IF 5 3区生物学

RNA Pub Date : 2025-09-16 DOI: 10.1261/rna.080414.125

Michael H D'Souza, Higor S Pereira, Scott Tersteeg, Gunjan Vasudeva, M Quadir Siddiqui, Temi Agunlejika, Liam Kerr, Dylan Girodat, Trushar R Patel

{"title":"Solution structure determination and biophysical characterization studies of 7SK RNP and 7SL SRP RNAs.","authors":"Michael H D'Souza, Higor S Pereira, Scott Tersteeg, Gunjan Vasudeva, M Quadir Siddiqui, Temi Agunlejika, Liam Kerr, Dylan Girodat, Trushar R Patel","doi":"10.1261/rna.080414.125","DOIUrl":"10.1261/rna.080414.125","url":null,"abstract":"Long noncoding RNAs perform integral roles in eukaryotic life cycles, particularly the 7SK snRNA, which is responsible for RNA polymerase II transcription modulation and progression when interacting with P-TEFb, and the 7SL RNA involved in signal recognition particle mediation of cotranslation activities of endoplasmic reticulum bound proteins. These RNAs retain important secondary structures that interact with proteins involved in transcription and translation regulation. RNA-protein interactions involving the RNA stem-loops have been previously characterized using chemical probing techniques, cryo-electron microscopy, and nuclear magnetic resonance. However, complete three-dimensional structures of the full-length 7SK and 7SL have not been resolved, limiting our understanding of these RNAs' tertiary landscapes and mechanisms. Our study bridges this gap in knowledge by using small-angle X-ray scattering and coarse-grained computational modeling of previously determined secondary structures through SimRNA to produce full-sequence, three-dimensional atomistic models of both 7SK and 7SL RNAs. We used size exclusion chromatography connected with light scattering and circular dichroism spectroscopy to verify RNA size, and compare previously identified secondary structures in solution. We additionally used all-atom, structure-based potential simulations to generate optimized models within our calculated SAXS envelopes. 7SK's total morphology is thus presented as a highly versatile structure whose well-defined stem-loops interact with each other in three-dimensional space. 7SL RNA is presented as a tightly wound and somewhat rigid structure, with significant base-pairing features in its Alu-domain whereupon it forms likely scaffolds for signal recognition peptide formation.","PeriodicalId":21401,"journal":{"name":"RNA","volume":" ","pages":"1436-1459"},"PeriodicalIF":5.0,"publicationDate":"2025-09-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12439597/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144761164","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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

Accurate in silico predictions of modified RNA interactions to a prototypical RNA-binding protein with λ-dynamics. 用λ动力学准确预测修饰RNA与典型RNA结合蛋白的相互作用。

IF 5 3区生物学

RNA Pub Date : 2025-09-16 DOI: 10.1261/rna.080367.124

Murphy Angelo, Yash Bhargava, Elzbieta Kierzek, Ryszard Kierzek, Ryan L Hayes, Wen Zhang, Jonah Z Vilseck, Scott Takeo Aoki

{"title":"Accurate in silico predictions of modified RNA interactions to a prototypical RNA-binding protein with λ-dynamics.","authors":"Murphy Angelo, Yash Bhargava, Elzbieta Kierzek, Ryszard Kierzek, Ryan L Hayes, Wen Zhang, Jonah Z Vilseck, Scott Takeo Aoki","doi":"10.1261/rna.080367.124","DOIUrl":"10.1261/rna.080367.124","url":null,"abstract":"RNA-binding proteins shape biology through their widespread functions in RNA biochemistry. Their function requires the recognition of specific RNA motifs for targeted binding. These RNA-binding elements can be composed of both unmodified and chemically modified RNAs, of which over 170 chemical modifications have been identified in biology. Unmodified RNA sequence preferences for RNA-binding proteins have been widely studied, with numerous methods available to identify their preferred sequence motifs. However, only a few techniques can detect preferred RNA modifications, and no current method can comprehensively screen the vast array of hundreds of natural RNA modifications. Prior work demonstrated that λ-dynamics is an accurate in silico method to predict RNA base binding preferences of an RNA-binding antibody. This work extends that effort by using λ-dynamics to predict unmodified and modified RNA-binding preferences of human Pumilio, a prototypical RNA-binding protein. A library of RNA modifications was screened at eight nucleotide positions along the RNA to identify modifications predicted to affect Pumilio binding. Computed binding affinities were compared with experimental data to reveal high predictive accuracy. In silico force field accuracies were also evaluated between CHARMM36 and Amber RNA force fields to determine the best parameter set to use in binding calculations. This work demonstrates that λ-dynamics can predict RNA interactions to a bona fide RNA-binding protein without the requirements of chemical reagents or new methods to experimentally test binding at the bench. Advancing in silico methods like λ-dynamics will unlock new frontiers in understanding how RNA modifications shape RNA biochemistry.","PeriodicalId":21401,"journal":{"name":"RNA","volume":" ","pages":"1460-1471"},"PeriodicalIF":5.0,"publicationDate":"2025-09-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12439594/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144761162","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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

Stress granules promote quiescence by enhancing p21 levels and reducing phospho-Rb. 应激颗粒通过提高p21水平和降低磷酸rb来促进静止。

IF 5 3区生物学

RNA Pub Date : 2025-09-16 DOI: 10.1261/rna.080635.125

Anthony Khong, Nina Ripin, Luisa Macedo de Vasconcelos, Victor Passanisi, Sabrina Spencer, Roy Parker

{"title":"Stress granules promote quiescence by enhancing p21 levels and reducing phospho-Rb.","authors":"Anthony Khong, Nina Ripin, Luisa Macedo de Vasconcelos, Victor Passanisi, Sabrina Spencer, Roy Parker","doi":"10.1261/rna.080635.125","DOIUrl":"10.1261/rna.080635.125","url":null,"abstract":"During the integrated stress response (ISR), most mRNAs exit translation and some condense into stress granules. Stress granules that form during chemotherapy can promote cancer cell survival and chemoresistance by an unknown mechanism. Cells can also spontaneously trigger the ISR at low levels, which promotes cellular quiescence where cells exit the cell cycle and are resistant to therapeutic agents. We hypothesized that the ability of cells to form stress granules might be a critical signal to drive cells into quiescence. Herein, we provide several observations that suggest stress granules enhance cell survival and chemoresistance by promoting cellular quiescence. The mechanism by which stress granules promote quiescence is by stimulating p21 expression, leading to inhibition of Rb phosphorylation. These results demonstrate that stress granule formation is sufficient to trigger cellular quiescence and argue that inhibitors of stress granules may be effective in combination with chemotherapy to limit the development of chemoresistance in treating human tumors.","PeriodicalId":21401,"journal":{"name":"RNA","volume":" ","pages":"1472-1487"},"PeriodicalIF":5.0,"publicationDate":"2025-09-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12439591/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144754149","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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

m⁶A methylation inhibits recruitment of the Dand5 3'UTR to the left-right determinant Bicc1. M6A甲基化抑制Dand5 3'UTR向左右行列式Bicc1的募集。

IF 5 3区生物学

RNA Pub Date : 2025-09-16 DOI: 10.1261/rna.080526.125

Benjamin Rothé, Mateusz Mendel, Simon Fortier, Daniel B Constam

引用次数: 0

Corrigendum: Copy number determination of sperm-borne small RNAs implied in the intergenerational inheritance of metabolic syndromes. 更正：代谢综合征代际遗传中隐含的精子携带小rna的拷贝数测定。

IF 5 3区生物学

RNA Pub Date : 2025-09-16 DOI: 10.1261/rna.080709.125

Lisa König, Victoria Guggenberger, Kristeli Eleftheriou, Zsuzsanna Pinter, Alessandro Marotto, Christoph R Kreutz, Mark Wossidlo, Virginie Marchand, Yuri Motorin, Matthias R Schaefer

引用次数: 0

Profiling RNA subcellular localization in situ by TATA-seq. 利用TATA-seq分析RNA亚细胞原位定位。

IF 5 3区生物学

RNA Pub Date : 2025-09-16 DOI: 10.1261/rna.080670.125

Junjie Li, Chu Xu, Xiao Jiang, Xing Huang, Dan Ye, Lulu Hu

{"title":"Profiling RNA subcellular localization in situ by TATA-seq.","authors":"Junjie Li, Chu Xu, Xiao Jiang, Xing Huang, Dan Ye, Lulu Hu","doi":"10.1261/rna.080670.125","DOIUrl":"10.1261/rna.080670.125","url":null,"abstract":"Membrane-less organelles, dynamic subcellular structures formed by RNA and RNA-binding proteins (RBPs) undergoing liquid-liquid phase separation (LLPS), play key roles in biological processes such as RNA degradation in processing bodies (P-bodies), translation inhibition in stress granules, and RNA splicing in nuclear speckles. However, the study of RNA species within these organelles has been hindered by the absence of simple, sensitive, and specific methodologies. Here, we introduce target transcript amplification and sequencing (TATA-seq), a novel strategy for precisely profiling RNA in membrane-less organelles via in situ targeted transcription and linear amplification. TATA-seq uses a primary antibody against a marker protein of the target organelle to recruit a secondary antibody conjugated with streptavidin, which binds an oligonucleotide containing a T7 promoter. This initiates in situ RNA reverse transcription, followed by amplification with T7 RNA polymerase to generate sufficient material for sequencing, ensuring a duplication rate of no more than 25% and a mapping ratio of ∼90%. An IgG control is used to subtract background noise during data analysis. We demonstrate the method's utility by profiling RNA in stress granules induced by sodium arsenite in HeLa cells, with validation through FISH and immunofluorescence colocalization. TATA-seq offers a simple, highly sensitive, and accurate tool for studying RNA dynamics in membrane-less organelles, advancing the capabilities of RNA research.","PeriodicalId":21401,"journal":{"name":"RNA","volume":" ","pages":"1523-1535"},"PeriodicalIF":5.0,"publicationDate":"2025-09-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12439595/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144765345","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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

Caenorhabditis elegans FBF-1 and FBF-2 C-terminal intrinsically disordered regions differentially regulate RNA-binding affinity. 秀丽隐杆线虫FBF-1和FBF-2 c端本序紊乱区对rna结合亲和力的调节存在差异。

IF 5 3区生物学

RNA Pub Date : 2025-09-16 DOI: 10.1261/rna.080578.125

Hope R Hawthorne, Chen Qiu, Traci M Tanaka Hall

{"title":"Caenorhabditis elegans FBF-1 and FBF-2 C-terminal intrinsically disordered regions differentially regulate RNA-binding affinity.","authors":"Hope R Hawthorne, Chen Qiu, Traci M Tanaka Hall","doi":"10.1261/rna.080578.125","DOIUrl":"10.1261/rna.080578.125","url":null,"abstract":"PUF proteins (named for Drosophila melanogaster Pumilio and Caenorhabditis elegans fem-3 mRNA binding factor or FBF) are a family of RNA-binding proteins. C. elegans FBF is a collective term for two PUF proteins, FBF-1 and FBF-2, that maintain germline stem cells. FBF binds the 3'UTR of target RNAs and together with partner proteins represses translation of mRNAs that promote differentiation. Until recently, little was known about the functions of the FBF C-terminal intrinsically disordered regions that follow the RNA-binding domain (RBD). Despite high overall protein sequence conservation (91% identical residues), the FBF-1 and FBF-2 C-terminal tails (CTs) are distinct, and the FBF-2 CT is essential for its function. The FBF-2 CT contains a PUF-interacting motif (PIM) that binds its own RBD and autoinhibits RNA-binding affinity. Here we investigated whether differences in the FBF-1 and FBF-2 CTs impact molecular function. Unlike FBF-2, the FBF-1 CT had no impact on RNA binding. Despite this, a crystal structure of FBF-1 demonstrated that a PIM in the FBF-1 CT binds to its RBD, like FBF-2. By creating FBF-1/FBF-2 chimeric proteins, we discovered that the FBF-2 CT can autoinhibit FBF-1 RNA binding, and substitution of the FBF-1 PIM for the FBF-2 PIM diminished FBF-2 autoinhibition. Our results exemplify how RBP paralogs diverge to fine-tune their RNA-binding activities.","PeriodicalId":21401,"journal":{"name":"RNA","volume":" ","pages":"1391-1402"},"PeriodicalIF":5.0,"publicationDate":"2025-09-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12439598/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144795276","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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

Development of a modified RNA circularization system to improve circRNA-based protein expression in mammalian cells. 一种改良的RNA循环系统的开发，以提高哺乳动物细胞中基于环状RNA的蛋白表达。

IF 5 3区生物学

RNA Pub Date : 2025-09-11 DOI: 10.1261/rna.080733.125

Mingting Cui, Shunran Li, Yuhang Han, Minchao Li, Zirong Han, Jun Qian, Zhi Xie, Caijun Sun

{"title":"Development of a modified RNA circularization system to improve circRNA-based protein expression in mammalian cells.","authors":"Mingting Cui, Shunran Li, Yuhang Han, Minchao Li, Zirong Han, Jun Qian, Zhi Xie, Caijun Sun","doi":"10.1261/rna.080733.125","DOIUrl":"https://doi.org/10.1261/rna.080733.125","url":null,"abstract":"Circular RNA (circRNA) is emerging as a highly promising technology in various biomedical applications, offering advantages over traditional linear RNA. The Tornado (Twister-optimized RNA for durable overexpression) system has been widely investigated for generating circRNAs in mammalian cells; however, the use of Tornado system for large RNA inserts, especially those containing IRES sequences, is hindered by low circularization efficiency and limited circRNA abundance. Therefore, developing novel strategies to enhance RNA circularization in cells is of critical importance. In this study, we present a modified Tornado system that significantly improves circRNA-based protein expression by incorporating an optimal distance between the internal ribosome entry site (IRES) and the upstream CMV promoter. Furthermore, we elucidated the dual roles of HRV-B3 IRES in mammalian cells, demonstrating its negative regulatory effect on RNA abundance and its positive contribution to RNA circularization. Additionally, the integration of a truncated 5' long terminal repeat (LTR) from HIV-1 upstream of the HRV-B3 IRES, combined with the woodchuck hepatitis virus posttranscriptional regulatory element (WPRE), further enhanced transcriptional efficiency in the Tornado system. This modified system holds great potential for advancing circRNA-based therapeutics and vaccines, and these findings provide valuable insights for refining the Tornado system and designing regulatory elements in synthetic biology applications.","PeriodicalId":21401,"journal":{"name":"RNA","volume":" ","pages":""},"PeriodicalIF":5.0,"publicationDate":"2025-09-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145041324","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Genetic suppression of precocious transcription termination identifies mutations in essential subunits of the fission yeast cleavage and polyadenylation machinery. 基因抑制的早熟转录终止鉴定突变的基本亚单位的分裂酵母切割和聚腺苷酸化机制。

IF 5 3区生物学

RNA Pub Date : 2025-09-04 DOI: 10.1261/rna.080664.125

Aleksei Innokentev, Ana M Sanchez, Lauren Bednor, Jill Babor, Beate Schwer, Stewart Shuman

{"title":"Genetic suppression of precocious transcription termination identifies mutations in essential subunits of the fission yeast cleavage and polyadenylation machinery.","authors":"Aleksei Innokentev, Ana M Sanchez, Lauren Bednor, Jill Babor, Beate Schwer, Stewart Shuman","doi":"10.1261/rna.080664.125","DOIUrl":"https://doi.org/10.1261/rna.080664.125","url":null,"abstract":"The fission yeast phosphate acquisition (PHO) regulon is repressed under phosphate-replete conditions by upstream lncRNA-mediated transcriptional interference. Inositol-1-pyrophosphates control phosphate homeostasis via their action as agonists of precocious PHO lncRNA 3'-processing/termination. Inositol pyrophosphatase-inactivating mutations that increase inositol-1-pyrophosphates elicit derepression of the PHO genes and a severe growth defect in YES medium. Previous studies demonstrated suppression of inositol pyrophosphate toxicosis by targeted deletion or loss-of-function mutations in the nonessential Ssu72, Ppn1, Swd22, and Ctf1 subunits of the fission yeast Cleavage and Polyadenylation Factor (CPF) complex. Here we conducted a screen for spontaneous mutations that suppress the precocious PHO lncRNA termination underlying the sickness of asp1-STF pyrophosphatase mutants. We thereby recovered and characterized novel hypomorphic missense mutations in five essential CPF subunits: Ysh1 (the cleavage endonuclease), Pta1 (an Armadillo/HEAT-repeat protein), Pfs2 (a WD repeat protein), Cft1 (a WD repeat protein), and Msi2 (a tandem RRM RNA-binding protein). The screen also yielded an intron branchpoint mutation in the gene encoding essential CPF subunit Iss1. In addition, we found that asp1-STF toxicosis was suppressed by a missense mutation in the active site of Pla1, the essential poly(A) polymerase subunit of CPF. Genetic crosses revealed a hierarchy of mutational synergies between the essential CPF subunits, the inessential CPF subunits, termination factor Rhn1, the Thr4 \"letter\" of the RNA polymerase II CTD code, and the Asp1 kinase that synthesizes inositol-1-pyrophosphates. The synthetic lethality of msi2-G252E with ctf1∆, swd22∆, ppn1∆, ssu72-C13S, rpb1-CTD-T4A, and asp1∆ establishes Msi2 as a central agent of 3'-processing/termination, functioning in parallel to inositol-1-pyrophosphates.","PeriodicalId":21401,"journal":{"name":"RNA","volume":" ","pages":""},"PeriodicalIF":5.0,"publicationDate":"2025-09-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145001437","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

From activator to suppressor: PACT is joining the company of PKR negative regulators. 从激活剂到抑制剂：PACT加入PKR负调控剂的行列。

IF 5 3区生物学

RNA Pub Date : 2025-09-03 DOI: 10.1261/rna.080743.125

Francisco M M Acosta, Christian K Pfaller

引用次数: 0

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