🧬 PubMed RNA Editing Daily Digest

The clustered regularly interspaced short palindromic repeats CRISPR-associated protein 9 (CRISPR/Cas9) system has emerged as a versatile platform for genome editing, transcriptional regulation, and chromosomal imaging. Recent advances in synthetic biology have enabled the engineering of single guide RNA (sgRNA) to confer conditional responsiveness on the CRISPR/Cas9 system. By integrating functional nucleic acid elements, such as aptamers, ribozymes, and aptazymes, into specific structural regions of the sgRNA, researchers have developed systems that respond to a variety of molecular signals, including small molecules, proteins, and endogenous metabolites. These engineered sgRNAs enable spatiotemporal control of gene editing, activation, repression, and imaging in both prokaryotic and eukaryotic cells. This review summarizes the structural principles, design strategies, and applications of condition-responsive CRISPR/Cas9 systems, highlighting their potential in synthetic biology, disease modeling, and therapeutic development. Current challenges and future directions for improving the specificity, efficiency, and applicability of these systems are also discussed.

Lipid nanoparticles (LNPs) have emerged as a leading nonviral delivery system for ribonucleic acid (RNA)-based therapeutics, offering a scalable and efficient platform for both prophylactic and therapeutic applications. Compared to traditional excipients and delivery methods, LNPs provide several distinct advantages that make them particularly well-suited for nucleic acid delivery. These include enhanced cellular uptake, potential for reduced systemic toxicity, organ-specific biodistribution and protection of RNA cargo from enzymatic degradation. LNPs also facilitate endosomal escape, a critical barrier in the intracellular delivery of RNA molecules, thereby improving the overall transfection efficiency and therapeutic index. Their tunable composition allows for precise control over pharmacokinetics, immunogenicity and tissue targeting, which are critical in the development of vaccines, gene therapies, other RNA-based interventions and precision medicines. In addition, LNP-based nonviral delivery methods may offer several advantages over viral vectors, including cost-effectiveness, scalable and rapid manufacturing and greater adaptability for developing personalized gene therapy products. These features align well with platform technology designations and support a bespoke, umbrella trial approach. This review provides a comprehensive overview of the critical components and design considerations of LNP-based delivery systems, with a focus on messenger RNA, guide RNA payloads and lipid components. We discuss the differences between prophylactic and therapeutic formulations, key quality and purity attributes required for safe and effective use. Particular attention is given to the complexities of LNP formulation, including the characterization, production and quality control of both RNA cargo and lipid components. We also highlight current regulatory challenges, including the lack of standardized testing methods, unclear excipient classification and the absence of compendial monographs for the ionizable and pegylated lipid components. Essential questions for optimizing LNP formulations are discussed along with the evolving regulatory landscape and current clinical applications. As the field evolves to incorporate novel RNA modalities such as short-interference RNA, RNAi, self-replicating RNA and circular RNAs, the need for robust quality standards and regulatory clarity becomes more vital. This review underscores the urgent need for collaboration between industry and regulatory to establish harmonized framework for LNP development to ensure safety, efficacy and rapid translation of RNA-LNP therapeutics across diverse disease areas.

Maintaining genome integrity is essential, yet how DNA repair is balanced across life stages remains poorly understood. Here we uncover an epitranscriptomic mechanism in the fungal pathogen Fusarium graminearum that alleviates a trade-off between heat-stress adaptation and sexual reproduction. We show that FgMus81 acts independently of its nuclease activity and canonical partner FgMms4, and has dosage-dependent, stage-specific functions: restrained levels support meiosis, whereas elevated levels promote heat-stress survival. We identify a sexual stage-specific A-to-I RNA editing event that recodes FgMus81 (N420D) and tunes its abundance to meet meiotic demands without compromising stress resilience. Notably, both pre-editing and post-editing isoforms support meiotic interhomolog crossovers, but the post-editing isoform impairs mitotic recombination. Conservation of this editing across Sordariomycetes suggests evolutionary selection for stage-specific control. Together, these findings reveal an epitranscriptomic switch that partitions Mus81 functions across life stages and identify adaptive RNA editing as a regulator of homologous recombination in fungal pathogens.

Functional mitochondrial mRNAs in Trypanosoma brucei are generated by the post-transcriptional guide RNA (gRNA) directed insertion and deletion of uridine residues, called RNA editing, that is catalyzed by three closely related multiprotein RNA Editing Catalytic Complexes (RECCs). These RECCs contain a common set of 12 proteins including KREPA6 which is largely comprised of an oligonucleotide binding (OB)-fold domain with a predicted intrinsically disordered region (IDR) at its C-terminus. Here we show that certain single amino acid substitutions throughout KREPA6 or deletion of the IDR inhibit the growth and viability of bloodstream form (BF) parasites. These mutations variously impact RECC structure, many alter but do not eliminate RNA editing, and some result in differential utilization of gRNAs. The results indicate that KREPA6 protein has multiple functions some of which stem from its interactions with multiple RECC proteins and perhaps with substrate RNA in each of the three different RECCs. These functions likely involve dynamic interactions of KREPA6 with key domains of other RECC proteins, other editing proteins, and with messenger RNA/gRNA substrates during the multiple catalytic and noncatalytic steps that occur during the complicated editing process.