{"doi":"10.1016/j.ceca.2025.103081","title":"CPVT1 point mutations in RyR2 S5 and S6 segments and their Ca2+ signaling consequence","abstract":"• RyR-R4822H mutation located at S5-S6 luminal loop and RyR-L4865V mutation located at S6 segment are identified to associated with CPVT1. • RyR2-R4822H mutation shows deficient CICR, and caffeine triggered Ca 2+ release while L4865V mutation doesn't affect Ca 2+ signaling significantly. • Spontaneous beating persists in R4822H expressing hiPSC CMs even though SR Ca 2+ release is completely suppressed, suggestive of cardiac EC-coupling remodeling. • Spontaneous Ca 2+ spark igniting frequency in both R4822H and L4865V mutant myocytes were like those measured in WT myocytes, but their spark durations were significantly shorter. • Even though both R4822H and L4865V mutations are located at the transmembrane hot spot of CPVT1 mutations, their Ca 2+ signaling phenotypes differ greatly. Precise activation of cardiac ryanodine receptor (RyR2) by small influx of Ca 2+ during the action potential triggers the release of SR Ca 2+ that activates contraction, a process known as Ca 2+ -induced Ca 2+ release (CICR). Missense mutations in RyR2 often cause aberrant and unregulated Ca 2+ releases that are associated with catecholaminergic polymorphic ventricular tachycardia (CPVT), often lethal arrhythmias. Here using CRISPR/Cas9 gene editing in human induced pluripotent stem cells (hiPSCs), we extended our previous studies to include two new arrhythmogenic mutations one, R4822H, located in S5-S6 transmembrane luminal loop near RyR2 selective filter and the other, L4865V, located on S6 segment. TIRF-imaging of voltage-clamped mutant myocytes showed that I Ca and caffeine-triggered cytosolic Ca 2+ rise (Fura-2 signal) or ER-GCaMP6 SR Ca 2+ release signals were significantly suppressed in R4822H but not in L4865V myocytes. Spontaneous Ca 2+ transients, however, persisted in both mutant lines activating both Fura-2 and ER-GCaMP6 Ca 2+ transients in L4865V cells, but only Fura-2 Ca 2+ transients in R4822H mutant. Spontaneous Ca 2+ sparks igniting frequencies were similar in both mutants, but spark durations were significantly shorter. Although both of these mutations are located at S5 and S6 transmembrane regions of RyR2, their phenotypes diverge markedly. L4865V mutant does not show suppressed E-C coupling function, while R4822H mutant has completely suppressed CICR suggesting that the spontaneous beating in R4822H mutant results from remodeling of dormant Ca 2+ signaling pathway expressed in hiPSC CMs.","journal":"Cell Calcium","year":2025,"id":547035,"datarank":0.10397207708399181,"base_score":0.6931471805599453,"endowment":0.6931471805599453,"self_citation_contribution":0.10397207708399181,"citation_network_contribution":0.0,"self_endowment_contribution":0.10397207708399181,"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":0.9491,"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":1439472,"name":"Grace Ellen Donch","orcid":null,"position":1,"is_corresponding":false},{"id":533148,"name":"Naohiro Yamaguchi","orcid":"0000-0001-5933-1507","position":2,"is_corresponding":false},{"id":392358,"name":"Martin Morad","orcid":"0000-0002-9054-0195","position":3,"is_corresponding":false},{"id":805151,"name":"Xiaohua Zhang","orcid":"0000-0003-0102-6352","position":0,"is_corresponding":true}],"reference_count":27,"raw_metadata":{"citation_network_status":"fetched"},"created_at":"2026-07-19T02:53:36.567932Z","pmid":"40974955","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":[]}