{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,3]],"date-time":"2026-09-03T13:23:13Z","timestamp":1788441793096,"version":"build-2803163510"},"reference-count":58,"publisher":"Ovid Technologies (Wolters Kluwer Health)","issue":"23","content-domain":{"domain":["www.ahajournals.org"],"crossmark-restriction":true},"short-container-title":["Circulation"],"published-print":{"date-parts":[[2020,12,8]]},"abstract":"<jats:sec>\n            <jats:title>Background:<\/jats:title>\n            <jats:p>\n              Pathogenic\n              <jats:italic toggle=\"yes\">TNNT2<\/jats:italic>\n              variants are a cause of hypertrophic and dilated cardiomyopathies, which promote heart failure by incompletely understood mechanisms. The precise functional significance for 87% of\n              <jats:italic toggle=\"yes\">TNNT2<\/jats:italic>\n              variants remains undetermined, in part, because of a lack of functional genomics studies. The knowledge of which and how\n              <jats:italic toggle=\"yes\">TNNT2<\/jats:italic>\n              variants cause hypertrophic and dilated cardiomyopathies could improve heart failure risk determination, treatment efficacy, and therapeutic discovery, and provide new insights into cardiomyopathy pathogenesis, as well.\n            <\/jats:p>\n          <\/jats:sec>\n          <jats:sec>\n            <jats:title>Methods:<\/jats:title>\n            <jats:p>\n              We created a toolkit of human induced pluripotent stem cell models and functional assays using CRISPR\/Cas9 to study\n              <jats:italic toggle=\"yes\">TNNT2<\/jats:italic>\n              variant pathogenicity and pathophysiology. Using human induced pluripotent stem cell\u2013derived cardiomyocytes in cardiac microtissue and single-cell assays, we functionally interrogated 51\n              <jats:italic toggle=\"yes\">TNNT2<\/jats:italic>\n              variants, including 30 pathogenic\/likely pathogenic variants and 21 variants of uncertain significance. We used RNA sequencing to determine the transcriptomic consequences of pathogenic\n              <jats:italic toggle=\"yes\">TNNT2<\/jats:italic>\n              variants and adapted CRISPR\/Cas9 to engineer a transcriptional reporter assay to assist prediction of\n              <jats:italic toggle=\"yes\">TNNT2<\/jats:italic>\n              variant pathogenicity. We also studied variant-specific pathophysiology using a thin filament\u2013directed calcium reporter to monitor changes in myofilament calcium affinity.\n            <\/jats:p>\n          <\/jats:sec>\n          <jats:sec>\n            <jats:title>Results:<\/jats:title>\n            <jats:p>\n              Hypertrophic cardiomyopathy\u2013associated\n              <jats:italic toggle=\"yes\">TNNT2<\/jats:italic>\n              variants caused increased cardiac microtissue contraction, whereas dilated cardiomyopathy\u2013associated variants decreased contraction.\n              <jats:italic toggle=\"yes\">TNNT2<\/jats:italic>\n              variant\u2013dependent changes in sarcomere contractile function induced graded regulation of 101 gene transcripts, including MAPK (mitogen-activated protein kinase) signaling targets,\n              <jats:italic toggle=\"yes\">HOPX<\/jats:italic>\n              , and\n              <jats:italic toggle=\"yes\">NPPB<\/jats:italic>\n              . We distinguished pathogenic\n              <jats:italic toggle=\"yes\">TNNT2<\/jats:italic>\n              variants from wildtype controls using a sarcomere functional reporter engineered by inserting tdTomato into the endogenous\n              <jats:italic toggle=\"yes\">NPPB<\/jats:italic>\n              locus. On the basis of a combination of\n              <jats:italic toggle=\"yes\">NPPB<\/jats:italic>\n              reporter activity and cardiac microtissue contraction, our study provides experimental support for the reclassification of 2 pathogenic\/likely pathogenic variants and 2 variants of uncertain significance.\n            <\/jats:p>\n          <\/jats:sec>\n          <jats:sec>\n            <jats:title>Conclusions:<\/jats:title>\n            <jats:p>\n              Our study found that hypertrophic cardiomyopathy\u2013associated\n              <jats:italic toggle=\"yes\">TNNT2<\/jats:italic>\n              variants increased cardiac microtissue contraction, whereas dilated cardiomyopathy\u2013associated variants decreased contraction, both of which paralleled changes in myofilament calcium affinity. Transcriptomic changes, including\n              <jats:italic toggle=\"yes\">NPPB<\/jats:italic>\n              levels, directly correlated with sarcomere function and can be used to predict\n              <jats:italic toggle=\"yes\">TNNT2<\/jats:italic>\n              variant pathogenicity.\n            <\/jats:p>\n          <\/jats:sec>","DOI":"10.1161\/circulationaha.120.047999","type":"journal-article","created":{"date-parts":[[2020,10,7]],"date-time":"2020-10-07T09:00:13Z","timestamp":1602061213000},"page":"2262-2275","update-policy":"https:\/\/doi.org\/10.1161\/crossmarkpolicy","source":"Crossref","is-referenced-by-count":60,"title":["Development of a Cardiac Sarcomere Functional Genomics Platform to Enable Scalable Interrogation of Human\n            <i>TNNT2<\/i>\n            Variants"],"prefix":"10.1161","volume":"142","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-5623-8315","authenticated-orcid":true,"given":"Anthony M.","family":"Pettinato","sequence":"first","affiliation":[{"name":"University of Connecticut Health Center (A.M.P., F.A.L., R.R., J.T.H.)."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Feria A.","family":"Ladha","sequence":"additional","affiliation":[{"name":"University of Connecticut Health Center (A.M.P., F.A.L., R.R., J.T.H.)."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6606-0274","authenticated-orcid":true,"given":"David J.","family":"Mellert","sequence":"additional","affiliation":[{"name":"The Jackson Laboratory for Genomic Medicine (D.J.M., N.L., R.C., K.T., Y.-S.C., J.T.H.)"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nicholas","family":"Legere","sequence":"additional","affiliation":[{"name":"The Jackson Laboratory for Genomic Medicine (D.J.M., N.L., R.C., K.T., Y.-S.C., J.T.H.)"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rachel","family":"Cohn","sequence":"additional","affiliation":[{"name":"The Jackson Laboratory for Genomic Medicine (D.J.M., N.L., R.C., K.T., Y.-S.C., J.T.H.)"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Robert","family":"Romano","sequence":"additional","affiliation":[{"name":"University of Connecticut Health Center (A.M.P., F.A.L., R.R., J.T.H.)."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ketan","family":"Thakar","sequence":"additional","affiliation":[{"name":"The Jackson Laboratory for Genomic Medicine (D.J.M., N.L., R.C., K.T., Y.-S.C., J.T.H.)"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7785-4181","authenticated-orcid":true,"given":"Yu-Sheng","family":"Chen","sequence":"additional","affiliation":[{"name":"The Jackson Laboratory for Genomic Medicine (D.J.M., N.L., R.C., K.T., Y.-S.C., J.T.H.)"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1455-113X","authenticated-orcid":true,"given":"J. Travis","family":"Hinson","sequence":"additional","affiliation":[{"name":"University of Connecticut Health Center (A.M.P., F.A.L., R.R., J.T.H.)."},{"name":"The Jackson Laboratory for Genomic Medicine (D.J.M., N.L., R.C., K.T., Y.-S.C., J.T.H.)"},{"name":"Calhoun Cardiology Center, UConn Health (J.T.H.), 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