{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,2,26]],"date-time":"2026-02-26T15:34:33Z","timestamp":1772120073002,"version":"3.50.1"},"reference-count":35,"publisher":"Wiley","issue":"1","license":[{"start":{"date-parts":[[2020,3,1]],"date-time":"2020-03-01T00:00:00Z","timestamp":1583020800000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"},{"start":{"date-parts":[[2020,3,1]],"date-time":"2020-03-01T00:00:00Z","timestamp":1583020800000},"content-version":"tdm","delay-in-days":0,"URL":"http:\/\/www.springer.com\/tdm"}],"funder":[{"DOI":"10.13039\/100000002","name":"National Institutes of Health","doi-asserted-by":"publisher","award":["1R01\u2010GM106107"],"award-info":[{"award-number":["1R01\u2010GM106107"]}],"id":[{"id":"10.13039\/100000002","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["link.springer.com"],"crossmark-restriction":false},"short-container-title":["Quant. Biol."],"published-print":{"date-parts":[[2020,3]]},"abstract":"<jats:sec>\n                    <jats:title>Background<\/jats:title>\n                    <jats:p>E2F1 protein, a major effector of the Rb\/E2F pathway plays a central role in regulating cell\u2010fate decisions involved in proliferation, apoptosis, and differentiation. Its expression is highly dynamic and tightly modulated through a combination of transcriptional, translational and posttranslational controls. However, the mechanisms by which its expression and activity can promote different cellular outcomes remain to be fully elucidated. To better document E2F1 expression in live cells, we have engineered a series of fluorescent E2F1 protein reporters that quantitatively capture E2F1 protein dynamics.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>\n                      Reporter constructs, under the control of the mouse or human E2F1 proximal promoter, were designed to express an E2F1\u2010Venus fusion protein incapable of binding DNA. In addition, constructs either included or excluded the 3\u2032 untranslated region (3\u2032UTR) of the\n                      <jats:italic>E2F1<\/jats:italic>\n                      gene. These constructs were introduced into fibroblasts and epithelial cells, and expression of the fusion reporter protein was validated and quantified in single cells using live imaging.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>\n                      In all cases, expression of the reporter protein effectively recapitulated the behavior of E2F1 under various conditions, including cell cycle progression and genotoxic stress. No or little fluorescent signal of the reporter was detected in G\n                      <jats:sub>0<\/jats:sub>\n                      , but as the cycle progressed, expression of the reporter protein steadily increased in the nucleus, peaking a few hours before cell division, but declining to baseline 2\u20133 h prior to the onset of mitosis. The absence of the E2F1 3\u2032UTR in the constructs led to considerably higher steady\u2010state levels of the fusion protein, which although normally regulated, exhibited a slightly less complex dynamic profile during the cell cycle or genotoxic stress. Lastly, the presence or absence of Rb failed to impact the overall detection and levels of the reporter proteins.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusions<\/jats:title>\n                    <jats:p>Our validated E2F1 protein reporters complement nicely other reporters of the Rb\/E2F pathway and provide a unique tool to follow the complex dynamics of E2F1 expression in real time in single cells.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.1007\/s40484-019-0193-6","type":"journal-article","created":{"date-parts":[[2020,3,5]],"date-time":"2020-03-05T23:51:26Z","timestamp":1583452286000},"page":"20-30","update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Quantifying E2F1 protein dynamics in single cells"],"prefix":"10.1002","volume":"8","author":[{"given":"Bernard","family":"Mathey\u2010Prevot","sequence":"first","affiliation":[{"name":"<!--1--> Department of Pharmacology and Cancer Biology Duke University School of Medicine Durham NC 27710 USA"},{"name":"<!--7--> Department of Pediatrics Duke University School of Medicine Durham NC 27708 USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Bao\u2010Tran","family":"Parker","sequence":"additional","affiliation":[{"name":"<!--1--> Department of Pharmacology and Cancer Biology Duke University School of Medicine Durham NC 27710 USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Carolyn","family":"Im","sequence":"additional","affiliation":[{"name":"<!--1--> Department of Pharmacology and Cancer Biology Duke University School of Medicine Durham NC 27710 USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Cierra","family":"Hong","sequence":"additional","affiliation":[{"name":"<!--2--> Duke University School of Medicine Durham NC 27710 USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Peng","family":"Dong","sequence":"additional","affiliation":[{"name":"<!--1--> Department of Pharmacology and Cancer Biology Duke University School of Medicine Durham NC 27710 USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Guang","family":"Yao","sequence":"additional","affiliation":[{"name":"<!--3--> Molecular and Cellular Biology University of Arizona Tucson AZ 85721 USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Lingchong","family":"You","sequence":"additional","affiliation":[{"name":"<!--4--> Department of Biomedical Engineering Duke University Durham NC 27705 USA"},{"name":"<!--5--> Center for Genomic and Computational Biology Duke University Durham NC 27705 USA"},{"name":"<!--6--> Department of Molecular Genetics and Microbiology Duke University School of Medicine Durham NC 27708 USA"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2020,3]]},"reference":[{"key":"e_1_2_9_2_2","doi-asserted-by":"publisher","DOI":"10.1080\/10409238.2017.1360836"},{"key":"e_1_2_9_3_2","doi-asserted-by":"publisher","DOI":"10.1038\/nrm3629"},{"key":"e_1_2_9_4_2","doi-asserted-by":"publisher","DOI":"10.1101\/gad.949802"},{"key":"e_1_2_9_5_2","doi-asserted-by":"publisher","DOI":"10.1038\/nrm2469"},{"key":"e_1_2_9_6_2","doi-asserted-by":"publisher","DOI":"10.1186\/1747\u20101028\u20107\u201010"},{"key":"e_1_2_9_7_2","doi-asserted-by":"publisher","DOI":"10.1101\/gad.1256504"},{"key":"e_1_2_9_8_2","doi-asserted-by":"publisher","DOI":"10.1038\/nrc2696"},{"key":"e_1_2_9_9_2","doi-asserted-by":"publisher","DOI":"10.1038\/ncomms5750"},{"key":"e_1_2_9_10_2","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0185637"},{"key":"e_1_2_9_11_2","doi-asserted-by":"publisher","DOI":"10.1038\/sj.onc.1208612"},{"key":"e_1_2_9_12_2","doi-asserted-by":"publisher","DOI":"10.1128\/MCB.20.1.273\u2010285.2000"},{"key":"e_1_2_9_13_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.celrep.2019.05.004"},{"key":"e_1_2_9_14_2","doi-asserted-by":"publisher","DOI":"10.1038\/nature03677"},{"key":"e_1_2_9_15_2","doi-asserted-by":"publisher","DOI":"10.1038\/onc.2008.372"},{"key":"e_1_2_9_16_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.ceb.2007.10.006"},{"key":"e_1_2_9_17_2","doi-asserted-by":"publisher","DOI":"10.4161\/23723548.2014.970480"},{"key":"e_1_2_9_18_2","doi-asserted-by":"publisher","DOI":"10.1101\/gad.13.6.666"},{"key":"e_1_2_9_19_2","doi-asserted-by":"publisher","DOI":"10.1038\/onc.2011.603"},{"key":"e_1_2_9_20_2","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/15.20.8125"},{"key":"e_1_2_9_21_2","doi-asserted-by":"publisher","DOI":"10.4161\/cc.10.18.17350"},{"key":"e_1_2_9_22_2","first-page":"1833","article-title":"Selective induction of E2F1 in response to DNA damage, mediated by ATM\u2010dependent phosphorylation.","volume":"15","author":"Lin W. C.","year":"2001","journal-title":"Genes Dev."},{"key":"e_1_2_9_23_2","doi-asserted-by":"publisher","DOI":"10.1242\/jcs.109.7.1717"},{"key":"e_1_2_9_24_2","doi-asserted-by":"publisher","DOI":"10.1242\/jcs.110.22.2819"},{"key":"e_1_2_9_25_2","doi-asserted-by":"publisher","DOI":"10.4161\/cc.6.17.4650"},{"key":"e_1_2_9_26_2","doi-asserted-by":"publisher","DOI":"10.1101\/gad.10.23.2949"},{"key":"e_1_2_9_27_2","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.94.6.2221"},{"key":"e_1_2_9_28_2","doi-asserted-by":"publisher","DOI":"10.1038\/ncb2309"},{"key":"e_1_2_9_29_2","doi-asserted-by":"publisher","DOI":"10.1038\/cdd.2017.12"},{"key":"e_1_2_9_30_2","doi-asserted-by":"publisher","DOI":"10.1186\/gb\u20102011\u201012\u20109\u2010r96"},{"key":"e_1_2_9_31_2","doi-asserted-by":"publisher","DOI":"10.1016\/0092\u20108674(92)90107\u2010N"},{"key":"e_1_2_9_32_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2011.02.013"},{"key":"e_1_2_9_33_2","doi-asserted-by":"publisher","DOI":"10.1038\/sj.gt.3301206"},{"key":"e_1_2_9_34_2","doi-asserted-by":"publisher","DOI":"10.1038\/nmeth.3047"},{"key":"e_1_2_9_35_2","doi-asserted-by":"publisher","DOI":"10.1126\/science.1247005"},{"key":"e_1_2_9_36_2","doi-asserted-by":"publisher","DOI":"10.1016\/0014\u20104827(65)90211\u20109"}],"container-title":["Quantitative Biology"],"original-title":[],"language":"en","link":[{"URL":"http:\/\/link.springer.com\/content\/pdf\/10.1007\/s40484-019-0193-6.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"http:\/\/link.springer.com\/article\/10.1007\/s40484-019-0193-6\/fulltext.html","content-type":"text\/html","content-version":"vor","intended-application":"text-mining"},{"URL":"http:\/\/link.springer.com\/content\/pdf\/10.1007\/s40484-019-0193-6.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"similarity-checking"},{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/pdf\/10.1007\/s40484-019-0193-6","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,1,3]],"date-time":"2025-01-03T04:57:31Z","timestamp":1735880251000},"score":1,"resource":{"primary":{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/10.1007\/s40484-019-0193-6"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2020,3]]},"references-count":35,"journal-issue":{"issue":"1","published-print":{"date-parts":[[2020,3]]}},"alternative-id":["10.1007\/s40484-019-0193-6"],"URL":"https:\/\/doi.org\/10.1007\/s40484-019-0193-6","archive":["Portico"],"relation":{"has-preprint":[{"id-type":"doi","id":"10.1101\/634956","asserted-by":"object"}]},"ISSN":["2095-4689","2095-4697"],"issn-type":[{"value":"2095-4689","type":"print"},{"value":"2095-4697","type":"electronic"}],"subject":[],"published":{"date-parts":[[2020,3]]},"assertion":[{"value":"23 June 2019","order":1,"name":"received","label":"Received","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"24 September 2019","order":2,"name":"revised","label":"Revised","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"16 October 2019","order":3,"name":"accepted","label":"Accepted","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"6 March 2020","order":4,"name":"first_online","label":"First Online","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"The authors Bernard Mathey-Prevot, Bao-Tran Parker, Carolyn Im, Cierra Hong, Peng Dong, Guang Yao and Lingchong You declare that they have no conflict of interests.","order":1,"name":"Ethics","group":{"name":"EthicsHeading","label":"Compliance with Ethics Guidelines"}},{"value":"All procedures performed in studies were in accordance with the ethical standards of the institutional and\/or national research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards.","order":2,"name":"Ethics","group":{"name":"EthicsHeading","label":"Compliance with Ethics Guidelines"}},{"value":"This content has been made available to all.","name":"free","label":"Free to read"}]}}