Decapping the message: a beginning or an end

2006 ◽  
Vol 34 (1) ◽  
pp. 35-38 ◽  
Author(s):  
H. Liu ◽  
M. Kiledjian
Keyword(s):  
Mrna Stability ◽  
Mrna Turnover ◽  
Decapping Enzyme ◽  
Decapping Enzymes

Removal of the mRNA 5′ cap is an important step in the regulation of mRNA stability. mRNAs are degraded by at least two distinct exonucleolytic decay pathways, one from the 5′ end, and the second from the 3′ end. Two major cellular decapping enzymes have been identified, and each primarily functions in one of the two decay pathways. The Dcp2 decapping enzyme utilizes capped mRNA as substrate and hydrolyses the cap to release m7GDP (N7-methyl GDP), while a scavenger decapping enzyme, DcpS, utilizes cap dinucleotides or capped oligonucleotides as substrate and releases m7GMP (N7-methyl GMP). In this review, we will highlight the function of different decapping enzymes and their role in mRNA turnover.

scholarly journals Scavenger Decapping Activity Facilitates 5′ to 3′ mRNA Decay

2005 ◽  
Vol 25 (22) ◽  
pp. 9764-9772 ◽  
Author(s):  
Hudan Liu ◽  
Megerditch Kiledjian
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mrna Stability ◽  
Mrna Decay ◽  
Hydrolytic Activity ◽  
Mrna Degradation ◽  
Mrna Turnover ◽  
Threefold Increase ◽  
Mechanistic Analysis ◽  
Decapping Enzyme ◽  
Mrna Half Life

ABSTRACT mRNA degradation occurs through distinct pathways, one primarily from the 5′ end of the mRNA and the second from the 3′ end. Decay from the 3′ end generates the m7GpppN cap dinucleotide, which is subsequently hydrolyzed to m7Gp and ppN in Saccharomyces cerevisiae by a scavenger decapping activity termed Dcs1p. Although Dcs1p functions in the last step of mRNA turnover, we demonstrate that its activity modulates earlier steps of mRNA decay. Disruption of the DCS1 gene manifests a threefold increase of the TIF51A mRNA half-life. Interestingly, the hydrolytic activity of Dcs1p was essential for the altered mRNA turnover, as Dcs1p, but not a catalytically inactive Dcs1p mutant, complemented the increased mRNA stability. Mechanistic analysis revealed that 5′ to 3′ exoribonucleolytic activity was impeded in the dcs1Δ strain, resulting in the accumulation of uncapped mRNA. These data define a new role for the Dcs1p scavenger decapping enzyme and demonstrate a novel mechanism whereby the final step in the 3′ mRNA decay pathway can influence 5′ to 3′ exoribonucleolytic activity.

scholarly journals A Novel NAD-RNA Decapping Pathway Discovered by Synthetic Light-Up NAD-RNAs

Biomolecules ◽  
2020 ◽  
Vol 10 (4) ◽  
pp. 513
Author(s):  
Florian Abele ◽  
Katharina Höfer ◽  
Patrick Bernhard ◽  
Julia Grawenhoff ◽  
Maximilian Seidel ◽  
...  
Keyword(s):  
High Throughput Screening ◽  
Rna Degradation ◽  
Enzymatic Reactions ◽  
Nicotinamide Mononucleotide ◽  
Decapping Enzyme ◽  
New Research ◽  
Rna Decapping ◽  
Decapping Enzymes

The complexity of the transcriptome is governed by the intricate interplay of transcription, RNA processing, translocation, and decay. In eukaryotes, the removal of the 5’-RNA cap is essential for the initiation of RNA degradation. In addition to the canonical 5’-N7-methyl guanosine cap in eukaryotes, the ubiquitous redox cofactor nicotinamide adenine dinucleotide (NAD) was identified as a new 5’-RNA cap structure in prokaryotic and eukaryotic organisms. So far, two classes of NAD-RNA decapping enzymes have been identified, namely Nudix enzymes that liberate nicotinamide mononucleotide (NMN) and DXO-enzymes that remove the entire NAD cap. Herein, we introduce 8-(furan-2-yl)-substituted NAD-capped-RNA (FurNAD-RNA) as a new research tool for the identification and characterization of novel NAD-RNA decapping enzymes. These compounds are found to be suitable for various enzymatic reactions that result in the release of a fluorescence quencher, either nicotinamide (NAM) or nicotinamide mononucleotide (NMN), from the RNA which causes a fluorescence turn-on. FurNAD-RNAs allow for real-time quantification of decapping activity, parallelization, high-throughput screening and identification of novel decapping enzymes in vitro. Using FurNAD-RNAs, we discovered that the eukaryotic glycohydrolase CD38 processes NAD-capped RNA in vitro into ADP-ribose-modified-RNA and nicotinamide and therefore might act as a decapping enzyme in vivo. The existence of multiple pathways suggests that the decapping of NAD-RNA is an important and regulated process in eukaryotes.

scholarly journals A non-canonical role for the EDC4 decapping factor in regulating MARF1-mediated mRNA decay

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
William R Brothers ◽  
Steven Hebert ◽  
Claudia L Kleinman ◽  
Marc R Fabian
Keyword(s):  
Essential Role ◽  
Mrna Decay ◽  
Rna Binding ◽  
Mrna Turnover ◽  
Core Component ◽  
P Bodies ◽  
Target Mrnas ◽  
Binding Domains ◽  
Bona Fide ◽  
Decapping Enzyme

EDC4 is a core component of processing (P)-bodies that binds the DCP2 decapping enzyme and stimulates mRNA decay. EDC4 also interacts with mammalian MARF1, a recently identified endoribonuclease that promotes oogenesis and contains a number of RNA binding domains, including two RRMs and multiple LOTUS domains. How EDC4 regulates MARF1 action and the identity of MARF1 target mRNAs is not known. Our transcriptome-wide analysis identifies bona fide MARF1 target mRNAs and indicates that MARF1 predominantly binds their 3’ UTRs via its LOTUS domains to promote their decay. We also show that a MARF1 RRM plays an essential role in enhancing its endonuclease activity. Importantly, we establish that EDC4 impairs MARF1 activity by preventing its LOTUS domains from binding target mRNAs. Thus, EDC4 not only serves as an enhancer of mRNA turnover that binds DCP2, but also as a repressor that binds MARF1 to prevent the decay of MARF1 target mRNAs.

scholarly journals ZBP1 regulates mRNA stability during cellular stress

2006 ◽  
Vol 175 (4) ◽  
pp. 527-534 ◽  
Author(s):  
Nadine Stöhr ◽  
Marcell Lederer ◽  
Claudia Reinke ◽  
Sylke Meyer ◽  
Mechthild Hatzfeld ◽  
...  
Keyword(s):  
Mrna Stability ◽  
Cellular Stress ◽  
Specific Protein ◽  
Mrna Turnover ◽  
Processing Bodies ◽  
Target Mrnas ◽  
Specific Mrnas ◽  
Mrna Binding ◽  
Mrna Targeting ◽  
Translational Suppression

An essential constituent of the integrated stress response (ISR) is a reversible translational suppression. This mRNA silencing occurs in distinct cytoplasmic foci called stress granules (SGs), which transiently associate with processing bodies (PBs), typically serving as mRNA decay centers. How mRNAs are protected from degradation in these structures remains elusive. We identify that Zipcode-binding protein 1 (ZBP1) regulates the cytoplasmic fate of specific mRNAs in nonstressed cells and is a key regulator of mRNA turnover during the ISR. ZBP1 association with target mRNAs in SGs was not essential for mRNA targeting to SGs. However, ZBP1 knockdown induced a selective destabilization of target mRNAs during the ISR, whereas forced expression increased mRNA stability. Our results indicate that although targeting of mRNAs to SGs is nonspecific, the stabilization of mRNAs during cellular stress requires specific protein–mRNA interactions. These retain mRNAs in SGs and prevent premature decay in PBs. Hence, mRNA-binding proteins are essential for translational adaptation during cellular stress by modulating mRNA turnover.

scholarly journals Sbp1p Affects Translational Repression and Decapping in Saccharomyces cerevisiae

2006 ◽  
Vol 26 (13) ◽  
pp. 5120-5130 ◽  
Author(s):  
Scott P. Segal ◽  
Travis Dunckley ◽  
Roy Parker
Keyword(s):  
Rna Binding ◽  
Regulation Of Gene Expression ◽  
Glucose Deprivation ◽  
Decay Rates ◽  
The Body ◽  
Translational Repression ◽  
Mrna Turnover ◽  
Major Pathway ◽  
Body Formation ◽  
Decapping Enzyme

ABSTRACT The relationship between translation and mRNA turnover is critical to the regulation of gene expression. One major pathway for mRNA turnover occurs by deadenylation, which leads to decapping and subsequent 5′-to-3′ degradation of the body of the mRNA. Prior to mRNA decapping, a transcript exits translation and enters P bodies to become a potential decapping substrate. To understand the transition from translation to decapping, it is important to identify the factors involved in this process. In this work, we identify Sbp1p (formerly known as Ssb1p), an abundant RNA binding protein, as a high-copy-number suppressor of a conditional allele in the decapping enzyme. Sbp1p overexpression restores normal decay rates in decapping-defective strains and increases P-body size and number. In addition, Sbp1p promotes translational repression of mRNA during glucose deprivation. Moreover, P-body formation is reduced in strains lacking Sbp1p. Sbp1p acts in conjunction with Dhh1p, as it is required for translational repression and P-body formation in pat1Δ strains under these conditions. These results identify Sbp1p as a new protein that functions in the transition of mRNAs from translation to an mRNP complex destined for decapping.

scholarly journals Analysis of Mutations in the Yeast mRNA Decapping Enzyme

Genetics ◽  
1999 ◽  
Vol 151 (4) ◽  
pp. 1273-1285 ◽  
Author(s):  
Sundaresan Tharun ◽  
Roy Parker
Keyword(s):  
Specific Activity ◽  
Temperature Sensitive ◽  
Loss Of Function ◽  
Alanine Scanning Mutagenesis ◽  
Mrna Decapping ◽  
Nonsense Codons ◽  
Decapping Enzyme ◽  
Decapping Enzymes

Abstract A major mechanism of mRNA decay in yeast is initiated by deadenylation, followed by mRNA decapping, which exposes the transcript to 5′ to 3′ exonucleolytic degradation. The decapping enzyme that removes the 5′ cap structure is encoded by the DCP1 gene. To understand the function of the decapping enzyme, we used alanine scanning mutagenesis to create 31 mutant versions of the enzyme, and we examined the effects of the mutations both in vivo and in vitro. Two types of mutations that affected mRNA decapping in vivo were identified, including a temperature-sensitive allele. First, two mutants produced decapping enzymes that were defective for decapping in vitro, suggesting that these mutated residues are required for enzymatic activity. In contrast, several mutants that moderately affected mRNA decapping in vivo yielded decapping enzymes that had at least the same specific activity as the wild-type enzyme in vitro. Combination of alleles within this group yielded decapping enzymes that showed a strong loss of function in vivo, but that still produced fully active enzymes in vitro. This suggested that interactions of the decapping enzyme with other factors may be required for efficient decapping in vivo, and that these particular mutations may be disrupting such interactions. Interestingly, partial loss of decapping activity in vivo led to a defect in normal deadenylation-dependent decapping, but it did not affect the rapid deadenylation-independent decapping triggered by early nonsense codons. This observation suggested that these two types of mRNA decapping differ in their requirements for the decapping enzyme.

scholarly journals Impact of Methods on the Measurement of mRNA Turnover

2017 ◽  
Author(s):  
Takeo Wada ◽  
Attila Becskei
Keyword(s):  
Mrna Stability ◽  
Rna Degradation ◽  
Mrna Turnover ◽  
Gene Control ◽  
Cellular Activity ◽  
Transcription Inhibition ◽  
Rna Molecules ◽  
Metabolic Labelling ◽  
Differential Stability ◽  
Made In

The turnover of the RNA molecules is determined by the rates of transcription and RNA degradation. Several methods have been developed to study mRNA turnover since the beginnings of molecular biology. Here we summarize the main methods to measure RNA half-life: transcription inhibition, gene control and metabolic labelling. These methods were used to detect the cellular activity of the mRNAs degradation machinery, including the exo-ribonuclease Xrn1 and the exosome. Less progress has been made in the study of the differential stability of mature RNAs because the different methods have often yielded inconsistent results so that an mRNA considered to be stable can be classified as unstable by another method. Recent advances in the systematic comparison of different method variants in yeast have permitted the identification of the least invasive methodologies that reflect half-lives the most faithfully, which is expected to open the way for a consistent quantitative analysis of the determinants of mRNA stability.

scholarly journals Is mRNA decapping activity of ApaH like phosphatases (ALPH’s) the reason for the loss of cytoplasmic ALPH’s in all eukaryotes but Kinetoplastida?

2020 ◽  
Author(s):  
Paula Andrea Castañeda Londoño ◽  
Nicole Banholzer ◽  
Bridget Bannermann ◽  
Susanne Kramer
Keyword(s):  
Catalytic Domain ◽  
Last Common Ancestor ◽  
Transmembrane Helices ◽  
Mrna Decapping ◽  
Nudix Hydrolases ◽  
Eukaryotic Proteomes ◽  
Decapping Enzyme ◽  
Decapping Enzymes ◽  
Intracellular Localisation

ABSTRACTBackgroundApaH like phosphatases (ALPHs) originate from the bacterial ApaH protein and are present in eukaryotes of all eukaryotic super-groups; still, only two proteins have been functionally characterised. One is ALPH1 from the Kinetoplastid Trypanosoma brucei that we recently found to be the mRNA decapping enzyme of the parasite. mRNA decapping by ALPHs is unprecedented in eukaryotes, which usually use nudix hydrolases, but the bacterial ancestor protein ApaH was recently found to decap non-conventional caps of bacterial mRNAs. These findings prompted us to explore whether mRNA decapping by ALPHs is restricted to Kinetoplastida or more widespread among eukaryotes.ResultsWe screened 824 eukaryotic proteomes with a newly developed Python-based algorithm for the presence of ALPHs and used the data to refine phylogenetic distribution, conserved features, additional domains and predicted intracellular localisation of ALPHs. We found that most eukaryotes have either no ALPH (500/824) or very short ALPHs, consisting almost exclusively of the catalytic domain. These ALPHs had mostly predicted non-cytoplasmic localisations, often supported by the presence of transmembrane helices and signal peptides and in two cases (one in this study) by experimental data. The only exceptions were ALPH1 homologues from Kinetoplastida, that all have unique C-terminal and mostly unique N-terminal extension, and at least the T. brucei enzyme localises to the cytoplasm. Surprisingly, despite of these non-cytoplasmic localisations, ALPHs from all eukaryotic super-groups had in vitro mRNA decapping activity.ConclusionsALPH was present in the last common ancestor of eukaryotes, but most eukaryotes have either lost the enzyme since, or use it exclusively outside the cytoplasm in organelles in a version consisting of the catalytic domain only. While our data provide no evidence for the presence of further mRNA decapping enzymes among eukaryotic ALPHs, the broad substrate range of ALPHs that includes mRNA caps provides an explanation for the selection against the presence of a cytoplasmic ALPH protein as a mean to protect mRNAs from unregulated degradation. Kinetoplastida succeeded to exploit ALPH as their mRNA decapping enzyme, likely using the Kinetoplastida-unique N- and C-terminal extensions for regulation.

Vaccinia virus D10 has broad decapping activity that is regulated by mRNA splicing

2021 ◽  
Author(s):  
Michael Ly ◽  
Hannah M. Burgess ◽  
Ian Mohr ◽  
Britt A Glaunsinger
Keyword(s):  
Vaccinia Virus ◽  
Dominant Role ◽  
Mrna Splicing ◽  
Mrna Turnover ◽  
Viral Mrna ◽  
Wild Type ◽  
Intronless Genes ◽  
Cell Transcriptome ◽  
Encoding Genes ◽  
Decapping Enzymes

The mRNA 5’ cap structure serves both to protect transcripts from degradation and promote their translation. Cap removal is thus an integral component of mRNA turnover that is carried out by cellular decapping enzymes, whose activity is tightly regulated and coupled to other stages of the mRNA decay pathway. The poxvirus vaccinia virus (VACV) encodes its own decapping enzymes, D9 and D10, that act on cellular and viral mRNA, but may be regulated differently than their cellular counterparts. Here, we evaluated the targeting potential of these viral enzymes using RNA sequencing from cells infected with wild-type and decapping mutant versions of VACV as well as in uninfected cells expressing D10. We found that D9 and D10 target an overlapping subset of viral transcripts but that D10 plays a dominant role in depleting the vast majority of human transcripts, although not in an indiscriminate manner. Unexpectedly, the splicing architecture of a gene influences how robustly its corresponding transcript is targeted by D10, as transcripts derived from intronless genes are less susceptible to enzymatic decapping by D10. As all VACV genes are intronless, preferential decapping of transcripts from intron-encoding genes provides an unanticipated mechanism for the virus to disproportionately deplete host transcripts and remodel the infected cell transcriptome.

Functions of microRNAs in Drosophila development

2010 ◽  
Vol 38 (4) ◽  
pp. 1137-1143 ◽  
Author(s):  
Christopher I. Jones ◽  
Sarah F. Newbury
Keyword(s):  
Mrna Stability ◽  
Molecular Mechanisms ◽  
Mrna Translation ◽  
Mrna Degradation ◽  
Mrna Turnover ◽  
Factors Affecting ◽  
Biologically Relevant ◽  
The Core ◽  
Cellular Factors

Control of mRNA translation and degradation has been shown to be key in the development of complex organisms. The core mRNA degradation machinery is highly conserved in eukaryotes and relies on processive degradation enzymes gaining access to the mRNA. Control of mRNA stability in eukaryotes is also intimately linked to the regulation of translation. A key question in the control of mRNA turnover concerns the mechanisms whereby particular mRNAs are specifically degraded in response to cellular factors. Recently, microRNAs have been shown to bind specifically to mRNAs and regulate their expression via repression of translation and/or degradation. To understand the molecular mechanisms during microRNA repression of mRNAs, it is necessary to identify their biologically relevant targets. However, computational methods have so far proved unreliable, therefore verification of biologically important targets at present requires experimental analysis. The present review aims to outline the mechanisms of mRNA degradation and then focus on the role of microRNAs as factors affecting particular Drosophila developmental processes via their post-transcriptional effects on mRNA degradation and translation. Examples of experimentally verified targets of microRNAs in Drosophila are summarized.

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