{"doi":"10.1002/cbic.202500735","title":"Discovery and Tuning of RNA Editing Guides via High‐Throughput Screening and Chemical Modification","abstract":"Adenosine deaminases acting on RNA (ADAR) catalyze the deamination of adenosine to inosine in double-stranded RNA. Because inosine is read as guanosine during translation, this process enables programmable A-to-G recoding at the transcript level. ADARs can be harnessed for therapeutic correction of pathogenic mutations through site-directed RNA editing with guide RNAs. To expand the design space of editing-enabling guides, we applied EMERGe, a high-throughput screening platform, to identify motifs targeting a premature termination codon in the MeCP2 transcript associated with Rett syndrome. This uncovered a guide RNA motif that supported efficient ADAR2-mediated editing in vitro, featuring a 5'-GUG-3' sequence predicted to form an asymmetric loop. To enable therapeutic application, structure-activity relationship studies and chemical optimization were performed, yielding a fully modified guide RNA with 2'-O-methyl, 2'-fluoro, and phosphorothioate linkages. This stabilized guide retained the activity of unmodified RNA and showed enhanced nuclease resistance. The optimized guide induces dose-dependent editing at two MECP2 loci in reporter assays in HEK293T cells, demonstrating that EMERGe-selected motifs can be rendered viable in cells through targeted chemical modification. These findings highlight the utility of EMERGe as a discovery platform and establish a pipeline for identifying and optimizing editing-enabling guide RNA features beyond traditional design rules.","journal":"ChemBioChem","year":2025,"id":548809,"datarank":0.0,"base_score":0.0,"endowment":0.0,"self_citation_contribution":0.0,"citation_network_contribution":0.0,"self_endowment_contribution":0.0,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":1,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9498,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2025-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":1442497,"name":"S. Lin","orcid":"0009-0002-4475-8289","position":1,"is_corresponding":false},{"id":1442498,"name":"Megan M. Chinn","orcid":"0009-0007-1594-3609","position":2,"is_corresponding":false},{"id":884767,"name":"Victorio Jauregui Matos","orcid":"0000-0003-2009-5521","position":3,"is_corresponding":false},{"id":884766,"name":"Aashrita Manjunath","orcid":"0009-0007-9999-6176","position":4,"is_corresponding":false},{"id":788936,"name":"I.V. Yang","orcid":"0009-0002-9492-4134","position":5,"is_corresponding":false},{"id":1085908,"name":"Casey S. Jacobsen","orcid":"0000-0002-0249-1398","position":6,"is_corresponding":false},{"id":245813,"name":"Peter A. Beal","orcid":"0000-0003-4855-7185","position":7,"is_corresponding":false},{"id":1085909,"name":"Prince J. Salvador","orcid":"0000-0003-4536-584X","position":0,"is_corresponding":true}],"reference_count":32,"raw_metadata":null,"created_at":"2026-07-19T02:54:03.053965Z","pmid":"41272938","pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}