{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,29]],"date-time":"2026-08-29T15:59:03Z","timestamp":1788019143528,"version":"build-2784847793"},"reference-count":31,"publisher":"S. Karger AG","issue":"5","license":[{"start":{"date-parts":[[2025,2,21]],"date-time":"2025-02-21T00:00:00Z","timestamp":1740096000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/karger.com\/pages\/terms-and-conditions"},{"start":{"date-parts":[[2025,2,21]],"date-time":"2025-02-21T00:00:00Z","timestamp":1740096000000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/karger.com\/pages\/terms-and-conditions"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Horm Res Paediatr"],"abstract":"<jats:p>Introduction: Non-autoimmune hyperthyroidism (NAH) is a rare genetic disorder caused by germline-activating variants in the TSH receptor (TSHR) gene. While most NAH-related TSHR variants are located in the seven-transmembrane domain (7TMD), three variants (p.Leu267Phe, p.Ser281Asn, and p.Asn406Ser) have been identified in the extracellular domain (ECD), with p.Leu267Phe previously not showing constitutively active in vitro. Methods: We searched for TSHR variants in a Japanese family with NAH using PCR-based direct sequencing. We created three ECD variants (p.Leu267Phe, p.Ser281Asn, and p.Asn406Ser) and a series of variants in which Leu267 was mutated to 18 amino acids other than Leu and Phe. Based on the cryo-electron microscopic structures, we evaluated the structure-function relationship of TSHR. We compared their cAMP-producing capacities to WT-TSHR in the luciferase activity assay using HEK293 cells. Western blot and fluorescence immunostaining were performed using HA-tagged TSHR vectors to compare Leu267Phe-TSHR with WT-TSHR. Results: A heterozygous TSHR variant (p.Leu267Phe) was identified. Comparison of the cryo-electron microscopy structures of the activated and inactivated TSHRs revealed a significant change in the ECD structure around Leu267. The ligand-independent cAMP-producing capacities compared to WT-TSHR were 238 \u00b1 20% (mean \u00b1 SEM) for Leu267Phe. Of all 19 possible variants created through systematic mutagenesis, only Leu267Phe-TSHR and Leu267Tyr-TSHR exhibited significantly higher ligand-independent cAMP-producing capacities. Immunoblotting and fluorescence immunostaining showed that the Leu267Phe variant did not affect protein expression levels and intracellular localization of TSHR. Discussion\/Conclusion: Leu267Phe-TSHR causes NAH. Substituting Leu267 to aromatic amino acids may shift the equilibrium of the TSHR state toward activation.<\/jats:p>","DOI":"10.1159\/000544836","type":"journal-article","created":{"date-parts":[[2025,2,21]],"date-time":"2025-02-21T12:02:58Z","timestamp":1740139378000},"page":"623-631","source":"Crossref","is-referenced-by-count":1,"title":["Familial Non-Autoimmune Hyperthyroidism Caused by an Extracellular Domain Variant (p.Leu267Phe) of the TSH Receptor"],"prefix":"10.1159","volume":"99","author":[{"given":"Kazuhiro","family":"Shimura","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yosuke","family":"Ichihashi","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kiyomi","family":"Abe","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7360-2465","authenticated-orcid":false,"given":"Tomohiro","family":"Ishii","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tomonobu","family":"Hasegawa","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Satoshi","family":"Narumi","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"127","published-online":{"date-parts":[[2025,2,21]]},"reference":[{"key":"ref1","doi-asserted-by":"publisher","DOI":"10.1159\/000342982"},{"key":"ref2","doi-asserted-by":"publisher","DOI":"10.1530\/acta.0.1000512"},{"key":"ref3","doi-asserted-by":"publisher","DOI":"10.1007\/BF03345784"},{"key":"ref4","doi-asserted-by":"publisher","DOI":"10.1089\/thy.2000.10.859"},{"key":"ref5","doi-asserted-by":"publisher","DOI":"10.1507\/endocrj.k06-090"},{"key":"ref6","doi-asserted-by":"publisher","DOI":"10.4158\/EP.14.4.479"},{"key":"ref7","doi-asserted-by":"publisher","DOI":"10.1186\/1756-6614-4-S1-S8"},{"key":"ref8","doi-asserted-by":"publisher","DOI":"10.1210\/edrv-13-3-596"},{"key":"ref9","doi-asserted-by":"publisher","DOI":"10.1210\/er.2008-0044"},{"key":"ref10","doi-asserted-by":"publisher","DOI":"10.1038\/ng0794-396"},{"key":"ref11","doi-asserted-by":"publisher","DOI":"10.1089\/thy.2013.0431"},{"key":"ref12","doi-asserted-by":"publisher","DOI":"10.1016\/s0014-5793(97)00532-2"},{"key":"ref13","doi-asserted-by":"publisher","DOI":"10.1210\/jcem.83.5.4776"},{"key":"ref14","doi-asserted-by":"publisher","DOI":"10.1111\/j.1365-2265.2011.04324.x"},{"key":"ref15","doi-asserted-by":"publisher","DOI":"10.4158\/ACCR-2019-0361"},{"key":"ref16","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M604770200"},{"key":"ref17","doi-asserted-by":"publisher","DOI":"10.1210\/jc.2008-1767"},{"key":"ref18","doi-asserted-by":"publisher","DOI":"10.1038\/s41586-022-05159-1"},{"key":"ref19","doi-asserted-by":"publisher","DOI":"10.1210\/clinem\/dgaa772"},{"key":"ref20","doi-asserted-by":"publisher","DOI":"10.1530\/EJE-16-1049"},{"key":"ref21","doi-asserted-by":"publisher","DOI":"10.1210\/jcem.86.9.7888"},{"key":"ref22","doi-asserted-by":"publisher","DOI":"10.1210\/jcem.81.6.8964822"},{"key":"ref23","doi-asserted-by":"publisher","DOI":"10.1111\/j.1365-2265.2004.02040.x"},{"key":"ref24","doi-asserted-by":"publisher","DOI":"10.4274\/jcrpe.v2i4.168"},{"key":"ref25","doi-asserted-by":"publisher","DOI":"10.1159\/000023312"},{"key":"ref26","doi-asserted-by":"publisher","DOI":"10.1089\/thy.2006.16.505"},{"key":"ref27","doi-asserted-by":"publisher","DOI":"10.1007\/s00431-007-0659-9"},{"key":"ref28","doi-asserted-by":"publisher","DOI":"10.1038\/pr.2014.34"},{"key":"ref29","doi-asserted-by":"publisher","DOI":"10.1210\/en.2003-0438"},{"key":"ref30","doi-asserted-by":"publisher","DOI":"10.7554\/eLife.50279"},{"key":"ref31","doi-asserted-by":"publisher","DOI":"10.1210\/en.2008-1800"}],"container-title":["Hormone Research in Paediatrics"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/karger.com\/hrp\/article-pdf\/doi\/10.1159\/000544836\/4342316\/000544836.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/karger.com\/article-pdf\/doi\/10.1159\/000544836","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,7,13]],"date-time":"2026-07-13T16:01:07Z","timestamp":1783958467000},"score":1,"resource":{"primary":{"URL":"https:\/\/karger.com\/article\/doi\/10.1159\/000544836"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,2,21]]},"references-count":31,"journal-issue":{"issue":"5","published-online":{"date-parts":[[2026,7,13]]},"published-print":{"date-parts":[[2026,7,22]]}},"URL":"https:\/\/doi.org\/10.1159\/000544836","archive":["Portico"],"relation":{},"ISSN":["1663-2818","1663-2826"],"issn-type":[{"value":"1663-2818","type":"print"},{"value":"1663-2826","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,2,21]]}}}