RESUMEN
The exosome functions in the degradation of diverse RNA species, yet how it is negatively regulated remains largely unknown. Here, we show that NRDE2 forms a 1:1 complex with MTR4, a nuclear exosome cofactor critical for exosome recruitment, via a conserved MTR4-interacting domain (MID). Unexpectedly, NRDE2 mainly localizes in nuclear speckles, where it inhibits MTR4 recruitment and RNA degradation, and thereby ensures efficient mRNA nuclear export. Structural and biochemical data revealed that NRDE2 interacts with MTR4's key residues, locks MTR4 in a closed conformation, and inhibits MTR4 interaction with the exosome as well as proteins important for MTR4 recruitment, such as the cap-binding complex (CBC) and ZFC3H1. Functionally, MID deletion results in the loss of self-renewal of mouse embryonic stem cells. Together, our data pinpoint NRDE2 as a nuclear exosome negative regulator that ensures mRNA stability and nuclear export.
Asunto(s)
Exosomas/genética , Exosomas/metabolismo , Proteínas Nucleares/fisiología , ARN Helicasas/metabolismo , Animales , Núcleo Celular/metabolismo , Células Madre Embrionarias , Células HEK293 , Células HeLa , Humanos , Ratones , Proteínas Nucleares/genética , Unión Proteica , Dominios Proteicos , Transporte de Proteínas/genética , Estabilidad del ARN/genéticaRESUMEN
A significant fraction of mRNAs are degraded by the nuclear exosome in normal cells. Here, we studied where and when these exosome target mRNAs are sorted away from properly exported ones in the cells. We show that upon exosome inactivation, polyA RNAs are apparently accumulated in nuclear foci that are distinct from nuclear speckles (NSs), and provide several lines of evidence supporting that these polyA RNAs mainly correspond to accumulating exosome target mRNAs. These results suggest that exosomal mRNA degradation mostly occurs outside of NSs. In support of this possibility, targeting exosome target mRNAs to NSs stabilizes them by preventing exosomal degradation. Furthermore, inhibiting mRNA release from NSs does not attenuate exosomal degradation in normal cells, and results in polyA RNA accumulation both inside and outside of NSs in exosome inactivated cells, suggesting that passage through NSs is not required for sorting mRNAs for degradation or export. Indeed, exosome target mRNAs that normally do not enter NSs are exported upon exosome inactivation. Together, our data suggest that exosome target mRNAs are mainly degraded in the nucleoplasm before entering NSs and rapid removal of these mRNAs is important for preventing their nuclear export.
Asunto(s)
Transporte Activo de Núcleo Celular/fisiología , Núcleo Celular/metabolismo , Exosomas/metabolismo , Estabilidad del ARN/fisiología , Transporte de ARN/fisiología , ARN Mensajero/metabolismo , Núcleo Celular/ultraestructura , ARN Helicasas DEAD-box/antagonistas & inhibidores , ARN Helicasas DEAD-box/fisiología , Complejo Multienzimático de Ribonucleasas del Exosoma/fisiología , Células HeLa , Humanos , Poli A/genética , ARN Helicasas/fisiología , Proteínas de Unión al ARN/fisiologíaRESUMEN
The exosome is a key RNA machine that functions in the degradation of unwanted RNAs. Here, we found that significant fractions of precursors and mature forms of mRNAs and long noncoding RNAs are degraded by the nuclear exosome in normal human cells. Exosome-mediated degradation of these RNAs requires its cofactor hMTR4. Significantly, hMTR4 plays a key role in specifically recruiting the exosome to its targets. Furthermore, we provide several lines of evidence indicating that hMTR4 executes this role by directly competing with the mRNA export adaptor ALYREF for associating with ARS2, a component of the cap-binding complex (CBC), and this competition is critical for determining whether an RNA is degraded or exported to the cytoplasm. Together, our results indicate that the competition between hMTR4 and ALYREF determines exosome recruitment and functions in creating balanced nuclear RNA pools for degradation and export.
Asunto(s)
Proteínas Nucleares/metabolismo , ARN Helicasas/metabolismo , Estabilidad del ARN , Transporte de ARN/genética , ARN Nuclear/metabolismo , Proteínas de Unión al ARN/metabolismo , Factores de Transcripción/metabolismo , Transporte Activo de Núcleo Celular/genética , Complejo Multienzimático de Ribonucleasas del Exosoma/genética , Complejo Multienzimático de Ribonucleasas del Exosoma/metabolismo , Exosomas/genética , Exosomas/metabolismo , Técnicas de Silenciamiento del Gen , Células HEK293 , Células HeLa , Humanos , Proteínas Nucleares/genética , Unión Proteica , ARN Helicasas/genética , Estabilidad del ARN/genética , ARN Largo no Codificante/metabolismo , ARN Mensajero/metabolismo , Proteínas de Unión al ARN/genética , Factores de Transcripción/genéticaRESUMEN
Low density lipoprotein receptor-related protein 1 (LRP1) C766T polymorphism (rs1799986) has been extensively investigated for Alzheimer's disease (AD) susceptibility. However, results in different studies have been contradictory. Therefore, we conducted a meta-analysis containing 6455 AD cases and 6304 controls from 26 independent case-control studies to determine whether there was an association between the LRP1 C766T polymorphism and AD susceptibility. The combined analysis showed that there was no significant association between LRP1 C766T polymorphism and AD susceptibility (TT + CT versus CC: OR = 0.920, 95% CI = 0.817-1.037, P = 0.172). In subgroup analysis, significant decreased AD susceptibility was found among Asian population in allele model (T versus C: OR = 0.786, 95% CI = 0.635-0.974, P = 0.028) and dominant model (TT + CT versus CC: OR = 0.800, 95% CI = 0.647-0.990, P = 0.040). Moreover, T allele of LRP1 C766T was statistically associated with late onset of AD (LOAD) (T versus C: OR = 0.858, 95% CI = 0.748-0.985, P = 0.029; TT + CT versus CC: OR = 0.871, 95% CI = 0.763-0.994, P = 0.040). In conclusion, our meta-analysis suggested that LRP1 C766T polymorphism was associated with lower risk of AD in Asian, and could reduce LOAD risk especially. Considering some limitations of our meta-analysis, further large-scale studies should be done to reach a more comprehensive understanding.
Asunto(s)
Enfermedad de Alzheimer/genética , Predisposición Genética a la Enfermedad/genética , Proteína 1 Relacionada con Receptor de Lipoproteína de Baja Densidad/genética , Polimorfismo de Nucleótido Simple , Alelos , Enfermedad de Alzheimer/etnología , Enfermedad de Alzheimer/patología , Pueblo Asiatico/genética , Estudios de Casos y Controles , Frecuencia de los Genes , Predisposición Genética a la Enfermedad/etnología , Genotipo , Humanos , Factores de RiesgoRESUMEN
Bladder cancer is a challenging and fatal malignancy and the improvement in prognosis is limited over years. Deep understanding the mechanism of bladder cancer tumorigenesis and progression will help to discover novel and effective treatment strategies. In this study, we identify non-canonical IkB kinase TBK1 is up-regulated in bladder cancer tissue and cell lines. Knockdown of TBK1 markedly inhibits cell proliferation and migration. Inhibition of TBK1 kinase activity by BX795 significantly attenuates bladder cancer cell proliferation and migration. Mechanistic study shows that overexpression of TBK1 promoted the phosphorylation of Akt, whereas knockdown of TBK1 reverses this action. Taken together, our data suggest that TBK1 modulates the malignant behaviors of bladder cancer cell via Akt signaling, revealing new insights in discovering new therapy target for bladder cancer.