scholarly journals P-Body Formation Is a Consequence, Not the Cause, of RNA-Mediated Gene Silencing

2007 ◽  
Vol 27 (11) ◽  
pp. 3970-3981 ◽  
Author(s):  
Ana Eulalio ◽  
Isabelle Behm-Ansmant ◽  
Daniel Schweizer ◽  
Elisa Izaurralde
Keyword(s):  
Gene Silencing ◽  
Mrna Decay ◽  
Cytoplasmic Domains ◽  
Body Formation ◽  
P Bodies ◽  
Posttranscriptional Gene Regulation ◽  
Nonsense Mediated Mrna Decay ◽  
Body Components ◽  
Silencing Pathways ◽  
Interfering Rna

ABSTRACT P bodies are cytoplasmic domains that contain proteins involved in diverse posttranscriptional processes, such as mRNA degradation, nonsense-mediated mRNA decay (NMD), translational repression, and RNA-mediated gene silencing. The localization of these proteins and their targets in P bodies raises the question of whether their spatial concentration in discrete cytoplasmic domains is required for posttranscriptional gene regulation. We show that processes such as mRNA decay, NMD, and RNA-mediated gene silencing are functional in cells lacking detectable microscopic P bodies. Although P bodies are not required for silencing, blocking small interfering RNA or microRNA silencing pathways at any step prevents P-body formation, indicating that P bodies arise as a consequence of silencing. Consistently, we show that releasing mRNAs from polysomes is insufficient to trigger P-body assembly: polysome-free mRNAs must enter silencing and/or decapping pathways to nucleate P bodies. Thus, even though P-body components play crucial roles in mRNA silencing and decay, aggregation into P bodies is not required for function but is instead a consequence of their activity.

scholarly journals Deadenylation is prerequisite for P-body formation and mRNA decay in mammalian cells

2008 ◽  
Vol 182 (1) ◽  
pp. 89-101 ◽  
Author(s):  
Dinghai Zheng ◽  
Nader Ezzeddine ◽  
Chyi-Ying A. Chen ◽  
Wenmiao Zhu ◽  
Xiangwei He ◽  
...  
Keyword(s):  
Messenger Rna ◽  
Mammalian Cells ◽  
Mrna Decay ◽  
Protein Complexes ◽  
Major Step ◽  
Body Formation ◽  
P Bodies ◽  
Body Components ◽  
Selective Decay ◽  
Dynamic Interplay

Deadenylation is the major step triggering mammalian mRNA decay. One consequence of deadenylation is the formation of nontranslatable messenger RNA (mRNA) protein complexes (messenger ribonucleoproteins [mRNPs]). Nontranslatable mRNPs may accumulate in P-bodies, which contain factors involved in translation repression, decapping, and 5′-to-3′ degradation. We demonstrate that deadenylation is required for mammalian P-body formation and mRNA decay. We identify Pan2, Pan3, and Caf1 deadenylases as new P-body components and show that Pan3 helps recruit Pan2, Ccr4, and Caf1 to P-bodies. Pan3 knockdown causes a reduction of P-bodies and has differential effects on mRNA decay. Knocking down Caf1 or overexpressing a Caf1 catalytically inactive mutant impairs deadenylation and mRNA decay. P-bodies are not detected when deadenylation is blocked and are restored when the blockage is released. When deadenylation is impaired, P-body formation is not restorable, even when mRNAs exit the translating pool. These results support a dynamic interplay among deadenylation, mRNP remodeling, and P-body formation in selective decay of mammalian mRNA.

EV71 2A Protease inhibits P-body formation to promote viral RNA synthesis

2021 ◽  
Author(s):  
Shanshan Fan ◽  
Zihang Xu ◽  
Pengfei Liu ◽  
Yali Qin ◽  
Mingzhou Chen
Keyword(s):  
Viral Replication ◽  
Rna Synthesis ◽  
Enterovirus 71 ◽  
Viral Rna ◽  
Processing Bodies ◽  
Body Formation ◽  
P Bodies ◽  
Body Components ◽  
2A Protease ◽  
Enterovirus 71 Infection

Several viruses were proved to inhibit the formation of RNA processing bodies (P-bodies); however, knowledge regarding whether enterovirus blocks P-body formation remains unclear, and the detailed molecular mechanisms and functions of picornavirus regulation of P-bodies are limited. Here we show the crucial role of 2A protease in inhibiting P-bodies to promote viral replication during enterovirus 71 infection. Moreover, we found that the activity of 2A protease is essential to inhibit P-body formation, which was proved by the result that infection of EV71-2A C110S , the 2A protease activity-inactivated recombinant virus, failed to block the formation of P-bodies. Furthermore, we showed DDX6, a scaffolding protein of P-bodies, interacted with viral RNA to facilitate viral replication rather than viral translation, by using a Renilla luciferase mRNA reporter system and capturing the nascent RNA assay. Altogether, our data firstly demonstrate that the 2A protease of enterovirus inhibits P-body formation to facilitate viral RNA synthesis by recruiting the P-body components to viral RNA. IMPORTANCE Processing bodies (P-bodies) are constitutively present in eukaryotic cells and play an important role in the mRNA cycle, including regulating gene expression and mRNA degradation. P-bodies are the structure that viruses to manipulate to facilitate their survival. Here, we show that the 2A protease alone was efficient to block P-body formation during enterovirus 71 infection and its activity was essential. When the assembly of P-bodies was blocked by 2A, DDX6 and 4E-T which were required for P-body formation bound to viral RNA to facilitate viral RNA synthesis. We propose a model revealing that EV71 manipulates P-body formation to generate an environment that is conducive to viral replication by facilitating viral RNA synthesis: 2A protease blocked P-body assembly to make it possible for virus to take advantage of P-body components.

Phosphorylation of Argonaute proteins: regulating gene regulators

2008 ◽  
Vol 413 (3) ◽  
pp. e7-e9 ◽  
Author(s):  
Sabine Rüdel ◽  
Gunter Meister
Keyword(s):  
Gene Silencing ◽  
Translational Regulation ◽  
Mitogen Activated Protein Kinase ◽  
Rna Turnover ◽  
Processing Bodies ◽  
P Bodies ◽  
Biochemical Journal ◽  
Mitogen Activated Protein ◽  
Silencing Pathways ◽  
Protein Kinase Signalling

Members of the Ago (Argonaute) protein family are the mediators of small RNA-guided gene-silencing pathways including RNAi (RNA interference), translational regulation by miRNAs (microRNAs) and transcriptional silencing. Recent findings by Zeng et al. in this issue of the Biochemical Journal demonstrate that Ago proteins are post-translationally modified by phosphorylation of Ser387. Mutating Ser387 to alanine leads to reduced localization of human Ago2 to cytoplasmic P-bodies (processing bodies), cellular sites where RNA turnover and, at least in part, miRNA-guided gene regulation occurs. Zeng et al. further show that a member of the MAPK (mitogen-activated protein kinase) signalling pathway phosphorylates Ago2 at Ser387, suggesting that Ago2-mediated gene silencing might be linked to distinct signalling pathways.

scholarly journals Decapping complex is essential for functional P-body formation and is buffered by nuclear localization

2020 ◽  
Author(s):  
Kiril Tishinov ◽  
Anne Spang
Keyword(s):  
Endoplasmic Reticulum ◽  
Phase Separation ◽  
Nuclear Localization ◽  
Mrna Decay ◽  
Dynamic Equilibrium ◽  
Local Concentration ◽  
Processing Bodies ◽  
Body Formation ◽  
P Bodies ◽  
Translational Repressor

AbstractmRNA decay is a key step in regulating the cellular proteome. Cytoplasmic mRNA is largely turned over in processing bodies (P-bodies). P-body units assemble to form P-body granules under stress conditions. How this assembly is regulated, however, remains still poorly understood. Here, we show that the translational repressor Scd6 and the decapping stimulator Edc3 act partially redundantly in P-body assembly by capturing the Dcp1/2 decapping complex and preventing it from becoming imported into the nucleus by the karyopherin ß Kap95. Nuclear Dcp1/2 does not drive mRNA decay and might be stored there as a ready releasable pool, indicating a dynamic equilibrium between cytoplasmic and nuclear Dcp1/2. Cytoplasmic Dcp1/2 is linked to Dhh1 via Edc3 and Scd6. Functional P-bodies are present at the endoplasmic reticulum where Dcp2 potentially acts to increase the local concentration of Dhh1 through interaction with Scd6 and Edc3 to drive phase separation and hence P-body formation.

scholarly journals Sam68 regulates EMT through alternative splicing–activated nonsense-mediated mRNA decay of the SF2/ASF proto-oncogene

2010 ◽  
Vol 191 (1) ◽  
pp. 87-99 ◽  
Author(s):  
Cristina Valacca ◽  
Serena Bonomi ◽  
Emanuele Buratti ◽  
Simona Pedrotti ◽  
Francisco Ernesto Baralle ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Mrna Decay ◽  
Epithelial To Mesenchymal Transition ◽  
Mesenchymal Transition ◽  
Splicing Regulator ◽  
Extracellular Stimuli ◽  
Nonsense Mediated Mrna Decay ◽  
Vitro Model ◽  
Interfering Rna

Epithelial-to-mesenchymal transition (EMT) and its reversal (MET) are crucial cell plasticity programs that act during development and tumor metastasis. We have previously shown that the splicing factor and proto-oncogene SF2/ASF impacts EMT/MET through production of a constitutively active splice variant of the Ron proto-oncogene. Using an in vitro model, we now show that SF2/ASF is also regulated during EMT/MET by alternative splicing associated with the nonsense-mediated mRNA decay pathway (AS-NMD). Overexpression and small interfering RNA experiments implicate the splicing regulator Sam68 in AS-NMD of SF2/ASF transcripts and in the choice between EMT/MET programs. Moreover, Sam68 modulation of SF2/ASF splicing appears to be controlled by epithelial cell–derived soluble factors that act through the ERK1/2 signaling pathway to regulate Sam68 phosphorylation. Collectively, our results reveal a hierarchy of splicing factors that integrate splicing decisions into EMT/MET programs in response to extracellular stimuli.

scholarly journals Inhibition of nonsense-mediated mRNA decay (NMD) by a new chemical molecule reveals the dynamic of NMD factors in P-bodies

2007 ◽  
Vol 178 (7) ◽  
pp. 1145-1160 ◽  
Author(s):  
Sébastien Durand ◽  
Nicolas Cougot ◽  
Florence Mahuteau-Betzer ◽  
Chi-Hung Nguyen ◽  
David S. Grierson ◽  
...  
Keyword(s):  
Quality Control ◽  
Mrna Decay ◽  
Control Mechanism ◽  
Premature Termination Codon ◽  
Processing Bodies ◽  
P Bodies ◽  
Molecule Inhibitor ◽  
Nonsense Mediated Mrna Decay ◽  
Quality Control Mechanism ◽  
Insight Into

In mammals, nonsense-mediated mRNA decay (NMD) is a quality-control mechanism that degrades mRNA harboring a premature termination codon to prevent the synthesis of truncated proteins. To gain insight into the NMD mechanism, we identified NMD inhibitor 1 (NMDI 1) as a small molecule inhibitor of the NMD pathway. We characterized the mode of action of this compound and demonstrated that it acts upstream of hUPF1. NMDI 1 induced the loss of interactions between hSMG5 and hUPF1 and the stabilization of hyperphosphorylated isoforms of hUPF1. Incubation of cells with NMDI 1 allowed us to demonstrate that NMD factors and mRNAs subject to NMD transit through processing bodies (P-bodies), as is the case in yeast. The results suggest a model in which mRNA and NMD factors are sequentially recruited to P-bodies.

UPF1 P-body localization

2008 ◽  
Vol 36 (4) ◽  
pp. 698-700 ◽  
Author(s):  
Saverio Brogna ◽  
Preethi Ramanathan ◽  
Jikai Wen
Keyword(s):  
Mrna Decay ◽  
Translation Termination ◽  
Processing Bodies ◽  
Cytoplasmic Granules ◽  
P Bodies ◽  
The Core ◽  
Nonsense Mediated Mrna Decay ◽  
Premature Translation Termination

NMD (nonsense-mediated mRNA decay) is a mechanism that degrades transcripts containing PTCs (premature translation termination codons). NMD is a translation-associated process that is expected to take place throughout the cytoplasm. However, recent studies have indicated that the core NMD factors UPF1 (up-frameshift-1), UPF2 and UPF3 can associate with P-bodies (processing bodies), which are large cytoplasmic granules replete with proteins involved in general mRNA decay and related processes. It has been proposed that UPF1 directs PTC-containing mRNAs to P-bodies and triggers decay. Here, we discuss the link between P-bodies and NMD in view of recent studies that suggest that P-bodies are not required for NMD in Drosophila.

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