🧬 PubMed RNA Editing Daily Digest

This study aims to develop a label-free electrochemical biosensor for the detection of chikungunya virus (CHIKV) RNA, based on CRISPR-Cas13a integrated with a cerium oxide (ceria)-modified screen-printed carbon electrode (SPCE). The ceria film was deposited through cathodic electrodeposition, forming a uniform, needle-like film, as observed by SEM, and a crystalline fluorite structure was confirmed by XRD with characteristic (111), (200), and (220) reflections. The results showed that Raman spectroscopy demonstrated a dominant F2g band at ∼463 cm

CRISPR activation (CRISPRa) offers a powerful approach to upregulate endogenous genes; yet, existing systems in plants can be complex or difficult to integrate with CRISPR interference (CRISPRi). Here, we present a streamlined and flexible CRISPRa platform that enables robust gene activation. Using a dual-luciferase reporter, we benchmarked a range of guide RNA scaffolds, effector proteins, and promoters. We developed a novel single-guide RNA (sgRNA) architecture, harboring two MS2 aptamers inserted into the tetraloop and driven by a composite Pol II/Pol III promoter, as the most efficient configuration. This scaffold outperformed gR2.0- and SunTag-based constructs, reaching up to 100-fold activation of a minimal 35S promoter and up to 215-fold induction of three endogenous rice genes in protoplast assays. In contrast, scaffold RNAs (scRNAs) with aptamers at the 3' end or in excessive copy numbers were ineffective. Exploratory AlphaFold modeling supports a possible role for aptamer positioning and MCP-VP64 dimerization, although this remains a working hypothesis. This modular design enables tunable gene regulation in rice protoplasts and provides a practical platform for high-throughput screening and synthetic gene circuit prototyping in plants. Given that scRNA geometry and promoter architecture are universal features of CRISPR-based transcriptional modulation, the system is expected to be broadly portable across species. While the architecture is intended to be compatible with CRISPRi, future studies will be needed to establish its practical use in combined CRISPRa/i settings.

Heterologous expression systems have been instrumental in furthering our understanding of plant RNA editing proteins. In this commentary, we discuss how the establishment of yeast as a model for studying plant RNA editing by Ramanathan et al. could advance the engineering of pentatricopeptide repeat proteins, and how in return pentatricopeptide repeat proteins might be used to advance yeast engineering.

Primate genomes harbor over a million Alu retrotransposons, yet how cells limit their impact on transcriptome stability during lineage specification remains unclear. Here, we show that the chromatin-associated ILF2/3 complex suppresses Alu hyper-editing, a function required for gastrulation and adult stem-cell differentiation in primates but not mice. Mechanistically, ILF2/3 directly binds Alu elements in nascent RNAs and shields them from ADAR1-mediated adenosine-to-inosine (A-to-I) editing. Acute ILF2/3 degradation increases A-to-I editing at Alu elements, but not murine retrotransposons, leading to aberrant splicing and nonsense-mediated decay of transcripts encoding key chromatin regulators in primate cells. This in turn destabilizes the epigenetic landscape and blocks lineage commitment across all three germ layers. Re-expression of correctly spliced chromatin regulators rescues differentiation defects in ILF2/3-deficient cells, functionally linking Alu editing control to chromatin regulation and cell fate. These findings define an evolutionary mechanism that restrains retrotransposon-associated RNA editing to preserve proteome integrity and enable primate-specific developmental programs.

CRISPy-web is an interactive web-based platform designed for rational guide RNA design in CRISPR-Cas9-based genome editing systems. Here, we present CRISPy-web 3.0 (https://crispy.secondarymetabolites.org/), an upgraded and versatile version that extends its functionality beyond classical Cas9-based systems. This version integrates support for diverse genome editing systems, including Cas9, CRISPR interference (CRISPRi), and TnpB/ωRNA. The redesigned interface enables users to toggle between multiple editing modes, select target regions such as ORFs or 5' UTRs, and visualize strand orientation, off-targets, and predicted mutation outcomes. It also incorporates scoring systems that evaluate guide RNA efficiency and specificity based on mismatch tolerance, positional context, and PAM requirements for CRISPR-Cas9-based applications. With an improved user interface design, enhanced backend scalability, and modular support for customizable inputs, CRISPy-web 3.0 provides a comprehensive and extensible platform for guide RNA design, enabling genome editing across a broader range of prokaryotic systems, including both CRISPR and TnpB.