receptores de Quinase C Ativada/genética , Ribossomos/genética , Ribossomos/metabolismoRESUMO
Many metabotropic receptors in the nervous system act through signaling pathways that result in the inhibition of voltage-dependent calcium channels. Our previous findings showed that activation of seven-transmembrane receptors results in the internalization of calcium channels. This internalization takes place within a few seconds, raising the question of whether the endocytic machinery is in close proximity to the calcium channel to cause such rapid internalization. Here we show that voltage-dependent calcium channels are pre-associated with arrestin, a protein known to play a role in receptor trafficking. Upon GABAB receptor activation, receptors are recruited to the arrestin-channel complex and internalized. beta-Arrestin 1 selectively binds to the SNARE-binding region of the calcium channel. Peptides containing the arrestin-binding site of the channel disrupt agonist-induced channel internalization. Taken together these data suggest a novel neuronal role for arrestin.
Assuntos
Arrestina/fisiologia , Canais de Cálcio/metabolismo , Animais , Arrestina/metabolismo , Sítios de Ligação , Canais de Cálcio Tipo N/metabolismo , Embrião de Galinha , Endocitose , Neurônios/metabolismo , Peptídeos/química , Ligação Proteica , Transporte Proteico , Transdução de SinaisRESUMO
Calcium channels are well known targets for inhibition by G protein-coupled receptors, and multiple forms of inhibition have been described. Here we report a novel mechanism for G protein-mediated modulation of neuronal voltage-dependent calcium channels that involves the destabilization and subsequent removal of calcium channels from the plasma membrane. Imaging experiments in living sensory neurons show that, within seconds of receptor activation, calcium channels are cleared from the membrane and sequestered in clathrin-coated vesicles. Disruption of the L1-CAM-ankyrin B complex with the calcium channel mimics transmitter-induced trafficking of the channels, reduces calcium influx, and decreases exocytosis. Our results suggest that G protein-induced removal of plasma membrane calcium channels is a consequence of disrupting channel-cytoskeleton interactions and might represent a novel mechanism of presynaptic inhibition.