Nucleic Acids Research最新文献_第8页

In silico nanoscope to study the interplay of genome organization and transcription regulation. 研究基因组组织和转录调控相互作用的硅纳米镜。

IF 16.6 2区生物学

Nucleic Acids Research Pub Date : 2025-03-20 DOI: 10.1093/nar/gkaf189

Soundhararajan Gopi, Giovanni B Brandani, Cheng Tan, Jaewoon Jung, Chenyang Gu, Azuki Mizutani, Hiroshi Ochiai, Yuji Sugita, Shoji Takada

{"title":"In silico nanoscope to study the interplay of genome organization and transcription regulation.","authors":"Soundhararajan Gopi, Giovanni B Brandani, Cheng Tan, Jaewoon Jung, Chenyang Gu, Azuki Mizutani, Hiroshi Ochiai, Yuji Sugita, Shoji Takada","doi":"10.1093/nar/gkaf189","DOIUrl":"10.1093/nar/gkaf189","url":null,"abstract":"In eukaryotic genomes, regulated access and communication between cis-regulatory elements (CREs) are necessary for enhancer-mediated transcription of genes. The molecular framework of the chromatin organization underlying such communication remains poorly understood. To better understand it, we develop a multiscale modeling pipeline to build near-atomistic models of the 200 kb Nanog gene locus in mouse embryonic stem cells comprising nucleosomes, transcription factors, co-activators, and RNA polymerase II-mediator complexes. By integrating diverse experimental data, including protein localization, genomic interaction frequencies, cryo-electron microscopy, and single-molecule fluorescence studies, our model offers novel insights into chromatin organization and its role in enhancer-promoter communication. The models equilibrated by high-performance molecular dynamics simulations span a scale of ∼350 nm, revealing an experimentally consistent local and global organization of chromatin and transcriptional machinery. Our models elucidate that the sequence-regulated chromatin accessibility facilitates the recruitment of transcription regulatory proteins exclusively at CREs, guided by the contrasting nucleosome organization compared to other regions. By constructing an experimentally consistent near-atomic model of chromatin in the cellular environment, our approach provides a robust framework for future studies on nuclear compartmentalization, chromatin organization, and transcription regulation.","PeriodicalId":19471,"journal":{"name":"Nucleic Acids Research","volume":"53 6","pages":""},"PeriodicalIF":16.6,"publicationDate":"2025-03-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11925733/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143670486","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Small nucleolar RNAs promote the restoration of muscle differentiation defects in cells from myotonic dystrophy type 1.

IF 16.6 2区生物学

Nucleic Acids Research Pub Date : 2025-03-20 DOI: 10.1093/nar/gkaf232

Baptiste Bogard, Hélène Bonnet, Ekaterina Boyarchuk, Gilles Tellier, Denis Furling, Vincent Mouly, Claire Francastel, Florent Hubé

{"title":"Small nucleolar RNAs promote the restoration of muscle differentiation defects in cells from myotonic dystrophy type 1.","authors":"Baptiste Bogard, Hélène Bonnet, Ekaterina Boyarchuk, Gilles Tellier, Denis Furling, Vincent Mouly, Claire Francastel, Florent Hubé","doi":"10.1093/nar/gkaf232","DOIUrl":"10.1093/nar/gkaf232","url":null,"abstract":"Recently, the repertoire of human small nucleolar noncoding RNAs (snoRNAs) and their potential functions has expanded with the discovery of new snoRNAs and messenger RNA (mRNA) targets, for which snoRNA-guided modifications may influence their stability, translatability, and splicing. We previously identified snoRNAs that are abundant in healthy human muscle progenitor cells. In this study, we demonstrated that SNORA40 and SNORA70 loss-of-function impairs myogenic differentiation. Interestingly, gain-of-function can rescue impaired differentiation muscle progenitor cells in myotonic dystrophy type 1 (DM1). We identified cyclin D3 (CCND3) mRNA, which is partially located in the nucleolus, as a target for SNORA40 and SNORA70, which are required for its pseudouridylated status. Expression of the CCND3 protein is required for muscle progenitors to exit the cell-cycle when they are induced to differentiate. We revealed that this switch requires SNORA40/70. Finally, we observed that DM1 cells show reduced levels of SNORA40/70 and undetectable CCND3 protein. However, restoring normal levels of SNORA40/70 partially restored CCND3 protein expression, coinciding with improved cell fusion capacity in DM1 muscle progenitors. Collectively, these data suggest that this effect may stem from SNORA40/70-dependent pseudouridylation of CCND3 mRNA, emphasizing snoRNAs as key players in normal and pathological muscle differentiation.","PeriodicalId":19471,"journal":{"name":"Nucleic Acids Research","volume":"53 6","pages":""},"PeriodicalIF":16.6,"publicationDate":"2025-03-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11954525/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143743407","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Exploring the relationship between aptamer binding thermodynamics, affinity, and specificity.

IF 16.6 2区生物学

Nucleic Acids Research Pub Date : 2025-03-20 DOI: 10.1093/nar/gkaf219

Obtin Alkhamis, Caleb Byrd, Juan Canoura, Adara Bacon, Ransom Hill, Yi Xiao

{"title":"Exploring the relationship between aptamer binding thermodynamics, affinity, and specificity.","authors":"Obtin Alkhamis, Caleb Byrd, Juan Canoura, Adara Bacon, Ransom Hill, Yi Xiao","doi":"10.1093/nar/gkaf219","DOIUrl":"10.1093/nar/gkaf219","url":null,"abstract":"Aptamers are oligonucleotide-based bioreceptors that are selected in vitro from randomized libraries to bind specific molecules with high affinity, and are proving popular for applications in diagnostics, bioimaging, and therapeutics. A better understanding of aptamer-ligand interactions could facilitate sequence engineering efforts to improve aptamer binding properties, and perhaps eventually allow for the direct design of high-quality aptamers. To date, however, there have been very few comprehensive studies exploring the relationship between aptamer binding properties and thermodynamics. Isothermal titration calorimetry (ITC) is a gold-standard method for studying the thermodynamics of ligand-receptor interactions. In this work, we have compiled ITC-derived thermodynamic binding data from 317 small-molecule-binding DNA aptamers, along with specificity profiles for ∼6000 aptamer-ligand pairs, and performed systematic analysis of the resulting datasets. This analysis revealed a variety of interesting patterns and trends. For example, ligand binding for most aptamers is generally driven solely by enthalpy, and aptamers with the highest binding enthalpy and greatest entropic binding penalties consistently have high specificity. We envision that the expansion and further analysis of such datasets will yield a far better understanding of the complex interplay between the various non-covalent interactions underlying aptamer-ligand recognition.","PeriodicalId":19471,"journal":{"name":"Nucleic Acids Research","volume":"53 6","pages":""},"PeriodicalIF":16.6,"publicationDate":"2025-03-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11952966/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143743431","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Methyl-GP: accurate generic DNA methylation prediction based on a language model and representation learning.

IF 16.6 2区生物学

Nucleic Acids Research Pub Date : 2025-03-20 DOI: 10.1093/nar/gkaf223

Hao Xie, Leyao Wang, Yuqing Qian, Yijie Ding, Fei Guo

{"title":"Methyl-GP: accurate generic DNA methylation prediction based on a language model and representation learning.","authors":"Hao Xie, Leyao Wang, Yuqing Qian, Yijie Ding, Fei Guo","doi":"10.1093/nar/gkaf223","DOIUrl":"10.1093/nar/gkaf223","url":null,"abstract":"Accurate prediction of DNA methylation remains a challenge. Identifying DNA methylation is important for understanding its functions and elucidating its role in gene regulation mechanisms. In this study, we propose Methyl-GP, a general predictor that accurately predicts three types of DNA methylation from DNA sequences. We found that the conservation of sequence patterns among different species contributes to enhancing the generalizability of the model. By fine-tuning a language model on a dataset comprising multiple species with similar sequence patterns and employing a fusion module to integrate embeddings into a high-quality comprehensive representation, Methyl-GP demonstrates satisfactory predictive performance in methylation identification. Experiments on 17 benchmark datasets for three types of DNA methylation (4mC, 5hmC, and 6mA) demonstrate the superiority of Methyl-GP over existing predictors. Furthermore, by utilizing the attention mechanism, we have visualized the sequence patterns learned by the model, which may help us to gain a deeper understanding of methylation patterns across various species.","PeriodicalId":19471,"journal":{"name":"Nucleic Acids Research","volume":"53 6","pages":""},"PeriodicalIF":16.6,"publicationDate":"2025-03-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11952970/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143743440","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Light-dependent flavin redox and adduct states control the conformation and DNA-binding activity of the transcription factor EL222. 光依赖性黄素氧化还原和加合物状态控制转录因子 EL222 的构象和 DNA 结合活性。

IF 16.6 2区生物学

Nucleic Acids Research Pub Date : 2025-03-20 DOI: 10.1093/nar/gkaf215

Aditya S Chaudhari, Adrien Favier, Zahra Aliakbar Tehrani, Tomáš Kovaľ, Inger Andersson, Bohdan Schneider, Jan Dohnálek, Jiří Černý, Bernhard Brutscher, Gustavo Fuertes

{"title":"Light-dependent flavin redox and adduct states control the conformation and DNA-binding activity of the transcription factor EL222.","authors":"Aditya S Chaudhari, Adrien Favier, Zahra Aliakbar Tehrani, Tomáš Kovaľ, Inger Andersson, Bohdan Schneider, Jan Dohnálek, Jiří Černý, Bernhard Brutscher, Gustavo Fuertes","doi":"10.1093/nar/gkaf215","DOIUrl":"10.1093/nar/gkaf215","url":null,"abstract":"The activity of the light-oxygen-voltage/helix-turn-helix (LOV-HTH) photoreceptor EL222 is regulated through protein-protein and protein-DNA interactions, both triggered by photo-excitation of its flavin mononucleotide (FMN) cofactor. To gain molecular-level insight into the photocycle of EL222, we applied complementary methods: macromolecular X-ray crystallography (MX), nuclear magnetic resonance (NMR) spectroscopy, optical spectroscopies (infrared and UV-visible), molecular dynamics/metadynamics (MD/metaD) simulations, and protein engineering using noncanonical amino acids. Kinetic experiments provided evidence for two distinct EL222 conformations (lit1 and lit2) that become sequentially populated under illumination. These two lit states were assigned to covalently bound N5 protonated, and noncovalently bound hydroquinone forms of FMN, respectively. Only subtle structural differences were observed between the monomeric forms of all three EL222 species (dark, lit1, and lit2). While the dark state is largely monomeric, both lit states undergo monomer-dimer exchange. Furthermore, molecular modeling revealed differential dynamics and interdomain separation times arising from the three FMN states (oxidized, adduct, and reduced). Unexpectedly, all three EL222 species can associate with DNA, but only upon blue-light irradiation, a high population of stable complexes is obtained. Overall, we propose a model of EL222 activation where photoinduced changes in the FMN moiety shift the population equilibrium toward an open conformation that favors self-association and DNA-binding.","PeriodicalId":19471,"journal":{"name":"Nucleic Acids Research","volume":"53 6","pages":""},"PeriodicalIF":16.6,"publicationDate":"2025-03-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11928941/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143677059","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Targeted translation inhibition of chloroplast and mitochondrial mRNAs by designer pentatricopeptide repeat proteins.

IF 16.6 2区生物学

Nucleic Acids Research Pub Date : 2025-03-20 DOI: 10.1093/nar/gkaf222

Nikolay Manavski, Serena Schwenkert, Hans-Henning Kunz, Dario Leister, Jörg Meurer

{"title":"Targeted translation inhibition of chloroplast and mitochondrial mRNAs by designer pentatricopeptide repeat proteins.","authors":"Nikolay Manavski, Serena Schwenkert, Hans-Henning Kunz, Dario Leister, Jörg Meurer","doi":"10.1093/nar/gkaf222","DOIUrl":"10.1093/nar/gkaf222","url":null,"abstract":"Pentatricopeptide repeat (PPR) proteins are crucial for organellar gene expression. To establish a tool for gene expression manipulation in Arabidopsis plastids and genetically inaccessible mitochondria, we engineered designer (dPPR) proteins to specifically inhibit the translation of organellar mRNAs by masking their start codons. Unlike prior methods for targeted downregulation of gene expression, which rely on re-targeting native PPR proteins to RNA sequences closely related to their original targets, our approach employs a synthetic P-type PPR scaffold that can be designed to bind any RNA sequence of interest. Here, using dPPR-psbK and dPPR-nad7, we targeted the psbK mRNA in chloroplasts and the nad7 mRNA in mitochondria, respectively. dPPR-psbK effectively bound to psbK mRNA and inhibited its translation with high specificity, resulting in disrupted PSII supercomplexes and reduced photosynthetic efficiency. dPPR-nad7 suppressed nad7 translation, affecting NADH oxidase activity in complex I and growth retardation. Comparing phenotypes with tobacco psbK knockouts and nad7 knockdown bir6-2 mutants, along with quantitative proteomics, showed no clear evidence of physiologically relevant off-target effects. Our findings establish dPPR proteins as precise tools for targeted translation inhibition, facilitating functional studies of organellar genes and offering a novel approach with potential for manipulating organellar gene expression in diverse plant species.","PeriodicalId":19471,"journal":{"name":"Nucleic Acids Research","volume":"53 6","pages":""},"PeriodicalIF":16.6,"publicationDate":"2025-03-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11941472/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143730782","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Mapping small molecule-RNA binding sites via Chem-CLIP synergized with capillary electrophoresis and nanopore sequencing.

IF 16.6 2区生物学

Nucleic Acids Research Pub Date : 2025-03-20 DOI: 10.1093/nar/gkaf231

Xueyi Yang, Jielei Wang, Noah A Springer, Patrick R A Zanon, Yilin Jia, Xiaoxuan Su, Matthew D Disney

{"title":"Mapping small molecule-RNA binding sites via Chem-CLIP synergized with capillary electrophoresis and nanopore sequencing.","authors":"Xueyi Yang, Jielei Wang, Noah A Springer, Patrick R A Zanon, Yilin Jia, Xiaoxuan Su, Matthew D Disney","doi":"10.1093/nar/gkaf231","DOIUrl":"10.1093/nar/gkaf231","url":null,"abstract":"Target validation and identification of binding sites are keys to the development of bioactive small molecules that target RNA. Herein, we describe optimized protocols to profile small molecule-RNA interactions and to define binding sites of the small molecules in RNAs using covalent chemistry. Various reactive modules appended to an RNA-binding small molecule were studied for cross-linking to the RNA target. Electrophilic modules, whether N-chloroethyl aniline or diazirine, have reactive profiles consistent with induced proximity; however, probes with N-chloroethyl aniline were more reactive and more specific than those with a diazirine cross-linking moiety. Depending upon the identity of the cross-linking module, covalent adducts with different nucleotides that are proximal to a small molecule's binding site were formed. The nucleotides where cross-linking occurred were elucidated by using two different platforms: (i) automated capillary electrophoresis that identified a binding site by impeding reverse transcriptase, or \"RT stops\"; and (ii) nanopore sequencing where the cross-link produces mutations in the corresponding complementary DNA formed by reverse transcriptase-polymerase chain reaction amplification of the cross-linked RNA. These approaches are broadly applicable to aid in the advancement of chemical probes targeting RNA, including identifying binding sites and using covalent chemistry to screen for RNA-binding molecules in a high throughput format.","PeriodicalId":19471,"journal":{"name":"Nucleic Acids Research","volume":"53 6","pages":""},"PeriodicalIF":16.6,"publicationDate":"2025-03-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11952968/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143743434","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Structural basis for control of integrative and conjugative element excision and transfer by the oligomeric winged helix-turn-helix protein RdfS.

IF 16.6 2区生物学

Nucleic Acids Research Pub Date : 2025-03-20 DOI: 10.1093/nar/gkaf249

Callum J Verdonk, Mark Agostino, Karina Yui Eto, Drew A Hall, Charles S Bond, Joshua P Ramsay

{"title":"Structural basis for control of integrative and conjugative element excision and transfer by the oligomeric winged helix-turn-helix protein RdfS.","authors":"Callum J Verdonk, Mark Agostino, Karina Yui Eto, Drew A Hall, Charles S Bond, Joshua P Ramsay","doi":"10.1093/nar/gkaf249","DOIUrl":"10.1093/nar/gkaf249","url":null,"abstract":"Winged helix-turn-helix (wHTH) proteins are diverse DNA-binding proteins that often oligomerize on DNA and participate in DNA recombination and transcriptional regulation. wHTH recombination directionality factors (RDFs) associated with tyrosine recombinases, stimulate excision of prophage and integrative and conjugative elements (ICEs). RdfS is required for excision and conjugation of the Mesorhizobium japonicum R7A ICE, ICEMlSymR7A, which carries genes for nitrogen-fixing symbiosis. We show RdfS binds to DNA regions within the IntS attachment site (attP) and within the rdfS promoter, enabling RdfS to coordinate rdfS/intS expression and stimulate RdfS/IntS-mediated ICEMlSymR7A excision. Several RdfS DNA-binding sites were identified. However, no consensus motif was apparent and no individual nucleotide substitutions in attP prevented RdfS binding. RdfS forms extensive helical filaments in crystals, with subunits contacting via a novel α1-helix absent in other wHTH-RDFs. RdfS oligomerized in solution in the absence of DNA. Molecular dynamics simulations supported a role for the α1-helix in oligomerization and compaction of nucleoprotein complexes. Removal of RdfS-α1 did not eliminate DNA-binding in vitro but reduced oligomerization and abolished RdfS-mediated ICEMlSymR7A excision and conjugative transfer. We propose the novel RdfS-α1 mediated oligomerization enables RdfS to specifically recognize larger DNA regions with low primary sequence conservation through an indirect readout mechanism.","PeriodicalId":19471,"journal":{"name":"Nucleic Acids Research","volume":"53 6","pages":""},"PeriodicalIF":16.6,"publicationDate":"2025-03-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11963761/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143773019","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

BmNPV interacts with super-enhancer regions of the host chromatin to hijack cellular transcription machinery.

IF 16.6 2区生物学

Nucleic Acids Research Pub Date : 2025-03-20 DOI: 10.1093/nar/gkaf188

Shudi Zhao, Xingyang Wang, Tian Yang, Xinyu Zhu, Xiaofeng Wu

{"title":"BmNPV interacts with super-enhancer regions of the host chromatin to hijack cellular transcription machinery.","authors":"Shudi Zhao, Xingyang Wang, Tian Yang, Xinyu Zhu, Xiaofeng Wu","doi":"10.1093/nar/gkaf188","DOIUrl":"10.1093/nar/gkaf188","url":null,"abstract":"Effective transcriptional activation relies on the spatial interaction between specific DNA elements. DNA interactions have also been observed between DNA viruses and their hosts, with limited understanding of the involved details. Baculovirus is a representative species of DNA virus and has been reported to interact with the host genome in our previous study. However, the biological significance of the baculovirus-host trans-species DNA interaction and its underlying mechanisms remain elusive. Here, using Bombyx mori nucleopolyhedrovirus (BmNPV) as the model virus, we combine epigenome, transcriptome, and biochemical assays to investigate the baculovirus-host DNA interaction. Our data show that BmNPV hijacks the transcriptional regulatory capacity of host super-enhancers (SEs) by physically interacting with these regions on the host genome. This results in the usurpation of the activating capacity of an SE-binding transcription factor GATA by the virus, thereby impairing the SE-induced specific transcriptional activation of the target antiviral genes. Moreover, the hijacked regulatory capacity is spread on BmNPV genome through cis-interaction of viral DNA, leading to enhanced viral gene expression. Overall, our results provide novel insights into the intricate interplay of viruses with host gene expression regulatory networks and broaden the vision in the mechanisms of viral exploitation on cellular machinery.","PeriodicalId":19471,"journal":{"name":"Nucleic Acids Research","volume":"53 6","pages":""},"PeriodicalIF":16.6,"publicationDate":"2025-03-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11934923/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143710806","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Efficient DNA- and virus-free engineering of cellular transcriptomic states using dCas9 ribonucleoprotein (dRNP) complexes.

IF 16.6 2区生物学

Nucleic Acids Research Pub Date : 2025-03-20 DOI: 10.1093/nar/gkaf235

Tobias Schmidt, Maximilian Wiesbeck, Luisa Egert, Thi-Tram Truong, Anna Danese, Lukas Voshagen, Simon Imhof, Matilde Iraci Borgia, Deeksha, Andrea M Neuner, Anna Köferle, Arie Geerlof, André Santos Dias Mourão, Stefan H Stricker

{"title":"Efficient DNA- and virus-free engineering of cellular transcriptomic states using dCas9 ribonucleoprotein (dRNP) complexes.","authors":"Tobias Schmidt, Maximilian Wiesbeck, Luisa Egert, Thi-Tram Truong, Anna Danese, Lukas Voshagen, Simon Imhof, Matilde Iraci Borgia, Deeksha, Andrea M Neuner, Anna Köferle, Arie Geerlof, André Santos Dias Mourão, Stefan H Stricker","doi":"10.1093/nar/gkaf235","DOIUrl":"10.1093/nar/gkaf235","url":null,"abstract":"For genome editing, the use of CRISPR ribonucleoprotein (RNP) complexes is well established and often the superior choice over plasmid-based or viral strategies. RNPs containing dCas9 fusion proteins, which enable the targeted manipulation of transcriptomes and epigenomes, remain significantly less accessible. Here, we describe the production, delivery, and optimization of second generation CRISPRa RNPs (dRNPs). We characterize the transcriptional and cellular consequences of dRNP treatments in a variety of human target cells and show that the uptake is very efficient. The targeted activation of genes demonstrates remarkable potency, even for genes that are strongly silenced, such as developmental master transcription factors. In contrast to DNA-based CRISPRa strategies, gene activation is immediate and characterized by a sharp temporal precision. We also show that dRNPs allow very high-target multiplexing, enabling undiminished gene activation of multiple genes simultaneously. Applying these insights, we find that intensive target multiplexing at single promoters synergistically elevates gene transcription. Finally, we demonstrate in human stem and differentiated cells that the preferable features of dRNPs allow to instruct and convert cell fates efficiently without the need for DNA delivery or viral vectors.","PeriodicalId":19471,"journal":{"name":"Nucleic Acids Research","volume":"53 6","pages":""},"PeriodicalIF":16.6,"publicationDate":"2025-03-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11952960/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143743428","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0