{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,1,15]],"date-time":"2026-01-15T20:51:28Z","timestamp":1768510288227,"version":"3.49.0"},"posted":{"date-parts":[[2024,3,19]]},"group-title":"Biophysics","reference-count":57,"publisher":"openRxiv","content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2024,3,19]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                <jats:p>\n                  Many transcription factors (TFs) have been shown to bind RNA, leading to open questions regarding the mechanism(s) of this RNA binding and its role in regulating TF activities. Here we use biophysical assays to interrogate the k\n                  <jats:sub>on<\/jats:sub>\n                  , k\n                  <jats:sub>off<\/jats:sub>\n                  , and K\n                  <jats:sub>d<\/jats:sub>\n                  for DNA and RNA binding of two model human transcription factors, ER\u03b1 and Sox2. Unexpectedly, we found that both proteins exhibited multiphasic nucleic acid binding kinetics. We propose that Sox2 RNA and DNA multiphasic binding kinetics could be explained by a conventional model for sequential Sox2 monomer association and dissociation. In contrast, ER\u03b1 nucleic acid binding exhibited biphasic dissociation paired with novel triphasic association behavior, where two apparent binding transitions are separated by a 10-20 min \u201clag\u201d phase depending on protein concentration. We considered several conventional models for the observed kinetic behavior, none of which adequately explained all the ER\u03b1 nucleic acid binding data. Instead, simulations with a model incorporating sequential ER\u03b1 monomer association, ER\u03b1 nucleic acid complex isomerization, and product \u201cfeedback\u201d on isomerization rate recapitulated the general kinetic trends for both ER\u03b1 DNA and RNA binding. Collectively, our findings reveal that Sox2 and ER\u03b1 bind RNA and DNA with previously unappreciated multiphasic binding kinetics, and that their reaction mechanisms differ with ER\u03b1 binding nucleic acids via a novel reaction mechanism.\n                <\/jats:p>","DOI":"10.1101\/2024.03.18.585577","type":"posted-content","created":{"date-parts":[[2024,3,19]],"date-time":"2024-03-19T14:45:17Z","timestamp":1710859517000},"source":"Crossref","is-referenced-by-count":0,"title":["Transcription factors ER\u03b1 and Sox2 have differing multiphasic DNA and RNA binding mechanisms"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-8136-1574","authenticated-orcid":false,"given":"Wayne O.","family":"Hemphill","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5212-1945","authenticated-orcid":false,"given":"Halley R.","family":"Steiner","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Jackson R.","family":"Kominsky","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Deborah S.","family":"Wuttke","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Thomas R.","family":"Cech","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"54368","reference":[{"key":"2024032112401022000_2024.03.18.585577v1.1","doi-asserted-by":"publisher","DOI":"10.1186\/s13058-014-0494-7"},{"key":"2024032112401022000_2024.03.18.585577v1.2","doi-asserted-by":"publisher","DOI":"10.1101\/gad.224503"},{"key":"2024032112401022000_2024.03.18.585577v1.3","doi-asserted-by":"publisher","DOI":"10.1210\/me.2004-0486"},{"key":"2024032112401022000_2024.03.18.585577v1.4","doi-asserted-by":"publisher","DOI":"10.1111\/cge.12275"},{"key":"2024032112401022000_2024.03.18.585577v1.5","doi-asserted-by":"publisher","DOI":"10.1128\/MCB.25.14.6031-6046.2005"},{"key":"2024032112401022000_2024.03.18.585577v1.6","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0143160"},{"key":"2024032112401022000_2024.03.18.585577v1.7","doi-asserted-by":"crossref","first-page":"5306","DOI":"10.1021\/acs.biochem.5b00693","article-title":"Glucocorticoid Receptor\u2013DNA Dissociation Kinetics Measured in Vitro Reveal Exchange on the Second Time Scale","volume":"54","year":"2015","journal-title":"Biochemistry"},{"key":"2024032112401022000_2024.03.18.585577v1.8","doi-asserted-by":"publisher","DOI":"10.1172\/JCI27987"},{"key":"2024032112401022000_2024.03.18.585577v1.9","doi-asserted-by":"publisher","DOI":"10.1038\/s41586-020-2195-y"},{"key":"2024032112401022000_2024.03.18.585577v1.10","doi-asserted-by":"crossref","first-page":"462","DOI":"10.1038\/ng1120","article-title":"Mutations in SOX2 cause anophthalmia","volume":"33","year":"2003","journal-title":"Nat Genet"},{"key":"2024032112401022000_2024.03.18.585577v1.11","doi-asserted-by":"publisher","DOI":"10.1038\/nn.2397"},{"key":"2024032112401022000_2024.03.18.585577v1.12","unstructured":"Fersht A . 1985. 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