RESUMO
The positive-sense, single-stranded RNA alphaviruses pose a potential epidemic threat. Understanding the complex interactions between the viral and the host cell proteins is crucial for elucidating the mechanisms underlying successful virus replication strategies and for developing specific antiviral interventions. Here we present the first comprehensive protein-protein interaction map between the proteins of Semliki Forest Virus (SFV), a mosquito-borne member of the alphaviruses, and host cell proteins. Among the many identified cellular interactors of SFV proteins, the enrichment of factors involved in translation and nonsense-mediated mRNA decay (NMD) was striking, reflecting the virus' hijacking of the translation machinery and indicating viral countermeasures for escaping NMD by inhibiting NMD at later time points during the infectious cycle. In addition to observing a general inhibition of NMD about 4 hours post infection, we also demonstrate that transient expression of the SFV capsid protein is sufficient to inhibit NMD in cells, suggesting that the massive production of capsid protein during the SFV reproduction cycle is responsible for NMD inhibition.
Assuntos
Infecções por Alphavirus/virologia , Proteínas do Capsídeo/metabolismo , Interações Hospedeiro-Patógeno , Degradação do RNAm Mediada por Códon sem Sentido/genética , Vírus da Floresta de Semliki/fisiologia , Proteínas do Capsídeo/genética , Células HeLa , Humanos , Vírus da Floresta de Semliki/genética , Replicação ViralRESUMO
To gain insight into the mechanistic link between translation termination and nonsense-mediated mRNA decay (NMD), we depleted the ribosome recycling factor ABCE1 in human cells, resulting in an upregulation of NMD-sensitive mRNAs. Suppression of NMD on these mRNAs occurs prior to their SMG6-mediated endonucleolytic cleavage. ABCE1 depletion caused ribosome stalling at termination codons (TCs) and increased ribosome occupancy in 3' UTRs, implying enhanced TC readthrough. ABCE1 knockdown indeed increased the rate of readthrough and continuation of translation in different reading frames, providing a possible explanation for the observed NMD inhibition, since enhanced readthrough displaces NMD activating proteins from the 3' UTR. Our results indicate that stalling at TCs triggers ribosome collisions and activates ribosome quality control. Collectively, we show that improper translation termination can lead to readthrough of the TC, presumably due to ribosome collisions pushing the stalled ribosomes into the 3' UTR, where it can resume translation in-frame as well as out-of-frame.
Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Códon de Terminação/genética , Degradação do RNAm Mediada por Códon sem Sentido/genética , Telomerase/genética , Regiões 3' não Traduzidas/genética , Mudança da Fase de Leitura do Gene Ribossômico/genética , Humanos , Biossíntese de Proteínas/genética , RNA Mensageiro/genética , Ribossomos/genéticaRESUMO
Eukaryotic cells have evolved a number of survival tactics and quality control pathways that are able to counter intrinsic error-prone mechanisms and allow for maintenance of cellular homeostasis in the face of external stresses. This review will discuss the role of two cellular eukaryotic processes that are vital for maintenance of cellular homeostasis - 1) the nonsense-mediated mRNA decay (NMD) pathway and 2) the transient formation of stress granules (SG) - and explore the current literature on their roles in antiviral defence. Within the NCCR RNA & Disease, the laboratories of Proffs. O. Mühlemann and Volker Thiel teamed up to unravel the roles of NMD and SGs, and their interconnections in defending cells from alphavirus and coronavirus infections.
Assuntos
Degradação do RNAm Mediada por Códon sem Sentido , Antivirais , RNARESUMO
Nonsense-mediated mRNA decay (NMD) has traditionally been described as a quality control system that rids cells of aberrant mRNAs with crippled protein coding potential. However, transcriptome-wide profiling of NMD deficient cells identified a plethora of seemingly intact mRNAs coding for functional proteins as NMD targets. This led to the view that NMD constitutes an additional post-transcriptional layer of gene expression control involved in the regulation of many different biological pathways. Here, we review our current knowledge about the role of NMD in embryonic development and tissue-specific cell differentiation. We further summarize how NMD contributes to balancing of the integrated stress response and to cellular homeostasis of splicing regulators and NMD factors through auto-regulatory feedback loops. In addition, we discuss recent evidence that suggests a role for NMD as an innate immune response against several viruses. Altogether, NMD appears to play an important role in a broad spectrum of biological pathways, many of which still remain to be discovered.
Assuntos
Regulação da Expressão Gênica , Homeostase/genética , Degradação do RNAm Mediada por Códon sem Sentido , RNA Mensageiro/genética , Animais , Diferenciação Celular/genética , Desenvolvimento Embrionário/genética , Humanos , Imunidade Inata/genética , Controle de Qualidade , RNA Mensageiro/metabolismoRESUMO
Due to the high heterogeneity of breast cancers, numerous recent patents describe improved methods of detection and classification which promise better patient prognosis and treatment. In particular, there has been a shift towards more effective genetic screening to identify specific mutations associated with breast tumours, which may lead to "personalised medicine" with improved outcomes. Two challenging areas of breast cancer research involve the development of treatments for the highly aggressive triple negative breast cancer subtype as well as the chemotherapy-resistant cancer stem cell subpopulation. In addition, despite numerous recent advances in breast cancer treatment in woman, male breast cancer remains poorly understood and there are limited therapies available which are developed specifically for men. This review serves to report on important developments in the treatment of breast malignancies patented in the past two years as well as to highlight the current gaps in the field of breast cancer therapeutics and areas which require further study.
Assuntos
Antineoplásicos/uso terapêutico , Neoplasias da Mama Masculina/tratamento farmacológico , Neoplasias da Mama/tratamento farmacológico , Medicina de Precisão , Neoplasias da Mama/metabolismo , Neoplasias da Mama Masculina/metabolismo , Feminino , Humanos , Masculino , Patentes como Assunto , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/metabolismoRESUMO
The Hsp90/Hsp70 organising protein (Hop) is a co-chaperone that mediates the interaction of Hsp90 and Hsp70 molecular chaperones during assembly of Hsp90 complexes in cells. Formation of Hsp90 complexes is a key intermediate step in the maturation and homeostasis of oncoproteins and several hormone receptors. In this paper, we demonstrate that knockdown of Hop decreased migration of Hs578T and MDA-MB-231 breast cancer cells. Hop was identified in isolated pseudopodia fractions; it colocalised with actin in lamellipodia, and co-sedimented with purified actin in vitro. Knockdown of Hop caused a decrease in the level of RhoC GTPase, and significantly inhibited pseudopodia formation in Hs578T cells. Our data suggest that Hop regulates directional cell migration by multiple unknown mechanisms.
