U1 interference (U1i) for Antiviral Approaches

Blázquez, Lorea; Fortes, Puri

doi:10.1007/978-1-4939-2432-5_3

Cited by 11 publications

(8 citation statements)

References 56 publications

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“…With the exception of the case of F30 insulators, experiments obtained thus far suggest that stronger inhibitions by U1 snRNP could result in better inducibility after TC-mediated structural shifts. We have previously shown that multiplexing U1 BSs leads to synergistic inhibitions (reviewed in refs and ). Thus, we combined U1in and U1out riboswitches in the reporter cassette with the aim of increasing the dynamic range of the system (Figure a).…”

Section: Resultsmentioning

confidence: 99%

Engineering U1-Based Tetracycline-Inducible Riboswitches to Control Gene Expression in Mammals

Rovira,

Moreno,

Razquin

et al. 2023

ACS Nano

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Synthetic riboswitches are promising regulatory devices due to their small size, lack of immunogenicity, and ability to fine-tune gene expression in the absence of exogenous trans-acting factors. Based on a gene inhibitory system developed at our lab, termed U1snRNP interference (U1i), we developed tetracycline (TC)-inducible riboswitches that modulate mRNA polyadenylation through selective U1 snRNP recruitment. First, we engineered different TC-U1i riboswitches, which repress gene expression unless TC is added, leading to inductions of gene expression of 3-to-4-fold. Second, we developed a technique called Systematic Evolution of Riboswitches by Exponential Enrichment (SEREX), to isolate riboswitches with enhanced U1 snRNP binding capacity and activity, achieving inducibilities of up to 8-fold. Interestingly, by multiplexing riboswitches we increased inductions up to 37-fold. Finally, we demonstrated that U1i-based riboswitches are dose-dependent and reversible and can regulate the expression of reporter and endogenous genes in culture cells and mouse models, resulting in attractive systems for gene therapy applications. Our work probes SEREX as a much-needed technology for the in vitro identification of riboswitches capable of regulating gene expression in vivo.

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Section: Resultsmentioning

confidence: 99%

Engineering U1-Based Tetracycline-Inducible Riboswitches to Control Gene Expression in Mammals

Rovira,

Moreno,

Razquin

et al. 2023

ACS Nano

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“…Although the C25G base change did not result in an improvement in the splicing of the Dup51p reporter, the improvement in binding of U1-70K was significant. Modified U1 snRNAs carrying base changes to their 5′-regions are widely used for correction of adverse effects of mutant 5′-ss, and it may be worthwhile to test if the C25G base change in the engineered U1 snRNAs enhances splicing of other targets ( Blázquez and Fortes 2015 ).…”

Section: Discussionmentioning

confidence: 99%

Examining the capacity of human U1 snRNA variants to facilitate pre-mRNA splicing

Wong,

Yellamaty,

Gallante

et al. 2023

RNA

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The human U1 snRNA is encoded by a multigene family consisting of transcribed variants and defective pseudogenes. Many variant U1 (vU1) snRNAs have been demonstrated to not only be transcribed but also processed by the addition of a trimethylated guanosine cap, packaged into snRNPs, and assembled into spliceosomes, however, their capacity to facilitate pre-mRNA splicing has, so far, not been tested. A recent systematic analysis of the human snRNA genes identified 178 U1 snRNA genes that are present in the genome as either tandem arrays or single genes on multiple chromosomes. Of these, 15 were found to be expressed in human tissues and cell lines, although at significantly low levels from their endogenous loci, less than 0.001% of the canonical U1 snRNA. In this study, we found that placing the variants in the context of the regulatory elements of theRNU1-1gene improves expression of many variants to levels comparable to the canonical U1 snRNA. Application of a previously established HeLa cell based minigene reporter assay to examine the capacity of the vU1 snRNAs to support pre-mRNA splicing revealed that even though the exogenously expressed variant snRNAs were enriched in the nucleus, only a few had a measurable effect on splicing.

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“…Although the experience concerning therapeutic targeting of splicing is far more advanced, the general proof-of-concept of targeting specific polyadenylation sites for redirection of processing based on analogous approaches has been made. This includes the use of ASOs [ 179 ] and siRNAs [ 178 ] as well as modified U1 snRNP, which interacts with a target gene upstream of its PAS to regulate gene expression [ 14 ].…”

Section: Targeting Mrna 3′ End Formation As a Novel Therapymentioning

confidence: 99%

Processing and transcriptome expansion at the mRNA 3′ end in health and disease: finding the right end

Ogorodnikov

Kargapolova

Danckwardt

2016

Pflugers Arch - Eur J Physiol

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The human transcriptome is highly dynamic, with each cell type, tissue, and organ system expressing an ensemble of transcript isoforms that give rise to considerable diversity. Apart from alternative splicing affecting the “body” of the transcripts, extensive transcriptome diversification occurs at the 3′ end. Transcripts differing at the 3′ end can have profound physiological effects by encoding proteins with distinct functions or regulatory properties or by affecting the mRNA fate via the inclusion or exclusion of regulatory elements (such as miRNA or protein binding sites). Importantly, the dynamic regulation at the 3′ end is associated with various (patho)physiological processes, including the immune regulation but also tumorigenesis. Here, we recapitulate the mechanisms of constitutive mRNA 3′ end processing and review the current understanding of the dynamically regulated diversity at the transcriptome 3′ end. We illustrate the medical importance by presenting examples that are associated with perturbations of this process and indicate resulting implications for molecular diagnostics as well as potentially arising novel therapeutic strategies.

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U1 interference (U1i) for Antiviral Approaches

Cited by 11 publications

References 56 publications

Engineering U1-Based Tetracycline-Inducible Riboswitches to Control Gene Expression in Mammals

Engineering U1-Based Tetracycline-Inducible Riboswitches to Control Gene Expression in Mammals

Examining the capacity of human U1 snRNA variants to facilitate pre-mRNA splicing

Processing and transcriptome expansion at the mRNA 3′ end in health and disease: finding the right end

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