16,220 results on '"Decapping"'
Search Results
2. RNA degradation triggered by decapping is largely independent of initial deadenylation
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Audebert, Léna, Feuerbach, Frank, Zedan, Mostafa, Schürch, Alexandra P, Decourty, Laurence, Namane, Abdelkader, Permal, Emmanuelle, Weis, Karsten, Badis, Gwenaël, and Saveanu, Cosmin
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- 2024
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3. Human DCP1 is crucial for mRNA decapping and possesses paralog-specific gene regulating functions
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Ting-Wen Chen, Hsiao-Wei Liao, Michelle Noble, Jing-Yi Siao, Yu-Hsuan Cheng, Wei-Chung Chiang, Yi-Tzu Lo, and Chung-Te Chang
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mRNA decapping ,DCP1 paralogs ,DCP2 ,mRNA decay pathway ,post-transcriptional regulation ,Medicine ,Science ,Biology (General) ,QH301-705.5 - Abstract
The mRNA 5'-cap structure removal by the decapping enzyme DCP2 is a critical step in gene regulation. While DCP2 is the catalytic subunit in the decapping complex, its activity is strongly enhanced by multiple factors, particularly DCP1, which is the major activator in yeast. However, the precise role of DCP1 in metazoans has yet to be fully elucidated. Moreover, in humans, the specific biological functions of the two DCP1 paralogs, DCP1a and DCP1b, remain largely unknown. To investigate the role of human DCP1, we generated cell lines that were deficient in DCP1a, DCP1b, or both to evaluate the importance of DCP1 in the decapping machinery. Our results highlight the importance of human DCP1 in decapping process and show that the EVH1 domain of DCP1 enhances the mRNA-binding affinity of DCP2. Transcriptome and metabolome analyses outline the distinct functions of DCP1a and DCP1b in human cells, regulating specific endogenous mRNA targets and biological processes. Overall, our findings provide insights into the molecular mechanism of human DCP1 in mRNA decapping and shed light on the distinct functions of its paralogs.
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- 2024
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4. Toll/interleukin-1 receptor (TIR) domain-containing proteins have NAD-RNA decapping activity
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Wang, Xufeng, Yu, Dongli, Yu, Jiancheng, Hu, Hao, Hang, Runlai, Amador, Zachary, Chen, Qi, Chai, Jijie, and Chen, Xuemei
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- 2024
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5. Effect of the mRNA decapping enzyme scavenger (DCPS) inhibitor RG3039 on glioblastoma
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Duan, Hao, Xie, Yuan, Wu, Suwen, Zhao, Guangyin, Zeng, Zhen, Hu, Hongrong, Yu, Yanjiao, Hu, Wanming, Yang, Yuanzhong, Chen, Yukun, Xie, Haoqun, Chen, Zexin, Zhang, Gao, Flaherty, Keith T., Hu, Shanshan, Xu, Haineng, Ma, Wenjuan, and Mou, Yonggao
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- 2024
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6. Structure of the poxvirus decapping enzyme D9 reveals its mechanism of cap recognition and catalysis
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Peters, Jessica K, Tibble, Ryan W, Warminski, Marcin, Jemielity, Jacek, and Gross, John D
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Biochemistry and Cell Biology ,Biological Sciences ,Rare Diseases ,Emerging Infectious Diseases ,Genetics ,Infectious Diseases ,Small Pox ,1.1 Normal biological development and functioning ,Infection ,Catalysis ,Endoribonucleases ,Poxviridae ,RNA Caps ,RNA ,Double-Stranded ,RNA ,Messenger ,Vaccinia virus ,Viral Proteins ,cap recognition ,catalysis ,dsRNA decay ,host shutoff ,innate immunity ,mRNA decay ,poxvirus decapping enzyme D9 ,virus-host interactions ,Chemical Sciences ,Information and Computing Sciences ,Biophysics ,Biological sciences ,Chemical sciences - Abstract
Poxviruses encode decapping enzymes that remove the protective 5' cap from both host and viral mRNAs to commit transcripts for decay by the cellular exonuclease Xrn1. Decapping by these enzymes is critical for poxvirus pathogenicity by means of simultaneously suppressing host protein synthesis and limiting the accumulation of viral double-stranded RNA (dsRNA), a trigger for antiviral responses. Here we present a high-resolution structural view of the vaccinia virus decapping enzyme D9. This Nudix enzyme contains a domain organization different from other decapping enzymes in which a three-helix bundle is inserted into the catalytic Nudix domain. The 5' mRNA cap is positioned in a bipartite active site at the interface of the two domains. Specificity for the methylated guanosine cap is achieved by stacking between conserved aromatic residues in a manner similar to that observed in canonical cap-binding proteins VP39, eIF4E, and CBP20, and distinct from eukaryotic decapping enzyme Dcp2.
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- 2022
7. Heat stress-induced decapping of WUSCHEL mRNA enhances stem cell thermotolerance in Arabidopsis
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Liu, Sumei, Wu, Haijun, and Zhao, Zhong
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- 2024
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8. A distinct RNA recognition mechanism governs Np₄ decapping by RppH
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Levenson-Palmer, Rose, Luciano, Daniel J., Vasilyev, Nikita, Nuthanakanti, Ashok, Serganov, Alexander, and Belasco, Joel G.
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- 2022
9. mRNA decapping : finding the right structures
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Charenton, Clément and Graille, Marc
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- 2018
10. PAT mRNA decapping factors are required for proper development in Arabidopsis
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Zuo, Zhangli, Roux, Milena Edna, Dagdas, Yasin F., Rodriguez, Eleazar, Petersen, Morten, Zuo, Zhangli, Roux, Milena Edna, Dagdas, Yasin F., Rodriguez, Eleazar, and Petersen, Morten
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Evolutionarily conserved protein associated with topoisomerase II (PAT1) proteins activate mRNA decay through binding mRNA and recruiting decapping factors to optimize posttranscriptional reprogramming. Here, we generated multiple mutants of pat1, pat1 homolog 1 (path1), and pat1 homolog 2 (path2) and discovered that pat triple mutants exhibit extremely stunted growth and all mutants with pat1 exhibit leaf serration while mutants with pat1 and path1 display short petioles. All three PATs can be found localized to processing bodies and all PATs can target ASYMMETRIC LEAVES 2-LIKE 9 transcripts for decay to finely regulate apical hook and lateral root development. In conclusion, PATs exhibit both specific and redundant functions during different plant growth stages and our observations underpin the selective regulation of the mRNA decay machinery for proper development., Evolutionarily conserved protein associated with topoisomerase II (PAT1) proteins activate mRNA decay through binding mRNA and recruiting decapping factors to optimize posttranscriptional reprogramming. Here, we generated multiple mutants of pat1, pat1 homolog 1 (path1), and pat1 homolog 2 (path2) and discovered that pat triple mutants exhibit extremely stunted growth and all mutants with pat1 exhibit leaf serration while mutants with pat1 and path1 display short petioles. All three PATs can be found localized to processing bodies and all PATs can target ASYMMETRIC LEAVES 2-LIKE 9 transcripts for decay to finely regulate apical hook and lateral root development. In conclusion, PATs exhibit both specific and redundant functions during different plant growth stages and our observations underpin the selective regulation of the mRNA decay machinery for proper development.
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- 2024
11. Decapping enzyme 2 is a novel immune-related biomarker that predicts poor prognosis in glioma.
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Mei, Yuran, Lv, Qiaoli, Tan, Zilong, Zhang, Zhe, Ji, Yulong, Chen, Shuhui, and Shen, Xiaoli
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This study analyzed sequencing and clinical data from the Cancer Genome Atlas (TCGA) and gene expression synthesis, and used Chinese glioma Genome Atlas (CGGA) data for external validation. The expression of DCP2 in normal brain and tumor tissue was compared. We analyzed the clinical and molecular characteristics and prognostic value of DCP2 in glioma. In addition, DCP2 expression levels were evaluated in 30 glioma tissue samples and upregulated in glioma samples compared to normal brain tissue (p < 0.001). Multivariate data analysis from TCGA showed that increased DCP2 expression was an independent risk factor for overall survival and prognosis of glioma patients. As indicated by the analysis of the TCGA data set. The expression level of DCP2 is closely related to tumor immunity, including tumor immune cell infiltration, immune score, and co-expression of multiple immune-related genes. In addition, DCP2 was positively correlated with IL-6 and IL-7. Glioma cell proliferation and invasion were evaluated using cell viability, colony formation, wound healing, and transwell assays.Apoptosis and cell cycle were detected by flow cytometry. DCP2 promoted the proliferation, invasion and migration of glioma cells T98G and U251, inhibited apoptosis and blocked the S phase of the cell cycle. As a result of the altered expression of DCP2, a new prognostic biomarker may be identified that can improve patient survival.These findings suggest DCP2 as a potential biomarker for the prognosis of glioma and a candidate immunotherapy target. [ABSTRACT FROM AUTHOR]
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- 2024
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12. Loss of SRSF3 in Cardiomyocytes Leads to Decapping of Contraction-Related mRNAs and Severe Systolic Dysfunction
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Ortiz-Sánchez, Paula, Villalba Orero, María, López-Olañeta, Marina M., Larrasa-Alonso, Javier, Sánchez-Cabo, Fátima, Martí-Gómez, Carlos, Camafeita, Emilio, Gómez-Salinero, Jesús M., Ramos-Hernández, Laura, Nielsen, Peter J., Vázquez, Jesús, Müller-McNicoll, Michaela, García-Pavía, Pablo, Lara-Pezzi, Enrique, Ortiz-Sánchez, Paula, Villalba Orero, María, López-Olañeta, Marina M., Larrasa-Alonso, Javier, Sánchez-Cabo, Fátima, Martí-Gómez, Carlos, Camafeita, Emilio, Gómez-Salinero, Jesús M., Ramos-Hernández, Laura, Nielsen, Peter J., Vázquez, Jesús, Müller-McNicoll, Michaela, García-Pavía, Pablo, and Lara-Pezzi, Enrique
- Abstract
This study was supported by grants from the European Union (CardioNeT-ITN-289600 and CardioNext-ITN-608027 to E. Lara-Pezzi), from the Spanish Ministerio de Economía y Competitividad (RTI2018-096961-B-I00, SAF2015-65722-R, and SAF2012-31451 to E. Lara-Pezzi; BIO2015-67580-P and PGC2018-097019-B-I00 to J. Vázquez), the Spanish Carlos III Institute of Health (CPII14/00027 to E. Lara-Pezzi, RD12/0042/066 to P. García-Pavía and E. Lara-Pezzi, and RD12/0042/0056, PRB2-IPT13/0001-ISCIII-SGEFI/FEDER, ProteoRed to J. Vázquez), the Madrid Regional Government (2010-BMD-2321 Fibroteam to E. Lara-Pezzi). This study was also supported by the Plan Estatal de I+D+I 2013–2016—European Regional Development Fund (ERDF) A way of making Europe, Spain. The CNIC is supported by the Ministerio de Ciencia, Innovación y Universidades (MCNU) and the Pro CNIC Foundation, and is a Severo Ochoa Center of Excellence (SEV-2015-0505)., Rationale: RBPs (RNA binding proteins) play critical roles in the cell by regulating mRNA transport, splicing, editing, and stability. The RBP SRSF3 (serine/arginine-rich splicing factor 3) is essential for blastocyst formation and for proper liver development and function. However, its role in the heart has not been explored. Objective: To investigate the role of SRSF3 in cardiac function. Methods and Results: Cardiac SRSF3 expression was high at mid gestation and decreased during late embryonic development. Mice lacking SRSF3 in the embryonic heart showed impaired cardiomyocyte proliferation and died in utero. In the adult heart, SRSF3 expression was reduced after myocardial infarction, suggesting a possible role in cardiac homeostasis. To determine the role of this RBP in the adult heart, we used an inducible, cardiomyocyte-specific SRSF3 knockout mouse model. After SRSF3 depletion in cardiomyocytes, mice developed severe systolic dysfunction that resulted in death within 8 days. RNA-Seq analysis revealed downregulation of mRNAs encoding sarcomeric and calcium handling proteins. Cardiomyocyte-specific SRSF3 knockout mice also showed evidence of alternative splicing of mTOR (mammalian target of rapamycin) mRNA, generating a shorter protein isoform lacking catalytic activity. This was associated with decreased phosphorylation of 4E-BP1 (eIF4E-binding protein 1), a protein that binds to eIF4E (eukaryotic translation initiation factor 4E) and prevents mRNA decapping. Consequently, we found increased decapping of mRNAs encoding proteins involved in cardiac contraction. Decapping was partially reversed by mTOR activation. Conclusions: We show that cardiomyocyte-specific loss of SRSF3 expression results in decapping of critical mRNAs involved in cardiac contraction. The molecular mechanism underlying this effect likely involves the generation of a short mTOR isoform by alternative splicing, resulting in reduced 4E-BP1 phosphorylation. The identification of mRNA decappin, Unión Europea, Ministerio de Economía y Competitividad, de la Comunidad de Madrid, Plan Estatal de I+D+I 2013–2016 European Regional Development Fund (ERDF), “A way to build Europe” de la European Regional Development Fund (ERDF)., The Pro CNIC Foundation, and is a Severo Ochoa Center of Excellence, Ministerio de Ciencia, Innovación y Universidades (MCNU), Depto. de Medicina y Cirugía Animal, Fac. de Veterinaria, TRUE, pub
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- 2024
13. Purification of Enzymatically Active Xrn1 for Removal of Non-capped mRNAs from In Vitro Transcription Reactions and Evaluation of mRNA Decapping Status In Vivo.
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Drążkowska K, Tomecki R, and Tudek A
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- RNA Stability, Recombinant Proteins genetics, Recombinant Proteins metabolism, Recombinant Proteins isolation & purification, Exoribonucleases genetics, Exoribonucleases metabolism, RNA Caps genetics, RNA Caps metabolism, RNA, Messenger genetics, RNA, Messenger isolation & purification, RNA, Messenger metabolism, Transcription, Genetic
- Abstract
The cap is a 7-methylguanosine attached to the first messenger RNA (mRNA) nucleotide with a 5'-5' triphosphate bridge. This conserved eukaryotic modification confers stability to the transcripts and is essential for translation initiation. The specific mechanisms that govern transcript cytoplasmic longevity and translatability were always of substantial interest. Multiple works aimed at modeling mRNA decay mechanisms, including the onset of decapping, which is the rate-limiting step of mRNA decay. Additionally, with the recent advances in RNA-based vaccines, the importance of efficient synthesis of fully functional mRNAs has increased. Non-capped mRNAs arising during in vitro transcription are highly immunogenic, and multiple approaches were developed to reduce their levels. Efficient and low-cost methods for elimination of non-capped mRNAs in vitro are therefore essential to basic sciences and to pharmaceutical applications. Here, we present a protocol for heterologous expression and purification of catalytically active recombinant Xrn1 from Thermothelomyces (Myceliophthora) thermophilus (Tt_Xrn1). We also describe protocols needed to verify the enzyme quality., (© 2025. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)
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- 2025
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14. Decapping NAD-RNAs: TIR domain-containing proteins stand out for specificity
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Cao, Dechang
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- 2024
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15. mRNA Decapping Activator Pat1 Is Required for Efficient Yeast Adaptive Transcriptional Responses via the Cell Wall Integrity MAPK Pathway
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Pulido, Verónica, Rodríguez-Peña, Jose M., Alonso, Graciela, Sanz, Ana Belén, Arroyo, Javier, and García, Raúl
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- 2024
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16. Changes in conformational equilibria regulate the activity of the Dcp2 decapping enzyme
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Wurm, Jan Philip, Holdermann, Iris, Overbeck, Jan H., Mayer, Philipp H. O., and Sprangers, Remco
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- 2017
17. Non-Redundant Roles for the Human mRNA Decapping Cofactor Paralogs DCP1a and DCP1b
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Vukovic, Ivana, Barnada, Samantha M., Ruffin, Jonathan W., Karlin, Jon, Lokareddy, Ravi Kumar, Cingolani, Gino, McMahon, Steven B., Vukovic, Ivana, Barnada, Samantha M., Ruffin, Jonathan W., Karlin, Jon, Lokareddy, Ravi Kumar, Cingolani, Gino, and McMahon, Steven B.
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Eukaryotic gene expression is regulated at the transcriptional and post-transcriptional levels, with disruption of regulation contributing significantly to human diseases. The 5' m7G mRNA cap is a central node in post-transcriptional regulation, participating in both mRNA stabilization and translation efficiency. In mammals, DCP1a and DCP1b are paralogous cofactor proteins of the mRNA cap hydrolase DCP2. As lower eukaryotes have a single DCP1 cofactor, the functional advantages gained by this evolutionary divergence remain unclear. We report the first functional dissection of DCP1a and DCP1b, demonstrating that they are non-redundant cofactors of DCP2 with unique roles in decapping complex integrity and specificity. DCP1a is essential for decapping complex assembly and interactions between the decapping complex and mRNA cap-binding proteins. DCP1b is essential for decapping complex interactions with protein degradation and translational machinery. DCP1a and DCP1b impact the turnover of distinct mRNAs. The observation that different ontological groups of mRNA molecules are regulated by DCP1a and DCP1b, along with their non-redundant roles in decapping complex integrity, provides the first evidence that these paralogs have qualitatively distinct functions.
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- 2024
18. CleanCap M6 inhibits decapping of exogenously delivered IVT mRNA.
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Mandell ZF, Ujita A, Henderson J, Truong A, Vo C, Rezvani F, Abolhassani N, Lebedev A, Xu C, Koukhareva I, Ramos S, Broderick K, Hudson B, and Coller J
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Co-transcriptional capping allows exogenous mRNAs to yield robust protein expression. Identifying additional mRNA modifications that further boost protein output will be crucial for developing more efficacious mRNA therapies. Using in vitro approaches, we found that the co-transcriptional cap analog
m7 G3'OMe pppm6 A2'OMe pG, CleanCap M6, resists enzymatic decapping and that this decreased susceptibility to decapping correlated with substantially increased protein expression in vivo compared to mRNAs capped using existing industry standards., Competing Interests: J.C. was funded in part by a grant from Maravai LifeSciences, parent company of TriLink BioTechnologies. Authors noted as being affiliated with TriLink BioTechnologies are employees of TriLink BioTechnologies, a subsidiary of Maravai LifeSciences Holdings, and are shareholders thereof. These authors may also be inventors on patents related to this work, a list of which is available at https://www.trilinkbiotech.com/legal-notices., (© 2025 The Author(s).)- Published
- 2025
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19. A unique surface on Pat1 C-terminal domain directly interacts with Dcp2 decapping enzyme and Xrn1 5′–3′ mRNA exonuclease in yeast
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Charenton, Clément, Gaudon-Plesse, Claudine, Fourati, Zaineb, Taverniti, Valerio, Back, Régis, Kolesnikova, Olga, Séraphin, Bertrand, and Graille, Marc
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- 2017
20. Emerging role and clinical implication of mRNA scavenger decapping enzyme in colorectal cancer
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Dimberg, Jan, Shamoun, Levar, Johansson, Gustaf, Landerholm, Kalle, Wagsater, Dick, Dimberg, Jan, Shamoun, Levar, Johansson, Gustaf, Landerholm, Kalle, and Wagsater, Dick
- Abstract
Background: Turnover of RNA is a regulated process that in part controls gene expression. This process is partly controlled by the scavenger decapping enzyme (DcpS). This study aimed to investigate the expression of DcpS in colorectal cancer (CRC) tissue, to evaluate its prognostic significance in patients with CRC and to investigate potentially targeted genes by DcpS.Methods: Immunohistochemical analysis was used to determine localization of DcpS in normal and CRC tissue, western blot analysis for quantification of protein expression and qPCR for mRNA expression in normal and CRC tissue and expression in cell lines after silencing using siRNA. Gene array analysis was used to study regulation of genes after silencing of DcpS. Proliferation was studied using BRDU.Results: DcpS expression was localized to the epithelial cells of both control and cancer tissue. Tumor and paired control tissue samples from 100 patients who underwent surgical resection for primary colorectal adenocarcinomas were utilized. mRNA and protein of DcpS was significantly up-regulated in the patients with CRC and the mRNA level was higher in rectal cancer tissue compared to colon cancer tissue (p < 0.05). Lowest tertile levels of DcpS mRNA in cancer tissue was associated with a decreased cancer-specific survival rate with a hazard ratio (HR) of 4.7 (95% CI=1.02-12.3), independent of disease stage. The low level of DcpS mRNA was a predictor of poorer survival in patients with rectal and disseminated cancer and in patients receiving adjuvant treatment (p < 0.05). After silencing DcpS in Caco-2 cancer cells, altered expression of several genes associated with RNA, cell cycle regulation, alternative splicing and microRNA was observed and resulted in 23% increase in proliferation.Conclusions: These results indicate that DcpS has potential as a prognostic factor for CRC but further studies in a broader cohort are warranted to evaluate the significance of the findings in the clinic., Funding Agencies|Division of Medical Diagnostics of Region Jonkoping County Sweden [Futurum-970572]; Swedish Research Council [2019-01673]
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- 2024
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21. T4 phage RNA is NAD-capped and alters the NAD-cap epitranscriptome of Escherichia coli during infection through a phage-encoded decapping enzyme
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Wolfram-Schauerte, Maik, Moskalchuk, Anastassiya, Pozhydaieva, Nadiia, Ramı́rez Rojas, Adán Andrés, Schindler, Daniel, Kaiser, Stefanie, Paczia, Nicole, Höfer, Katharina, Wolfram-Schauerte, Maik, Moskalchuk, Anastassiya, Pozhydaieva, Nadiia, Ramı́rez Rojas, Adán Andrés, Schindler, Daniel, Kaiser, Stefanie, Paczia, Nicole, and Höfer, Katharina
- Abstract
Nicotinamide adenine dinucleotide (NAD) serves as a cap-like structure on cellular RNAs (NAD-RNAs) in all domains of life including the bacterium Escherichia coli. NAD also acts as a key molecule in phage-host interactions, where bacterial immune systems deplete NAD to abort phage infection. Nevertheless, NAD-RNAs have not yet been identified during phage infections of bacteria and the mechanisms of their synthesis and degradation are unknown in this context. The T4 phage that specifically infects E. coli presents an important model to study phage infections, but a systematic analysis of the presence and dynamics of NAD-RNAs during T4 phage infection is lacking. Here, we investigate the presence of NAD-RNAs during T4 phage infection in a dual manner. By applying time-resolved NAD captureSeq, we identify NAD-capped host and phage transcripts and their dynamic regulation during phage infection. We provide evidence that NAD-RNAs are – as reported earlier – generated by the host RNA polymerase by initiating transcription with NAD at canonical transcription start sites. In addition, we characterize NudE.1 – a T4 phage-encoded Nudix hydrolase – as the first phage-encoded NAD-RNA decapping enzyme. T4 phages carrying inactive NudE.1 display a delayed lysis phenotype. This study investigates for the first time the dual epitranscriptome of a phage and its host, thereby introducing epitranscriptomics as an important field of phage research.
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- 2024
22. InsP₇ is a small-molecule regulator of NUDT3-mediated mRNA decapping and processing-body dynamics
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Sahu, Soumyadip, Wang, Zhenzhen, Jiao, Xinfu, Gu, Chunfang, Jork, Nikolaus, Wittwer, Christopher, Li, Xingyao, Hostachy, Sarah, Fiedler, Dorothea, Wang, Huanchen, Jessen, Henning J., Kiledjian, Megerditch, and Shears, Stephen B.
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- 2020
23. Molecular basis of the selective processing of short mRNA substrates by the DcpS mRNA decapping enzyme
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Fuchs, Anna-Lisa, Wurm, Jan Philip, Neu, Ancilla, and Sprangers, Remco
- Published
- 2020
24. Thermotolerance through trade-off: Decapping WUSCHEL mRNA in plant stem cells.
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Debnath P and Olas JJ
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- 2025
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25. Chikungunya virus nonstructural protein 1 is a versatile RNA capping and decapping enzyme
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Law, Michelle Cheok Yien, Zhang, Kuo, Tan, Yaw Bia, Nguyen, Trinh Mai, and Luo, Dahai
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- 2023
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26. Mechanisms of action of cytoplasmic miRNAs. Part 4. Recruitment of the DCP1-DCP2 decapping complex. Mechanisms of final mRNA degradation
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A.E. Abaturov and V.L. Babуch
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microrna ,mirna ,mir ,post-transcriptional silencing ,recruitment of the dcp1-dcp2 decapping complex ,мicrorna-mediated degradation of mrna ,review ,Pediatrics ,RJ1-570 - Abstract
This scientific review deals with the mechanisms of action of cytoplasmic microRNAs, namely post-transcriptional silencing: recruitment of the DCP1-DCP2 decapping complex and disruption of the interaction of mRNA with ribosomes. To write the article, information was searched using Scopus, Web of Science, MedLine, PubMed, Google Scholar, EMBASE, Global Health, The Cochrane Library, CyberLeninka databases. The authors indicate that the key process that determines both mRNA stability and expression efficiency is the removal of the 5’-terminal cap. Decapping of mRNA is controlled by several direct and indirect regulators. The DCP1-DCP2 complex can be recruited directly to mRNA and indirectly with the help of several decapping enhancers: PAT1 directly interacts with DCP1 and the decapping stimulator; EDC, DDX6. It is known that the protein DCP2 (Nudt20) is a representative of the conserved subfamily of Nudix hydrolases, which catalyze the hydrolysis of small nucleotide substrates. It is presented that the DCP1 protein is a small molecule that contains the EVH1 (enabled/vasodilator-stimulated phosphoprotein homology 1) domain, which usually acts as a protein-protein interaction module, and a C-terminal trimerization domain. It is known that the DCP1-DCP2 complex exists in an open and closed conformation, with the closed conformation having catalytic activity. DCP2 protein and its enhancer and cofactor partners accumulate in P-bodies. The authors indicate that in P-bodies, 5’-monophosphorylated mRNA is finally cleaved under the action of 5’-3’-exoribonuclease XRN1. XRN exoribonucleases are vital enzymes whose gene deletion is accompanied by intraembryonic lethality against the background of various abnormalities in the development of organs and systems. Thus, recruitment of the DCP1-DCP2 decapping complex and disruption of the interaction of mRNA with ribosomes in the cytoplasm of the cell are mechanisms of post-transcriptional silencing. The stability of the mRNA and the efficiency of expression determines the removal of the 5’ end cap. Termination of translation is caused by mRNA. MicroRNA-mediated degradation of this mRNA can be carried out both in the 3’-5’ and 5’-3’ directions of the molecule.
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- 2022
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27. Vaccinia virus D10 has broad decapping activity that is regulated by mRNA splicing
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Ly, Michael, Burgess, Hannah M, Shah, Sahil B, Mohr, Ian, and Glaunsinger, Britt A
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Biological Sciences ,Bioinformatics and Computational Biology ,Biomedical and Clinical Sciences ,Infectious Diseases ,Emerging Infectious Diseases ,Genetics ,Rare Diseases ,Small Pox ,2.2 Factors relating to the physical environment ,2.1 Biological and endogenous factors ,Infection ,Endoribonucleases ,Humans ,Poxviridae ,RNA Caps ,RNA ,Messenger ,Vaccinia virus ,Viral Proteins ,Microbiology ,Immunology ,Medical Microbiology ,Virology ,Medical microbiology - Abstract
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-containing genes provides an unanticipated mechanism for the virus to disproportionately deplete host transcripts and remodel the infected cell transcriptome.
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- 2022
28. The messenger RNA decapping and recapping pathway in Trypanosoma
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Ignatochkina, Anna V., Takagi, Yuko, Liu, Yancheng, Nagata, Kyosuke, and Ho, C. Kiong
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- 2015
29. Decapping factor Dcp2 controls mRNA abundance and translation to adjust metabolism and filamentation to nutrient availability
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Anil Kumar Vijjamarri, Xiao Niu, Matthew D Vandermeulen, Chisom Onu, Fan Zhang, Hongfang Qiu, Neha Gupta, Swati Gaikwad, Miriam L Greenberg, Paul J Cullen, Zhenguo Lin, and Alan G Hinnebusch
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decapping ,degradation ,translation ,Dcp2 ,Dhh1 ,NMD ,Medicine ,Science ,Biology (General) ,QH301-705.5 - Abstract
Degradation of most yeast mRNAs involves decapping by Dcp1/Dcp2. DEAD-box protein Dhh1 has been implicated as an activator of decapping, in coupling codon non-optimality to enhanced degradation, and as a translational repressor, but its functions in cells are incompletely understood. RNA-Seq analyses coupled with CAGE sequencing of all capped mRNAs revealed increased abundance of hundreds of mRNAs in dcp2Δ cells that appears to result directly from impaired decapping rather than elevated transcription. Interestingly, only a subset of mRNAs requires Dhh1 for targeting by Dcp2, and also generally requires the other decapping activators Pat1, Edc3, or Scd6; whereas most of the remaining transcripts utilize nonsense-mediated mRNA decay factors for Dcp2-mediated turnover. Neither inefficient translation initiation nor stalled elongation appears to be a major driver of Dhh1-enhanced mRNA degradation. Surprisingly, ribosome profiling revealed that dcp2Δ confers widespread changes in relative translational efficiencies (TEs) that generally favor well-translated mRNAs. Because ribosome biogenesis is reduced while capped mRNA abundance is increased by dcp2Δ, we propose that an increased ratio of mRNA to ribosomes increases competition among mRNAs for limiting ribosomes to favor efficiently translated mRNAs in dcp2Δ cells. Interestingly, genes involved in respiration or utilization of alternative carbon or nitrogen sources are upregulated, and both mitochondrial function and cell filamentation are elevated in dcp2Δ cells, suggesting that decapping sculpts gene expression post-transcriptionally to fine-tune metabolic pathways and morphological transitions according to nutrient availability.
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- 2023
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30. Identification of NAD-RNA species and ADPR-RNA decapping in Archaea
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Gomes-Filho, José Vicente, Breuer, Ruth, Morales-Filloy, Hector Gabriel, Pozhydaieva, Nadiia, Borst, Andreas, Paczia, Nicole, Soppa, Jörg, Höfer, Katharina, Jäschke, Andres, and Randau, Lennart
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- 2023
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31. Alterations in cellular RNA decapping dynamics affect tomato spotted wilt virus cap snatching and infection in Arabidopsis
- Author
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Ma, Xiaofang, Zhou, Yijun, and Moffett, Peter
- Published
- 2019
32. mRNA Decapping Factors LSM1 and PAT Paralogs Are Involved in Turnip Mosaic Virus Viral Infection
- Author
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Zhangli Zuo, Milena Roux, Eleazar Rodriguez, and Morten Petersen
- Subjects
LSM1 ,mRNA decapping ,PAT paralogs ,Turnip mosaic virus ,Microbiology ,QR1-502 ,Botany ,QK1-989 - Abstract
Turnip mosaic virus is a devastating potyvirus infecting many economically important brassica crops. In response to this, the plant host engages its RNA silencing machinery, involving AGO proteins, as a prominent strategy to restrain turnip mosaic virus (TuMV) infection. It has also been shown that the mRNA decay components DCP2 and VCS partake in viral infection suppression. Here, we report that the mRNA decapping components LSM1, PAT1, PATH1, and PATH2 are essential for TuMV infection. More specifically, lsm1a/lsm1b double mutants and pat1/path1/path2 triple mutants in summ2 background exhibit resistance to TuMV. Concurrently, we observed that TuMV interferes with the decapping function of LSM1 and PAT proteins as the mRNA-decay target genes UGT87A2 and ASL9 accumulate during TuMV infection. Moreover, as TuMV coat protein can be specifically found in complexes with PAT proteins but not LSM1, this suggests that TuMV “hijacks” decapping components via PAT proteins to support viral infection.[Graphic: see text] Copyright © 2022 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.
- Published
- 2022
- Full Text
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33. Aptamer-based assay for high-throughput substrate profiling of RNA decapping enzymes.
- Author
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Grab K, Fido M, Spiewla T, Warminski M, Jemielity J, and Kowalska J
- Subjects
- Substrate Specificity, RNA Caps metabolism, RNA Caps chemistry, Kinetics, Endoribonucleases metabolism, Endoribonucleases chemistry, Aptamers, Nucleotide chemistry, High-Throughput Screening Assays methods
- Abstract
Recent years have led to the identification of a number of enzymes responsible for RNA decapping. This has provided a basis for further research to identify their role, dependency and substrate specificity. However, the multiplicity of these enzymes and the complexity of their functions require advanced tools to study them. Here, we report a high-throughput fluorescence intensity assay based on RNA aptamers designed as substrates for decapping enzymes. Using a library of differently capped RNA probes we generated a decapping susceptibility heat map, which confirms previously reported substrate specificities of seven tested hydrolases and uncovers novel. We have also demonstrated the utility of our assay for evaluating inhibitors of viral decapping enzymes and performed kinetic studies of the decapping process. The assay may accelerate the characterization of new decapping enzymes, enable high-throughput screening of inhibitors and facilitate the development of molecular tools for a better understanding of RNA degradation pathways., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.)
- Published
- 2024
- Full Text
- View/download PDF
34. Reply to Graille, "InsPection of electron density maps supports wrongly modeled hexakisphosphate (InsP6) bound to African swine fever mRNA-decapping enzyme g5Rp".
- Author
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Yang Y, Li X, and Su D
- Published
- 2024
- Full Text
- View/download PDF
35. Biomolecular condensates amplify mRNA decapping by biasing enzyme conformation
- Author
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Tibble, Ryan W, Depaix, Anaïs, Kowalska, Joanna, Jemielity, Jacek, and Gross, John D
- Subjects
Generic health relevance ,Binding Sites ,Cloning ,Molecular ,Cytoplasmic Granules ,Escherichia coli ,Fluorescent Dyes ,Gene Expression ,Genetic Vectors ,Models ,Molecular ,Protein Binding ,Protein Conformation ,alpha-Helical ,Protein Conformation ,beta-Strand ,Protein Interaction Domains and Motifs ,RNA Caps ,RNA Stability ,Recombinant Proteins ,Schizosaccharomyces ,Schizosaccharomyces pombe Proteins ,Staining and Labeling ,Substrate Specificity ,Medicinal and Biomolecular Chemistry ,Biochemistry and Cell Biology ,Biochemistry & Molecular Biology - Abstract
Cells organize biochemical processes into biological condensates. P-bodies are cytoplasmic condensates that are enriched in enzymes important for mRNA degradation and have been identified as sites of both storage and decay. How these opposing outcomes can be achieved in condensates remains unresolved. mRNA decapping immediately precedes degradation, and the Dcp1/Dcp2 decapping complex is enriched in P-bodies. Here, we show that Dcp1/Dcp2 activity is modulated in condensates and depends on the interactions promoting phase separation. We find that Dcp1/Dcp2 phase separation stabilizes an inactive conformation in Dcp2 to inhibit decapping. The activator Edc3 causes a conformational change in Dcp2 and rewires the protein-protein interactions to stimulate decapping in condensates. Disruption of the inactive conformation dysregulates decapping in condensates. Our results indicate that the regulation of enzymatic activity in condensates relies on a coupling across length scales ranging from microns to ångstroms. We propose that this regulatory mechanism may control the functional state of P-bodies and related phase-separated compartments.
- Published
- 2021
36. The mRNA decapping machinery targets LBD3/ASL9 to mediate apical hook and lateral root development
- Author
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Zuo, Zhangli, Roux, Milena E., Chevalier, Jonathan R., Dagdas, Yasin F., Yamashino, Takafumi, Højgaard, Søren D., Knight, Emilie, Østergaard, Lars, Rodriguez, Eleazar, Petersen, Morten, Zuo, Zhangli, Roux, Milena E., Chevalier, Jonathan R., Dagdas, Yasin F., Yamashino, Takafumi, Højgaard, Søren D., Knight, Emilie, Østergaard, Lars, Rodriguez, Eleazar, and Petersen, Morten
- Abstract
Multicellular organisms perceive and transduce multiple cues to optimize development. Key transcription factors drive developmental changes, but RNA processing also contributes to tissue development. Here, we report that multiple decapping deficient mutants share developmental defects in apical hook, primary and lateral root growth. More specifically, LATERAL ORGAN BOUNDARIES DOMAIN 3 (LBD3)/ASYMMETRIC LEAVES 2-LIKE 9 (ASL9) transcripts accumulate in decapping deficient plants and can be found in complexes with decapping components. Accumulation of ASL9 inhibits apical hook and lateral root formation. Interestingly, exogenous auxin application restores lateral roots formation in both ASL9 over-expressors and mRNA decay–deficient mutants. Likewise, mutations in the cytokinin transcription factors type-B ARABIDOPSIS RESPONSE REGULATORS (B-ARRs) ARR10 and ARR12 restore the developmental defects caused by over-accumulation of capped ASL9 transcript upon ASL9 overexpression. Most importantly, loss-of-function of asl9 partially restores apical hook and lateral root formation in both dcp5-1 and pat triple decapping deficient mutants. Thus, the mRNA decay machinery directly targets ASL9 transcripts for decay, possibly to interfere with cytokinin/auxin responses, during development., Multicellular organisms perceive and transduce multiple cues to optimize development. Key transcription factors drive developmental changes, but RNA processing also contributes to tissue development. Here, we report that multiple decapping deficient mutants share developmental defects in apical hook, primary and lateral root growth. More specifically, LATERAL ORGAN BOUNDARIES DOMAIN 3 (LBD3)/ASYMMETRIC LEAVES 2-LIKE 9 (ASL9) transcripts accumulate in decapping deficient plants and can be found in complexes with decapping components. Accumulation of ASL9 inhibits apical hook and lateral root formation. Interestingly, exogenous auxin application restores lateral roots formation in both ASL9 over-expressors and mRNA decay-deficient mutants. Likewise, mutations in the cytokinin transcription factors type-B ARABIDOPSIS RESPONSE REGULATORS (B-ARRs) ARR10 and ARR12 restore the developmental defects caused by over-accumulation of capped ASL9 transcript upon ASL9 overexpression. Most importantly, loss-of-function of asl9 partially restores apical hook and lateral root formation in both dcp5-1 and pat triple decapping deficient mutants. Thus, the mRNA decay machinery directly targets ASL9 transcripts for decay, possibly to interfere with cytokinin/auxin responses, during development.
- Published
- 2023
37. Oomycete Nudix effectors display WY-Nudix conformation and mRNA decapping activity.
- Author
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Guo B, Hu Q, Wang B, Yao D, Wang H, Kong G, Han C, Dong S, Liu F, Xing W, and Wang Y
- Subjects
- RNA, Messenger metabolism, RNA, Messenger genetics, RNA Caps metabolism, Oomycetes physiology
- Abstract
Oomycete Nudix effectors have characteristics of independent evolution, but adopt a conserved WY-Nudix conformation. Furthermore, multiple oomycete Nudix effectors exhibit mRNA decapping activity., (© 2024 Institute of Botany, Chinese Academy of Sciences.)
- Published
- 2024
- Full Text
- View/download PDF
38. Recent insights into noncanonical 5′ capping and decapping of RNA
- Author
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Doamekpor, Selom K., Sharma, Sunny, Kiledjian, Megerditch, and Tong, Liang
- Published
- 2022
- Full Text
- View/download PDF
39. Application of a Schizosaccharomyces pombe Edc1-fused Dcp1–Dcp2 decapping enzyme for transcription start site mapping
- Author
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Paquette, David R, Mugridge, Jeffrey S, Weinberg, David E, and Gross, John D
- Subjects
Biological Sciences ,Bioinformatics and Computational Biology ,Genetics ,5' Untranslated Regions ,Cloning ,Molecular ,Escherichia coli ,Protein Engineering ,RNA Caps ,RNA-Binding Proteins ,Recombinant Fusion Proteins ,Schizosaccharomyces pombe Proteins ,Transcription Initiation Site ,mRNA caps ,mapping mRNA 5' ends ,decapping enzymes ,RppH ,Dcp2 ,transcript leaders ,mapping mRNA 5′ ends ,Biochemistry and Cell Biology ,Developmental Biology ,Biochemistry and cell biology - Abstract
Changes in the 5' leader of an mRNA can have profound effects on its translational efficiency with little effect on abundance. Sequencing-based methods to accurately map the 5' leader by identifying the first transcribed nucleotide rely on enzymatic removal of the 5' eukaryotic cap structure by tobacco acid pyrophosphatase (TAP). However, commercial TAP production has been problematic and has now been discontinued. RppH, a bacterial enzyme that can also cleave the 5' cap, and Cap-Clip, a plant-derived enzyme, have been marketed as TAP replacements. We have engineered a Schizosaccharomyces pombe Edc1-fused Dcp1-Dcp2 decapping enzyme that functions as a superior TAP replacement. It can be purified from E. coli overexpression in high yields using standard biochemical methods. This constitutively active enzyme is four orders of magnitude more catalytically efficient than RppH at 5' cap removal, compares favorably to Cap-Clip, and the 5' monophosphorylated RNA product is suitable for standard RNA cloning methods. This engineered enzyme is a better replacement for TAP treatment than the current marketed use of RppH and can be produced cost-effectively in a general laboratory setting, unlike Cap-Clip.
- Published
- 2018
40. Potential epigenetic regulation of RNA 5’-terminal NAD decapping associated with cellular energy status of postharvest Fragaria × ananassa in response to Botrytis cinerea invasion
- Author
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Ma, Quan, Xu, Yanqun, Li, Dong, Wu, Xinwei, Zhang, Xiaochen, Chen, Yanpei, Li, Li, and Luo, Zisheng
- Published
- 2022
- Full Text
- View/download PDF
41. Eukaryotic mRNA decapping factors: molecular mechanisms and activity.
- Author
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He, Feng and Jacobson, Allan
- Subjects
RNA regulation ,GENE silencing ,EUKARYOTIC cells ,MESSENGER RNA ,SACCHAROMYCES cerevisiae - Abstract
Decapping is the enzymatic removal of 5′ cap structures from mRNAs in eukaryotic cells. Cap structures normally enhance mRNA translation and stability, and their excision commits an mRNA to complete 5′–3′ exoribonucleolytic digestion and generally ends the physical and functional cellular presence of the mRNA. Decapping plays a pivotal role in eukaryotic cytoplasmic mRNA turnover and is a critical and highly regulated event in multiple 5′–3′ mRNA decay pathways, including general 5′–3′ decay, nonsense‐mediated mRNA decay (NMD), AU‐rich element‐mediated mRNA decay, microRNA‐mediated gene silencing, and targeted transcript‐specific mRNA decay. In the yeast Saccharomyces cerevisiae, mRNA decapping is carried out by a single Dcp1‐Dcp2 decapping enzyme in concert with the accessory activities of specific regulators commonly known as decapping activators or enhancers. These regulatory proteins include the general decapping activators Edc1, 2, and 3, Dhh1, Scd6, Pat1, and the Lsm1‐7 complex, as well as the NMD‐specific factors, Upf1, 2, and 3. Here, we focus on in vivo mRNA decapping regulation in yeast. We summarize recently uncovered molecular mechanisms that control selective targeting of the yeast decapping enzyme and discuss new roles for specific decapping activators in controlling decapping enzyme targeting, assembly of target‐specific decapping complexes, and the monitoring of mRNA translation. Further, we discuss the kinetic contribution of mRNA decapping for overall decay of different substrate mRNAs and highlight experimental evidence pointing to the functional coordination and physical coupling between events in mRNA deadenylation, decapping, and 5′–3′ exoribonucleolytic decay. [ABSTRACT FROM AUTHOR]
- Published
- 2023
- Full Text
- View/download PDF
42. Is mRNA decapping by ApaH like phosphatases present in eukaryotes beyond the Kinetoplastida?
- Author
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Castañeda Londoño, Paula Andrea, Banholzer, Nicole, Bannermann, Bridget, and Kramer, Susanne
- Published
- 2021
- Full Text
- View/download PDF
43. PAT mRNA decapping factors are required for proper development in Arabidopsis.
- Author
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Zuo Z, Roux ME, Dagdas YF, Rodriguez E, and Petersen M
- Subjects
- Gene Expression Regulation, Plant, Mutation, RNA Stability, Arabidopsis genetics, Arabidopsis growth & development, Arabidopsis metabolism, Arabidopsis Proteins genetics, Arabidopsis Proteins metabolism, RNA, Messenger genetics, RNA, Messenger metabolism
- Abstract
Evolutionarily conserved protein associated with topoisomerase II (PAT1) proteins activate mRNA decay through binding mRNA and recruiting decapping factors to optimize posttranscriptional reprogramming. Here, we generated multiple mutants of pat1, pat1 homolog 1 (path1), and pat1 homolog 2 (path2) and discovered that pat triple mutants exhibit extremely stunted growth and all mutants with pat1 exhibit leaf serration while mutants with pat1 and path1 display short petioles. All three PATs can be found localized to processing bodies and all PATs can target ASYMMETRIC LEAVES 2-LIKE 9 transcripts for decay to finely regulate apical hook and lateral root development. In conclusion, PATs exhibit both specific and redundant functions during different plant growth stages and our observations underpin the selective regulation of the mRNA decay machinery for proper development., (© 2024 The Authors. FEBS Letters published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)
- Published
- 2024
- Full Text
- View/download PDF
44. Two-headed tetraphosphate cap analogs are inhibitors of the Dcp1/2 RNA decapping complex
- Author
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Ziemniak, Marcin, Mugridge, Jeffrey S, Kowalska, Joanna, Rhoads, Robert E, Gross, John D, and Jemielity, Jacek
- Subjects
Biochemistry and Cell Biology ,Biological Sciences ,Genetics ,Drug Evaluation ,Preclinical ,RNA Cap Analogs ,RNA Cleavage ,RNA ,Messenger ,Schizosaccharomyces ,Schizosaccharomyces pombe Proteins ,Dcp1/Dcp2 ,RNA metabolism ,decapping enzymes ,inhibitors ,mRNA cap ,Developmental Biology ,Biochemistry and cell biology - Abstract
Dcp1/2 is the major eukaryotic RNA decapping complex, comprised of the enzyme Dcp2 and activator Dcp1, which removes the 5' m(7)G cap from mRNA, committing the transcript to degradation. Dcp1/2 activity is crucial for RNA quality control and turnover, and deregulation of these processes may lead to disease development. The molecular details of Dcp1/2 catalysis remain elusive, in part because both cap substrate (m(7)GpppN) and m(7)GDP product are bound by Dcp1/2 with weak (mM) affinity. In order to find inhibitors to use in elucidating the catalytic mechanism of Dcp2, we screened a small library of synthetic m(7)G nucleotides (cap analogs) bearing modifications in the oligophosphate chain. One of the most potent cap analogs, m(7)GpSpppSm(7)G, inhibited Dcp1/2 20 times more efficiently than m(7)GpppN or m(7)GDP. NMR experiments revealed that the compound interacts with specific surfaces of both regulatory and catalytic domains of Dcp2 with submillimolar affinities. Kinetics analysis revealed that m(7)GpSpppSm(7)G is a mixed inhibitor that competes for the Dcp2 active site with micromolar affinity. m(7)GpSpppSm(7)G-capped RNA undergoes rapid decapping, suggesting that the compound may act as a tightly bound cap mimic. Our identification of the first small molecule inhibitor of Dcp2 should be instrumental in future studies aimed at understanding the structural basis of RNA decapping and may provide insight toward the development of novel therapeutically relevant decapping inhibitors.
- Published
- 2016
45. Enhancer of mRNA Decapping protein 4 (EDC4) interacts with replication protein a (RPA) and contributes to Cisplatin resistance in cervical Cancer by alleviating DNA damage
- Author
-
Xiaoling Wu, Youwen Zhong, Qing Chen, Xin Zhang, and Hua Zhang
- Subjects
Cervical Cancer ,Cisplatin resistance ,Enhancer of mRNA decapping protein 4 (EDC4) ,Replication protein a (RPA) ,DNA damage ,Genetics ,QH426-470 - Abstract
Abstract Background Cervical cancer (CC) is the third most common gynecological malignancy around the world. Cisplatin is an effective drug, but cisplatin resistance is a vital factor limiting the clinical usage of cisplatin. Enhancer of mRNA decapping protein 4 (EDC4) is a known regulator of mRNA decapping, which was related with genome stability and sensitivity of drugs. This research was to investigate the mechanism of EDC4 on cisplatin resistance in CC. Two human cervical cancer cell lines, HeLa and SiHa, were used to investigate the role of EDC4 on cisplatin resistance in vitro. The knockdown or overexpression of EDC4 or replication protein A (RPA) in HeLa or SiHa cells was performed by transfection. Cell viability was analyzed by MTT assay. The growth of cancer cells was evaluated by colony formation assay. DNA damage was measured by γH2AX (a sensitive DNA damage response marker) immunofluorescent staining. The binding of EDC4 and RPA was analyzed by immunoprecipitation. Results EDC4 knockdown in cervical cancer cells (HeLa and SiHa) enhanced cisplatin sensitivity and cisplatin induced cell growth inhibition and DNA damage. EDC4 overexpression reduced DNA damage caused by cisplatin and enhanced cell growth of cervical cancer cells. EDC4 could interact with RPA and promote RPA phosphorylation. RPA knockdown reversed the inhibitory effect of EDC4 on cisplatin-induced DNA damage. Conclusion The present results indicated that EDC4 is responsible for the cisplatin resistance partly through interacting with RPA in cervical cancer by alleviating DNA damage. This study indicated that EDC4 or RPA may be novel targets to combat chemotherapy resistance in cervical cancer. Graphical abstract
- Published
- 2020
- Full Text
- View/download PDF
46. A Poxvirus Decapping Enzyme Colocalizes with Mitochondria To Regulate RNA Metabolism and Translation and Promote Viral Replication
- Author
-
Shuai Cao, Joshua A. Molina, Fernando Cantu, Candy Hernandez, and Zhilong Yang
- Subjects
decapping enzyme ,poxvirus ,vaccinia virus ,mitochondria ,translation ,RNA decay ,Microbiology ,QR1-502 - Abstract
ABSTRACT Decapping enzymes remove the 5′ cap of eukaryotic mRNA, leading to accelerated RNA decay. They are critical in regulating RNA homeostasis and play essential roles in many cellular and life processes. They are encoded in many organisms and viruses, including vaccinia virus, which was used as the vaccine to eradicate smallpox. Vaccinia virus encodes two decapping enzymes, D9 and D10, that are necessary for efficient viral replication and pathogenesis. However, the underlying molecular mechanisms regulating vaccinia decapping enzymes’ functions are still largely elusive. Here, we demonstrated that vaccinia D10 almost exclusively colocalized with mitochondria. As mitochondria are highly mobile cellular organelles, colocalization of D10 with mitochondria can concentrate D10 locally and mobilize it to efficiently decap mRNAs. Mitochondria were barely observed in “viral factories,” where viral transcripts are produced, suggesting that mitochondrial colocalization provides a spatial mechanism to preferentially decap cellular mRNAs over viral mRNAs. We identified three amino acids at the N terminus of D10 that are required for D10’s mitochondrial colocalization. Loss of mitochondrial colocalization significantly impaired viral replication, reduced D10’s ability to remove the RNA 5′ cap during infection, and diminished D10’s gene expression shutoff and mRNA translation promotion abilities. IMPORTANCE Decapping enzymes comprise many members from various organisms, ranging from plants, animals, and viruses. The mechanisms regulating their functions vary and are still largely unknown. Our study provides evidence that a vaccinia virus-encoded decapping enzyme, D10, colocalizes with mitochondria. Loss of mitochondrial colocalization significantly impairs viral replication, D10’s gene expression shutoff, and mRNA translation promotion ability. Overall, our results suggest that mitochondrial colocalization is a spatial mechanism to concentrate D10 locally and mobilize it to efficiently and preferentially target cellular mRNAs for decapping and promote viral mRNA translation. Our results have broad impacts for understanding the functions and regulatory mechanisms of decapping enzymes.
- Published
- 2022
- Full Text
- View/download PDF
47. mRNAs biotinylated within the 5ʹ cap and protected against decapping : new tools to capture RNA–protein complexes
- Author
-
Bednarek, Sylwia, Madan, Vanesa, Sikorski, Pawel J., Bartenschlager, Ralf, Kowalska, Joanna, and Jemielity, Jacek
- Published
- 2018
48. Interplay between bacterial 5′-NAD-RNA decapping hydrolase NudC and DEAD-box RNA helicase CsdA in stress responses
- Author
-
Milda Mickutė, Renatas Krasauskas, Kotryna Kvederavičiūtė, Gytė Tupikaitė, Aleksandr Osipenko, Algirdas Kaupinis, Monika Jazdauskaitė, Raminta Mineikaitė, Mindaugas Valius, Viktoras Masevičius, and Giedrius Vilkaitis
- Subjects
RNA modification ,RNA hydrolase ,DEAD-box helicase ,RNA cap ,NAD-RNA ,Microbiology ,QR1-502 - Abstract
ABSTRACT Both prokaryotic and eukaryotic RNAs can be 5′-capped by the metabolite nicotinamide adenine dinucleotide (NAD). Nudix hydrolases, such as bacterial NudC, specifically remove NAD-caps; however, the molecular and cellular functions of these epitranscriptomic modulators remain elusive. Here, we discuss the roles of NudC under stress conditions and the effects of extracellular cues on the NAD epitranscriptome. Our proteome-wide analysis detected the proteins associated with the RNA degradosome as well as ribosomes and stress-responsive proteins in a NudC interactome. Moreover, we confirmed the physical association of NudC with the cold shock DEAD-box RNA helicase CsdA and the RNA chaperone Hfq. Interestingly, knocking out csdA similar to ∆nudC leads to an increased number of identified 5′-NAD-RNA species compared to wild type, exposing CsdA as a new player in a potentially unexplored layer of NAD-epitranscriptomic landscape. Mass spectrometry analysis also revealed the drastic up-regulation of 5′-NAD-RNA in response to cold. Furthermore, the inactivation of NudC in bacteria changes the levels of sRNA and protein-coding transcripts associated with bacterial chemotaxis and flagellar assembly pathways. We experimentally demonstrate that the decapping hydrolase NudC suppresses the flagellar movement, while CsdA stimulates it. Thus, the interplay between NudC and CsdA regulates bacterial mobility and coordinates stress-avoidance behavior. IMPORTANCE Non-canonical 5′-caps removing RNA hydrolase NudC, along with stress-responsive RNA helicase CsdA, is crucial for 5′-NAD-RNA decapping and bacterial movement.
- Published
- 2023
- Full Text
- View/download PDF
49. Multi-transcriptomics identifies targets of the endoribonuclease DNE1 and highlights its coordination with decapping.
- Author
-
Pouclet, Aude, Pflieger, David, Merret, Rémy, Carpentier, Marie-Christine, Schiaffini, Marlene, Zuber, Hélène, Gagliardi, Dominique, and Garcia, Damien
- Subjects
- *
SMALL interfering RNA , *TRANSCRIPTOMES , *RNA editing , *QUADRUPLEX nucleic acids , *NUCLEOTIDE sequencing - Abstract
Decapping is a crucial step in mRNA degradation in eucaryotes and requires the formation of a holoenzyme complex between the decapping enzyme DECAPPING 2 (DCP2) and the decapping enhancer DCP1. In Arabidopsis (Arabidopsis thaliana), DCP1-ASSOCIATED NYN ENDORIBONUCLEASE 1 (DNE1) is a direct protein partner of DCP1. The function of both DNE1 and decapping is necessary to maintain phyllotaxis, the regularity of organ emergence in the apex. In this study, we combined in vivo mRNA editing, RNA degradome sequencing, transcriptomics, and small RNA-omics to identify targets of DNE1 and study how DNE1 and DCP2 cooperate in controlling mRNA fate. Our data reveal that DNE1 mainly contacts and cleaves mRNAs in the coding sequence and has sequence cleavage preferences. DNE1 targets are also degraded through decapping, and both RNA degradation pathways influence the production of mRNA-derived small interfering RNAs. Finally, we detected mRNA features enriched in DNE1 targets including RNA G-quadruplexes and translated upstream open reading frames. Combining these four complementary high-throughput sequencing strategies greatly expands the range of DNE1 targets and allowed us to build a conceptual framework describing the influence of DNE1 and decapping on mRNA fate. These data will be crucial to unveil the specificity of DNE1 action and understand its importance for developmental patterning. Multi-transcriptomics reveals a large spectrum of mRNAs targeted by the endoribonuclease DNE1 and provides a conceptual framework to understand its importance for mRNA fate. [ABSTRACT FROM AUTHOR]
- Published
- 2024
- Full Text
- View/download PDF
50. Eukaryotic mRNA Decapping Activation
- Author
-
Elva Vidya and Thomas F. Duchaine
- Subjects
mRNA decapping and decay ,Dcp1/Dcp2 ,Edc1 ,Edc3 ,Edc4 ,PatL1 ,Genetics ,QH426-470 - Abstract
The 5′-terminal cap is a fundamental determinant of eukaryotic gene expression which facilitates cap-dependent translation and protects mRNAs from exonucleolytic degradation. Enzyme-directed hydrolysis of the cap (decapping) decisively affects mRNA expression and turnover, and is a heavily regulated event. Following the identification of the decapping holoenzyme (Dcp1/2) over two decades ago, numerous studies revealed the complexity of decapping regulation across species and cell types. A conserved set of Dcp1/2-associated proteins, implicated in decapping activation and molecular scaffolding, were identified through genetic and molecular interaction studies, and yet their exact mechanisms of action are only emerging. In this review, we discuss the prevailing models on the roles and assembly of decapping co-factors, with considerations of conservation across species and comparison across physiological contexts. We next discuss the functional convergences of decapping machineries with other RNA-protein complexes in cytoplasmic P bodies and compare current views on their impact on mRNA stability and translation. Lastly, we review the current models of decapping activation and highlight important gaps in our current understanding.
- Published
- 2022
- Full Text
- View/download PDF
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