ArticleJournal of biological engineering2014
Modular assembly of designer PUF proteins for specific post-transcriptional regulation of endogenous RNA.
Article in Journal of biological engineering, 2014. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 24 papers.
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Who cites it
24 citing papers in PubMed, 57 citations in OpenAlex.
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- Multi-level precise regulation of gene transcription in the yeast Saccharomyces cerevisiae based on light-sensitive CRISPR/Cas systems.Nucleic acids research · 2026Article
- GRASP: a modular toolkit for building synthetic pentatricopeptide repeat RNA-binding proteins.Nucleic acids research · 2025Article
- Repurposing CRISPR-Cas13 systems for robust mRNA trans-splicing.Nature communications · 2024Article
- Spatiotemporal control of RNA metabolism and CRISPR-Cas functions using engineered photoswitchable RNA-binding proteins.Nature protocols · 2024Review
- CRISPR-associated type V proteins as a tool for controlling mRNA stability in S. cerevisiae synthetic gene circuits.Nucleic acids research · 2023Article
- Building Blocks of Artificial CRISPR-Based Systems beyond Nucleases.International journal of molecular sciences · 2022Review
- Article
- Optogenetic control of RNA function and metabolism using engineered light-switchable RNA-binding proteins.Nature biotechnology · 2022Article
- Enabling one-pot Golden Gate assemblies of unprecedented complexity using data-optimized assembly design.PloS one · 2020Article
- Programmable RNA manipulation in living cells.Cellular and molecular life sciences : CMLS · 2019Review
- Sequence-specific mRNA (New York, N.Y.) · 2019Article
- Enhanced CRISPR-based DNA demethylation by Casilio-ME-mediated RNA-guided coupling of methylcytosine oxidation and DNA repair pathways.Nature communications · 2019Article
- Article
- Epitranscriptomics: A New Regulatory Mechanism of Brain Development and Function.Frontiers in neuroscience · 2018Review
- Review
- Aio-Casilio: a robust CRISPR-Cas9-Pumilio system for chromosome labeling.Journal of molecular histology · 2017Article
- Making the Message Clear: Concepts for mRNA Imaging.ACS central science · 2017Review
- Inducible Control of mRNA Transport Using Reprogrammable RNA-Binding Proteins.ACS synthetic biology · 2017Article
Corrections and comments
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Authors and funding
3 authors at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
backgroundDue to their modular repeat structure, Pumilio/fem-3 mRNA binding factor (PUF) proteins are promising candidates for designer RNA-binding protein (RBP) engineering. To further facilitate the application of the PUF domain for the sequence-specific RBP engineering, a rapid cloning approach is desirable that would allow efficient introduction of multiple key amino acid mutations in the protein. Here, we report the implementation of the Golden Gate cloning method for an efficient one-step assembly of a designer PUF domain for RNA specificity engineering.
resultsWe created a repeat module library that is potentially capable of generating a PUF domain with any desired specificity. PUF domains with multiple repeat modifications for the recognition of altered RNA targets were obtained in a one-step assembly reaction, which was found to be highly efficient. The new PUF variants exhibited high in vitro binding efficiencies to cognate RNA sequences, corroborating the applicability of the modular approach for PUF engineering. To demonstrate the application of the PUF domain assembly method for RBP engineering, we fused the PUF domain to a post-transcriptional regulator and observed a sequence-specific reporter and endogenous gene repression in human cell lines.
conclusionsThe Golden Gate based cloning approach thus should allow greater flexibility and speed in implementing the PUF protein scaffold for engineering designer RBPs, and facilitate its use as a tool in basic and applied biology and medicine.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.