🧬 PubMed RNA Editing Daily Digest

R-loops play critical roles in gene regulation and genome stability. To elucidate the protein-mediated mechanisms governing R-loop dynamics, we leveraged APEX-based proximity labeling to profile the R-loop proximal proteome. By integrating the proteomic data, bioinformatic predictions, and biochemical characterizations, we identified YY1 as an R-loop-binding protein. Localized R-loop induction at specific genomic loci via a CRISPR-dCas9/gRNA targeting system enhanced the local YY1 occupancy. Furthermore, integrated ChIP-seq and RNA-seq analyses revealed that promoter-associated YY1-R-loop interactions broadly regulate transcription. Together, our work identifies YY1 as a novel R-loop interaction protein and establishes a role for this interaction in modulating gene expression.

RNA editing is a post-transcriptional modification that can have important effects on mRNA translation and, thereby, on the composition of the proteome. RNA editing exhibits complex responses to temperature and thus may be important in ectothermic species' capacities to cope with thermal stress. To investigate the temperature-dependent patterns and potential functions of RNA editing events in intertidal mollusks, we conducted a genome-wide identification and analysis of RNA editing sites in the mussel Mytilus galloprovincialis exposed to various temperatures. Our results showed that the number of sites where adenosine (A) was converted to inosine (I) decreased significantly with increasing temperatures and correlated positively with the expression level of the enzyme adenosine deaminase acting on RNA. Furthermore, the shared edited genes among all temperatures were enriched significantly in some key biological processes (e.g., negative regulation of apoptotic processes), and the unique edited genes at the sublethal temperature of 31°C were enriched significantly in some cellular signaling pathways (e.g., GTPase activator activity). Luciferase assays showed that protein translation efficiency was positively correlated with the number of edited A-to-I sites in the 3' untranslated region (3'UTR). Our study demonstrates that temperature-sensitive A-to-I editing events may play an important role in allowing mollusks like M. galloprovincialis to rapidly acclimatize to stressful thermal environments.

The growing prevalence of gluten-related disorders has driven the development of wheat varieties with reduced immunogenic gluten. This study aimed to integrate RNA interference (RNAi) and CRISPR genome editing within a doubled haploid (DH) platform to overcome challenges of gene redundancy and polyploidy in wheat gliadins. We generated DH lines from crosses between RNAi and CRISPR lines and elite wheat cultivars, enabling stable fixation of multiple genetic modifications in a single generation. Deep sequencing analysis of α-gliadin amplicons was conducted using a custom bioinformatics pipeline optimized for complex, repetitive gene families. Gluten protein profiles were evaluated using RP-HPLC and R5 monoclonal antibody. Several DH lines presented over 70% reduction in immunogenic epitopes in α-gliadins, with lines outperforming both parents. Editing frequency was influenced by single-guide RNA efficiency and parental background. Silencing and editing combined led to nearly depleted gliadins in some lines, often with compensatory increases in other storage proteins linked with bread-making quality, such as high-molecular-weight glutenin subunits. Kernel and specific weight traits were largely maintained. This work demonstrates that combining RNAi and CRISPR in a DH platform enables efficient, heritable reduction of immunogenic gluten, providing a viable strategy for breeding wheat lines safer for individuals with gluten-related disorders.

Genome editing techniques, especially clustered regularly interspaced short palindromic repeats (CRISPR), brought researchers into a new era of molecular plant breeding because it enabled them to make targeted modifications in plant genomes and transcriptomes. However, the successful incorporation of large DNA segments into plant genomes, necessary for high genetic gains and desired traits, remains a critical challenge. As there is an increasing demand for technologies that support chromosomal integration of large DNA inserts suitable for application in synthetic biology and plant breeding, PrimeRoot editors have presented a revolutionary solution to this issue, as they integrate enhanced prime editing guide RNA (PegRNA) designs, improved plant prime editor systems and advanced recombinases, helping in the precise insertion of DNA fragments of up to 11.1 kb into the plant genomes. Third-generation PrimeRoot editors further enhanced the precision and efficiency of transformation under different gene delivery systems. This technology holds enormous promise for accurately inserting long DNA sequences across different species of plants. This review highlights expected developments, opportunities, applications, advantages and challenges associated with PrimeRoot editors as well as the significance of expanding their applicability to more varieties of plants.